A recurring feature of innate immune receptor signaling is the self-assembly of signaling proteins into oligomeric complexes. The Myddosome is an oligomeric complex that is required to transmit inflammatory signals from TLR/IL1Rs and consists of MyD88 and IRAK family kinases. However, the molecular basis for how Myddosome proteins self-assemble and regulate intracellular signaling remains poorly understood. Here, we developed a novel assay to analyze the spatiotemporal dynamics of IL1R and Myddosome signaling in live cells. We found that MyD88 oligomerization is inducible and initially reversible. Moreover, the formation of larger, stable oligomers consisting of more than four MyD88s triggers the sequential recruitment of IRAK4 and IRAK1. Notably, genetic knockout of IRAK4 enhanced MyD88 oligomerization, indicating that IRAK4 controls MyD88 oligomer size and growth. MyD88 oligomer size thus functions as a physical threshold to trigger downstream signaling. These results provide a mechanistic basis for how protein oligomerization might function in cell signaling pathways.

Purpose

Pseudoexfoliation (PXF) is a unique form of glaucoma characterized by accumulation of exfoliative material in the eyes. Changes in tear profile in disease stages may give us insights into molecular mechanisms involved in causing glaucoma in the eye.

High-dimensional Imaging technologies allow to simultaneously measure the expression levels of dozens of proteins in biological tissue, providing insight into single-cell phenotypes and organization in situ. Imaging artifacts, resulting from the sample, assay or instrumentation complicate downstream analyses and require correction by domain experts. Here, we present MAUI, a series of graphical user interfaces that facilitate this data pre-processing, including the removal of channel crosstalk, noise and antibody aggregates. MAUI accelerates and automates these steps, such that reproducible, high-quality data will be the input for subsequent stages of analysis.

Bacterial cell division requires the synthesis of septal peptidoglycan by the widely conserved SEDS-bPBP protein complex FtsWI, but how the complex is activated during cell division is still poorly understood. Previous studies suggested that FtsN initiates a signaling cascade in the periplasm to activate FtsWI. Here we isolated and characterized activated FtsW and FtsI mutants and confirmed that the signaling cascade for FtsW activation goes from FtsN to FtsQLB to FtsI and then to FtsW. The residues corresponding to mutations affecting FtsWI activation are clustered to a small region of the interaction interface between the pedestal domain of FtsI and the extracellular loop 4 of FtsW, suggesting that this interaction mediates activation of FtsW. This is strikingly similar to the proposed activation mechanism for the RodA-PBP2 complex, another SEDS-bPBP complex required for cell elongation. Thus, the two homologous SEDS-bPBP complexes are activated similarly by completely unrelated activators that modulate the interaction interface between the SEDS proteins and the bPBPs.

Farnesyl pyrophosphate, a metabolic intermediate of the cholesterol synthesis pathway, functions as a novel danger signal to trigger acute cell necrosis, and collaborates with activation of the cation channel TRPM2 to play an important role in brain ischemic injury.

Roughly 290 million electrical action potentials occur every second in the human brain, facilitating the propagation of signals among cells in the nervous system and driving most of our daily operations. New methods in brain imaging are emerging that have the speed and resolution to capture events in the brain at the pace at which neurons typically communicate. These methods measure voltage in neurons by using light, and therefore can access very detailed brain signaling patterns. However, the adoption of these methods by a larger community, and not a restricted set of experts, is limited by the lack of computational tools, thereby greatly hindering progress in this field. In this paper, we present VolPy, a software framework that greatly facilitates the preprocessing of this new type of imaging datasets. This pipeline incorporates efficient and optimized algorithms that can identify neurons and extract their activity with great accuracy. The presented software will make this new imaging modality accessible to a wide audience.

Elucidating the complex relationship between nutrient-induced signaling and metabolism represents a key in understanding the onset of many different human diseases like obesity, type 3 diabetes, cancer, and many neurological disorders. In this work we proposed a hybrid modeling approach, combining Boolean representation of signaling pathways, like Snf1, TORC1, and PKA with the enzyme constrained model of metabolism linking them via the regulatory network. This allowed us to improve individual model predictions and elucidate how single components in the dynamic signaling layer affect steady-state metabolism. The model has been tested under respiration and fermentation, revealing novel connections and further reproducing the regulatory effects that are associated with the Crabtree effect and glucose repression. Finally, we show a connection between Snf1 signaling and chronological lifespan.

Ubiquitin ligases modify proteins post-translationally which is essential for a variety of cellular processes. UBR5 is an E3 ubiquitin ligase and in Drosophila is a regulator of Hedgehog signaling. In mammals, the Hedgehog (HH) signalling pathway, among many other roles, plays an essential role in tissue maintenance, a process called homeostasis. A murine genetic system was developed to specifically eliminate UBR5 function from embryonic limb tissue that subsequently forms bone and connective tissue (ligaments and tendons). This approach revealed that UBR5 operates as a potent suppressor of excessive growth of normal cartilage and bone and prevents formation of bone in ectopic sites in connective tissue near the knees and ankle joints. In contrast to abnormal growth, UBR5 inhibits degradation of the articular cartilage that cushions the knee joint leading to extensive exposure of underlying bone. Furthermore, Ubr5 interacts with smoothened, a component of the HH pathway, identifying UBR5 as a regulator of mammalian HH signaling in the postnatal musculoskeletal system. In summary, this work shows that UBR5 interacts with the HH pathway to regulate skeletal homeostasis in and around joints of the legs and identifies targets that may be harnessed for biomedical engineering and clinical applications.

Proteins of the ezrin, radixin, and moesin (ERM) family control cell and tissue morphogenesis. We previously reported that moesin, the only ERM in Drosophila, controls mitotic morphogenesis and epithelial integrity. We also found that the Pp1-87B phosphatase dephosphorylates moesin, counteracting its activation by the Ste20-like kinase Slik. To understand how this signaling pathway is itself regulated, we conducted a genome-wide RNAi screen, looking for new regulators of moesin activity. We identified that Slik is a new member of the striatin-interacting phosphatase and kinase complex (STRIPAK). We discovered that the phosphatase activity of STRIPAK reduces Slik phosphorylation to promote its cortical association and proper activation of moesin. Consistent with this finding, inhibition of STRIPAK phosphatase activity causes cell morphology defects in mitosis and impairs epithelial tissue integrity. Our results implicate the Slik–STRIPAK complex in the control of multiple morphogenetic processes.

Biological timekeeping enables the coordination and execution of complex cellular processes such as developmental programs, day/night organismal changes, intercellular signaling, and proliferative safeguards. While these systems are often considered separately owing to a wide variety of mechanisms, time frames, and outputs, all clocks are built by calibrating or delaying the rate of biochemical reactions and processes. In this review, we explore the common themes and core design principles of cellular clocks, giving special consideration to the challenges associated with building timers from biochemical components. We also outline how evolution has coopted time to increase the reliability of a diverse range of biological systems.

How rapid cell multiplication leads to cell differentiation in developing tissues is still enigmatic. This question is central to morphogenesis, cell number control, and homeostasis. Self-renewal epidermoid epithelia are continuously exposed to mutagens and are the most common target of cancer. Unknown mechanisms commit rapidly proliferating cells to post-mitotic terminal differentiation. We have over-activated or inhibited the endogenous DNA damage response (DDR) pathways by combinations of activating TopBP1 protein, specific shRNAs, or chemical inhibitors for ATR, ATM, and/or DNA-PK. The results dissect and demonstrate that these signals control keratinocyte differentiation in proliferating cells independently of actual DNA damage. The DDR limits keratinocyte multiplication upon hyperproliferative stimuli. Moreover, knocking down H2AX, a common target of the DDR pathways, inhibits the epidermoid phenotype. The results altogether show that the DDR is required to maintain the balance proliferation differentiation and suggest that is part of the squamous program. We propose a homeostatic model where genetic damage is automatically and continuously cleansed by cell-autonomous mechanisms.

In neurons, dendrites form the major sites of information receipt and integration. It is thus vital that, during development, the dendritic arbor is adequately formed to enable proper neural circuit formation and function. While several known processes shape the arbor, little is known of those that govern dendrite branching versus extension. Here, we report a new mechanism instructing dendrites to branch versus extend. In it, glutamate signaling activates mGluR5 receptors to promote Ckd5-mediated phosphorylation of the C-terminal PDZ-binding motif of delta-catenin. The phosphorylation state of this motif determines delta-catenin’s ability to bind either Pdlim5 or Magi1. Whereas the delta:Pdlim5 complex enhances dendrite branching at the expense of elongation, the delta:Magi1 complex instead promotes lengthening. Our data suggest that these complexes affect dendrite development by differentially regulating the small-GTPase RhoA and actin-associated protein Cortactin. We thus reveal a “phospho-switch” within delta-catenin, subject to a glutamate-mediated signaling pathway, that assists in balancing the branching versus extension of dendrites during neural development.

Secondary cell walls are critical for plant growth and of economic importance to humans as fiber, pulp for paper, and as a renewable source of energy. A master switch for secondary wall biosynthesis, MYB46, ensures the exquisite expression of the biosynthetic genes in the tissues where secondary walls are critical. However, suppression of secondary wall formation is needed during impermanent cessation of vegetative growth triggered by environmental stresses. Although the expression of MYB46 is upregulated by abiotic stresses, secondary wall formation may not occur. How is this opposing control achieved? The mitogen-activated protein kinase (MPK) cascade is among the most conserved signal transduction systems in eukaryotes and plays a crucial role in the regulation of biochemical and physiological changes associated with environmental stimuli and phytohormones. In this report, we show that MYB46 is negatively regulated by MPK6 during salt stress, providing a novel insight into a mechanism by which plants incorporate environmental signals into differential regulation of secondary wall biosynthesis.

A central problem in animal regeneration is how animals determine what body part to regenerate. Planarians are flatworms that can regenerate any missing body region, and are studied to identify mechanisms underlying regeneration. At transverse amputation planes, a poorly understood mechanism specifies regeneration of either a head or a tail. This head-versus-tail regeneration decision-making process is referred to as regeneration polarity and has been studied for over a century to identify mechanisms that specify what to regenerate. The gene notum, which encodes a Wnt antagonist, is induced within hours after injury preferentially at anterior-facing wounds, where it specifies head regeneration. We report that Activin signaling is required for regeneration polarity, and the underlying asymmetric activation of notum at anterior- over posterior-facing wounds. We propose that Activin signaling is involved in regeneration-specific responses broadly in the animal kingdom.

In many reptiles and fish, environment can determine, or influence, the sex of developing embryos. How this happens at a molecular level that has eluded resolution for half a century of intensive research. We studied the bearded dragon, a lizard that has sex chromosomes (ZZ male and ZW female), but in which that temperature can override ZZ sex chromosomes to cause male to female sex reversal. This provides an unparalleled opportunity to disentangle, in the same species, the biochemical pathways required to make a female by these two different routes. We sequenced the transcriptomes of gonads from developing ZZ reversed and normal ZW dragon embryos and discovered that different sets of genes are active in ovary development driven by genotype or temperature. Females whose sex was initiated by temperature showed a transcriptional profile consistent with the recently-proposed Calcium-Redox hypotheses of cellular temperature sensing. These findings are an important for understanding how the environment influences the development of sex, and more generally how the environment can epigenetically modify the action of genes.

Wingless (Wg)/Wnt proteins act as important signals between cells in many contexts. For decades, studies in the Drosophila wing established that Wg signals to distant cells, implying that Wg spreads extracellularly. However, studies in other tissues and organisms have found Wnt ligands signal in a juxtacrine manner, to neighboring cells. Recently the importance of Wg spreading was re-evaluated in the fly wing, spurred by the finding that membrane-tethered Wg, unable to spread from its source cell, can substitute for Wg. These findings fueled a search for other tissues where Wg extracellular spreading is required. The nature of Wg signaling in Drosophila oogenesis has been unclear. In the germarium a visible gradient of Wg spans ~50 μm, reaching from its source to follicle stem cells, but it has been argued that Wg signals from neighboring cells to the stem cells. In this study, we tested the role of Wg spreading by analyzing oogenesis in the tethered-Wg flies. Two copies of tethered Wg cause a non-specific Wg toxicity; however, even when the dose tethered Wg is reduced, ovaries have negligible follicle stem cell proliferation and produce few eggs. Thus, extracellular Wg spreading is essential for follicle stem cell proliferation and oogenesis.

G. duodenalis, one of the most common cause of diarrheal diseases, is widely existed in the contaminated water and threatening the public health especially in developing countries. Along with the increasing resistance to anti-G. duodenalis drugs occurs, new targets against giardiasis are of urgently needed. The innate immune system is the first defense line of organism to resist multiple pathogens invasion through recognizing pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs), termed Toll-like receptors (TLRs) on the surface of cell membrane and nucleotide oligomerization domain (Nod)-like receptors (NLRs) inside immune cells. Recently, extracellular vesicles have emerged as a ubiquitous mechanism participating in cells communications. In this study, EVs secreted by extracellular protozoan G. duodenalis were obtained and displayed typical cup-shaped structure with 150 nm in diameter. Moreover, GEVs could enter into primary mouse peritoneal macrophages and regulate host cell innate immunity by up-regulation of various inflammatory cytokines expression. Furthermore, TLR2 and NLRP3 inflammasome signaling pathways involved in this process. This study demonstrated that GEVs could be internalized into primary mouse peritoneal macrophages, regulate host cell innate immunity via TLR2 and NLRP3 inflammasome signaling pathways, and may provide new targets against giardiasis.

Toll/interleukin-1 receptor domain-containing protein encoded by E. coli (TcpC) is an important virulence factor in many strains of uropathogenic E. coli (UPEC). TcpC-mediated evasion of innate immunity plays an important role in the pathogenesis of UPEC caused urinary tract infection (UTI) including pyelonephritis. In the present study, we show TcpC is an E3 ubiquitin ligase that promotes ubiquitination and degradation of MyD88, hereby blocking the TLR signaling pathway. Our findings not only illuminate the novel biochemical mechanisms underlying TcpC-mediated evasion of innate immunity, but also provide the first example that bacterial pathogens can subvert TLR signaling pathway through virulence factors that function as MyD88-targeted E3 ubiquitin ligase.

A specific strain of Vibrio parahaemolyticus causing acute hepatopancreatic necrosis disease (AHPND) in shrimp or VP_AHPND produces a binary toxin (PirAB^vp toxin) that is previously known to induce cell death of stomach and hepatopancreas but the molecular mechanism has not been defined. Similar to Cry toxin receptor in insects, a novel aminopeptidase N1 protein from L. vannamei (LvAPN1) was identified as a putative receptor of VP_AHPND toxin. Suppression of LvAPN1 reduced the number of AHPND virulence plasmids in stomach and occurrence of AHPND clinical sign, sustained the number of total hemocyte count, and elevated the number of viable hemocyte. We demonstrated that VP_AHPND toxin challenge induces hemocyte cell damage and it interacts with LvAPN1 in vitro. Collectively, our finding suggested that not only stomach and hepatopancreas but also hemocyte are the VP_AHPND target tissues where LvAPN1 serves as a VP_AHPND toxin receptor. This study provides novel insight into the contributions of LvAPN1 receptor towards the AHPND pathogenesis in shrimp and may extend to the development of AHPND preventive measure in shrimp.

The tumor microenvironment (TME) is a very complex cellular ecosystem, composed of the cancer cells (carrying the disease-causing genetic alterations), immune cells and other stromal cells (such as fibroblasts), which contribute to disease progression and drug responses. Here, we investigated these complex cellular dynamics by reconstituting the tumor ecosystems in a very controlled manner within microfluidic devices, with multiple cell populations, generating the so-called ‘tumors-on-chip’, which can be visualized by video-microscopy and treated with anti-cancer drugs. The resulting videos contain a huge amount of information that requires advanced computational approaches to be extracted. In this work, we developed a novel method, named STAMP, that precisely measures the kinetics and the spatial maps of cancer cell deaths within tumor-on-chip. Two case studies are presented: breast cancer cells upon chemotherapy treatment (doxorubicin) and lung cancer cells upon killing by specific immune cells (tumor-infiltrating cytotoxic T lymphocytes). We generated spatio-temporal maps on cancer death uncovering unsuspected relations between death events. This indicates that dying cancer cells might release soluble factors that induce death of neighbor cancer cells. The STAMP method was suitable to study the capacity of fibroblasts to promote resistance of cancer cells to chemotherapy.

Adult organs often contain a stem cell niche that maintains stem cells necessary for the replenishment of different types of terminally differentiated cells that are continuously lost. This study reveals that various modes of Notch signaling activation induce the formation of the germline stem cell niche in Drosophila. We show for the first time that even among non-equipotent cells, Notch signaling can be trans-activated via distant induction mode, where the ligand Delta is delivered via cellular protrusions to the somatic stem cell niche precursors located several cell diameters away. Moreover, there is a second security mechanism controlled by the soma that additionally ensures that the stem cell niche is formed. In the stem cell niche precursors, Notch signaling can be locally inhibited by the somatic Delta. While Notch signaling trans-inhibition has been proposed via mathematical modelling, our findings show that a group of cells that have high Delta can be seen in a living organism, confirming that this mode of Notch signaling inhibition by trans-Delta exists in vivo. This work provides significant advances in the understanding of Notch signaling and the stem cell niche formation, which is important for the fields of stem cell biology and regenerative medicine.

NOTCH1 deregulation can favor hematological malignancies. In addition to RBP-Jκ/NICD/MAML1, other regulators are required for the measured activation of NOTCH target genes. IKAROS is a known repressor of many NOTCH targets. Since it can also favor transcriptional activation and control gene expression levels, we questioned whether IKAROS could participate to the activation of specific NOTCH target genes. We are reporting that upon NOTCH induction, the absence of IKAROS impairs the measured activation of two groups of NOTCH target genes: (i) those overexpressed and characterized by an additive effect imposed by the absence of IKAROS and NOTCH induction; and (ii) those ‘desensitized’ and no more activated by NOTCH. At genes of both groups, IKAROS controls the timely recruitment of the chromatin remodelers CHD4 and BRG1. IKAROS then influences the activation of these genes through the organization of chromatin and poised transcription or through transcriptional elongation control. The importance of the IKAROS controlled and measured activation of genes is not limited to NOTCH signaling as it also characterizes Cdkn1a expression upon genotoxic stress. Thus, these results provide a new perspective on the importance of IKAROS for the adequate cellular response to stress, whether imposed by NOTCH or genotoxic insults.

Human cytomegalovirus is a major cause of congenital birth defects. Yet, currently the best way to avoid cytomegalovirus disease is to prevent infection of pregnant women through hygiene measures. Once the mother is infected there is no approved treatment to block transmission to the fetus. One intensively researched option is to neutralize the virus produced by the infected mother with anti-HCMV antibodies. Yet, as the efficiency of this approach remains to be demonstrated, alternative approaches need to be considered. Similar to antibodies, PDGFRα-Fc binds to the virus and blocks infection, but it is more potent and has a broader activity of inhibition which makes it a promising alternative. A problem however is that PDGFRα-Fc can not only bind to the virus but also to PDGFs which are important growth factors involved in cell-cycle regulation and tissue development. The results of this study offer a solution. Combinations of mutations were identified that can be introduced in PDGFRα-Fc to abrogate sequestration of PDGFs. Thus, the potential side effects of PDGFRα-Fc can be circumvented while it remains active against HCMV. These results pave the way for development of PDGFRα-Fc as a promising HCMV inhibitor.

The coordinated replication of complex eukaryotic genomes is primarily regulated at the level of replication initiation. Therefore, how initiation is regulated in time and space is an active field of research. A leading hypothesis is that the stoichiometry of MCM, the replicative helicase, at replication origins is a significant determinant of initiation timing. However, how MCM stoichiometry at origins is regulated is unknown. We tested two hypothetical mechanisms for the control of MCM stoichiometry: Origin capacity, in which MCM stoichiometry is constrained by the varying capacity of individual origins to load MCM, and ORC activity, in which MCM stoichiometry is regulated by the varying activity of ORC, the MCM loader, at individual origins. We find that neither mechanism is consistent with our data, but that a hybrid of the two mechanisms is.

Anautogenous mosquitoes must consume vertebrate blood meals to complete oogenesis. Repeated blood feeding makes the mosquitoes efficient disease-transmitting vectors. The TOR pathway activated by ingested blood is known as an important regulator for vitellogenesis in mosquitoes. Herein, we show that the protein kinase TOR is involved in the regulation of mosquitoes’ susceptibility to Plasmodium infection. Inhibition of the TOR pathway by rapamycin upregulates the expression of REL2, a transcription factor controlling the expression of a variety of immune effectors. The enhanced immune responses in turn promote parasite elimination. Therefore, the TOR pathway plays a dual role in not only regulating mosquito reproduction but also in their vector potential.

TGF-β signaling is critical for promoting cancer metastasis, primarily via Smad-dependent regulation of the epithelial-mesenchymal transition; this study reveals that non-canonical TGF-β signaling stabilizes the E3 ubiquitin ligase FBXO3 to target ΔNp63α for degradation, resulting in downregulation of adhesion molecules and promotion of breast cancer metastasis.

Biomineralization is a widespread, fundamental process by which organisms use minerals to harden their tissues. Mineral-bearing vesicles were observed in biomineralizing cells and play an essential role in biomineralization, yet little is known about their three-dimensional (3D) dynamics. Here we quantify 3D-vesicle-dynamics during calcite skeleton formation in sea urchin larvae, using lattice-light-sheet microscopy. We discover that calcium vesicles perform a diffusive motion in both calcifying and non-calcifying cells of the embryo. The diffusion coefficient and vesicle speed are higher in the mesenchymal skeletogenic cells compared to the epithelial ectodermal cells. This difference is possibly due to the higher rigidity of the ectodermal cells as demonstrated by the enhanced signal of f-actin and myosinII activity in these cells compared to the skeletogenic cells. The motion of the vesicles in the skeletogenic cells, is not directed toward the biomineralization compartment but the vesicles slow down near it, possibly to deposit their content. Blocking skeletogenesis through the inhibition of Vascular Endothelial Growth Factor Receptor (VEGFR), increases vesicle volume but doesn’t change the diffusion mode and the cytoskeleton markers in the cells. Our studies reveal the active diffusive motion of mineral bearing vesicles that is apparently defined by the mechanical properties of the cells.

The sonic hedgehog (Shh) pathway is vital for normal development in a wide variety of species and its activity is strictly regulated to ensure correct spatiotemporal patterning of numerous developing tissues. Shh signalling requires the formation of Shh aggregates, formed on producing cells via a range of different mechanisms, that then diffuse to receiving cells. We formulate and analyse a mathematical model of the most well described mechanisms, namely monomer multimerisation, and recruitment of Shh by heparan sulfate proteoglycans and lipoproteins. Our results illustrate a distribution of the size and quantities of aggregates formed by these mechanisms. We found that as a consequence of competition between the mechanisms for Shh monomers the shape distribution of Shh aggregates resembles an exponential distribution. We also found the distribution to be robust to both parameter changes and variations to the processes by which mechanisms recruit Shh. We report that our approach and subsequent results demonstrate that these mechanisms act in synergy allowing Shh to aggregate in various quantities with diverse diffusive abilities. We postulate that this regulation contributes significantly to aid precision in signalling for Shh in areas of development.

Background/aims

Isoalantolactone (IATL) is one of multiple isomeric sesquiterpene lactones and is isolated from inula helenium. IATL has multiple functions such as antibacterial, antihelminthic and antiproliferative activities. IATL also inhibits pancreatic cancer proliferation and induces apoptosis by increasing ROS production. However, the detailed mechanism of IATL-mediated pancreatic cancer apoptosis remains largely unknown.

Background

Pancreatic adenocarcinoma (PAAD) is a pancreatic disease with a high mortality rate in the world. This present research intends to identify the function of lncRNA LINC00857/miR-340-5p/Transforming growth factor alpha (TGFA) in the progression of PAAD.

Cancer is one of leading causes of death in the world, but unfortunately, the mechanism of tumor metastasis remains unclear so far. Intuitively, the tumor metastasis starts from a tumor cell migrating towards the vessel wall (namely margination), then adhering to the vessel wall (namely adhesion), and finally extravasating from where it adheres onto. Hence, it is important to investigate the margination and adhesion of tumor cells for understanding the tumor metastasis. We implemented a three-dimensional simulation to directly reproduce these two processes at a cellular scale, where the dynamic behaviors, such as deformation, aggregation and adhesion, of a lot of cells in a very complex microvascular network are taken into account. The results suggest that the tumor cells may be prone to adhere at the microvascular bifurcations with low shear rate and moderate hematocrit, because of a high wall-directed force from the surrounding RBCs. This implies that the tumor may be more likely to extravasate at the microvascular bifurcations if the blood flow is slow and the hematocrit is moderate. Our work may provide new insights into the cancer pathophysiology and its diagnosis and therapy.

Each human red blood cell traverses narrow capillaries about 1000 to 2000 times per day. Cell deformability is essential for smooth transits and this requires tight control of cell volume well below spherical maxima. The pump-leak ion-flux balance controlling red cell volume involves sodium-potassium and calcium pumps and a constellation of passive membrane transporters. PIEZO1, a mechanosensitive ion channel plays a central role, activating transiently during capillary transits. The main question, not accessible to direct experimentation, is what happens to red cell volume and composition in vivo during each capillary transit. We attempted answering this question by applying novel extensions to a tried and tested model of red cell homeostasis that encodes the current knowledge on the subject. The results revealed an unexpected up-down biphasic volume response characterized by a sharp initial surge caused by CaCl₂ influx via PIEZO1, associated with osmotic fluid gain, followed by a slow reversal, a prediction in conflict with prevailing views. The infinitesimal amplitude of the predicted volume changes precludes them playing a significant role in flow dynamics. In the following paper we explore how the biphasic response helps explain the age-related changes red cells experience during their long circulatory lifespan.

Courtship behavior in male Drosophila melanogaster is controlled by a well-defined neural circuit that is labeled by the male-specific transcription factor Fruitless (Fru^M). While Fru^M is known to change the number, anatomy and connectivity of neurons which comprise the circuit and has been suggested to repress the expression of a few gene targets, the mechanism of how Fru^M regulates genes across many different kinds of neurons is unknown. Using an approach to identify gene regulatory elements based on their chromatin accessibility states (ATAC-seq), we identified a large set of chromatin accessibility changes downstream of Fruitless. By examining the activity of these regulatory elements in vivo, we found that their activity was 1) sexually dimorphic and 2) specific to a single class of Fru^M neurons, suggesting that Fru^M acts on different chromatin targets in different neuron classes comprising the courtship circuit. Further, we found a known Fru^M-regulated enhancer of the Fru^M-repressed gene Lgr3 to have closed chromatin specifically in Fru^M neurons. Combined with an enrichment of Fru^M motifs in regions which are closed in Fru^M neurons, we present a mechanism where Fru^M directs the decommissioning of sex-shared regulatory elements to masculinize neurons in a cell-type specific manner.

The mammalian target of rapamycin complex 1 (mTORC1) integrates mitogenic and stress signals to control growth and metabolism. Activation of mTORC1 by amino acids and growth factors involves recruitment of the complex to the lysosomal membrane and is further supported by lysosome distribution to the cell periphery. Here, we show that translocation of lysosomes toward the cell periphery brings mTORC1 into proximity with focal adhesions (FAs). We demonstrate that FAs constitute discrete plasma membrane hubs mediating growth factor signaling and amino acid input into the cell. FAs, as well as the translocation of lysosome-bound mTORC1 to their vicinity, contribute to both peripheral and intracellular mTORC1 activity. Conversely, lysosomal distribution to the cell periphery is dispensable for the activation of mTORC1 constitutively targeted to FAs. This study advances our understanding of spatial mTORC1 regulation by demonstrating that the localization of mTORC1 to FAs is both necessary and sufficient for its activation by growth-promoting stimuli.

In the mammalian dentition, the number of the tooth roots varies in a manner that is finely tailored to the physiological function of each tooth type. Specifically, the anterior incisors and canines, which are mainly responsible for lower occlusal pressure movements such as cutting or shearing, are single-rooted, while the more posterior premolars and molars are multi-rooted and used for motions that require higher bite force such as chewing, crushing and grinding. So far, we have very limited understanding of the molecular mechanism that precisely determines the number of tooth roots. The multi-rooted mouse molar presents an excellent model for us to study this question. Here, we show that the gene Lhx6 is a key determinant of mouse molar root number, as the absence of this gene results in the transformation of multi-rooted mouse molars into single-rooted ones. We further show how this gene achieves its functional specificity in determining the tooth root number by controlling the region-specific fate determination of progenitor cells. This finding not only provides clues that could inform future tooth regeneration strategies, but also serves to illustrate developmental processes shared with other mammalian organs.

Excessive accumulation of collagen leads to fibrosis. Integrin α1β1 (Itgα1β1) prevents kidney fibrosis by reducing collagen production through inhibition of the EGF receptor (EGFR) that phosphorylates cytoplasmic and nuclear proteins. To elucidate how the Itgα1β1/EGFR axis controls collagen synthesis, we analyzed the levels of nuclear tyrosine phosphorylated proteins in WT and Itgα1-null kidney cells. We show that the phosphorylation of the RNA-DNA binding protein fused in sarcoma (FUS) is higher in Itgα1-null cells. FUS contains EGFR-targeted phosphorylation sites and, in Itgα1-null cells, activated EGFR promotes FUS phosphorylation and nuclear translocation. Nuclear FUS binds to the collagen IV promoter, commencing gene transcription that is reduced by inhibiting EGFR, down-regulating FUS, or expressing FUS mutated in the EGFR-targeted phosphorylation sites. Finally, a cell-penetrating peptide that inhibits FUS nuclear translocation reduces FUS nuclear content and collagen IV transcription. Thus, EGFR-mediated FUS phosphorylation regulates FUS nuclear translocation and transcription of a major profibrotic collagen gene. Targeting FUS nuclear translocation offers a new antifibrotic therapy.

The tumor suppressor PTEN is essential for early development. Its lipid phosphatase activity converts PIP₃ to PIP₂ and antagonizes the PI3K–Akt pathway. In this study, we demonstrate that PTEN’s protein phosphatase activity is required for epiblast epithelial differentiation and polarization. This is accomplished by reconstitution of PTEN-null embryoid bodies with PTEN mutants that lack only PTEN’s lipid phosphatase activity or both PTEN’s lipid and protein phosphatase activities. Phosphotyrosine antibody immunoprecipitation and mass spectrometry were used to identify Abi1, a core component of the WASP-family verprolin homologous protein (WAVE) regulatory complex (WRC), as a new PTEN substrate. We demonstrate that PTEN dephosphorylation of Abi1 at Y213 and S216 results in Abi1 degradation through the calpain pathway. This leads to down-regulation of the WRC and reorganization of the actin cytoskeleton. The latter is critical to the transformation of nonpolar pluripotent stem cells into the polarized epiblast epithelium. Our findings establish a link between PTEN and WAVE-Arp2/3–regulated actin cytoskeletal dynamics in epithelial morphogenesis.

Athie et al. (2020. J. Cell Biol. https://doi.org/10.1083/jcb.201908078) identify ALAL-1, a lncRNA frequently amplified or overexpressed in lung cancer, as an oncogenic driver, capable of promoting the proliferation and altering the immunogenicity of lung cancer cells.

Natural killer (NK) cells have potent antitumor and antimetastatic activity. It is incompletely understood how cancer cells escape NK cell surveillance. Using ex vivo and in vivo models of metastasis, we establish that keratin-14⁺ breast cancer cells are vulnerable to NK cells. We then discovered that exposure to cancer cells causes NK cells to lose their cytotoxic ability and promote metastatic outgrowth. Gene expression comparisons revealed that healthy NK cells have an active NK cell molecular phenotype, whereas tumor-exposed (teNK) cells resemble resting NK cells. Receptor–ligand analysis between teNK cells and tumor cells revealed multiple potential targets. We next showed that treatment with antibodies targeting TIGIT, antibodies targeting KLRG1, or small-molecule inhibitors of DNA methyltransferases (DMNT) each reduced colony formation. Combinations of DNMT inhibitors with anti-TIGIT or anti-KLRG1 antibodies further reduced metastatic potential. We propose that NK-directed therapies targeting these pathways would be effective in the adjuvant setting to prevent metastatic recurrence.

When new blood vessels are created, the endothelial cells that make up these vessels migrate and rearrange in response to blood flow to remodel and optimise the vessel network. An essential part of this process is maintaining the branched structure of the network; however, it is unclear what cues cells consider at regions where vessels branch (i.e., bifurcations). In this research, we present a computer model of cell migration to interrogate the process of preserving bifurcations during remodelling. In this model, cells at bifurcations are influenced by both flow and force transmitted from neighbouring cells. We found that both cues (flow-based and collective-based) must be considered equally in order to preserve branching in the vessel network. In simulations with stable bifurcations, we demonstrated that these cues oscillate: a strong signal in one was accompanied by a weak signal in the other. Furthermore, we found that these cues naturally compete with each other due to the coupling between blood flow and the size of the blood vessels, i.e. larger vessels with more cells produce less flow signals and vice versa. Our research provides insight into how forces transmitted between neighbouring cells stabilise and preserve branching during remodelling, as well as implicates the disruption of this force transmission as a potential mechanism when remodelling goes wrong as in the case of vascular malformation.

Ion channels are proteins that control the flow of ions into the cell. The family of ion channels known as the pentameric ligand gated ion channels (pLGICS) open in response to the binding of a neurotransmitter, moving the channel from a resting state to an open state. The glycine receptor is a pLGIC whose structure has been resolved in different functional states. It is also known that the response of pLGICs can also be modified by different types of lipids found within the membrane itself but exactly how is unclear. Here, we used a realistic model of a neuronal membrane and performed molecular dynamics simulations to show various lipid-protein interactions that are dependent on the channel state. Our work also reveals previously unconsidered protein-lipid interactions at a key junction of the channel known to be critical for the transmission of the opening process. We also demonstrate that cholesterol interacts with the protein at a site already known to bind to another compound that modulates the channel, called ivermectin. The work should be useful for future design of allosteric modulators.

Objective

The current study investigated the mechanism underlying the therapeutic effects of berberine in the vasculature in hypertension.

Currently, RNA massive sequencing RNA-seq is the most extensively used technique for gene expression profiling in a single assay. The output of RNA-seq experiments contains millions of sequences, generated from cDNA libraries produced by the retro-transcription of RNA samples, that need to be processed by computational methods to be transformed into meaningful biological information. Thus, a number of bioinformatic workflows and pipelines have been proposed to produce different types of gene expression measurements, including in some cases, functional annotations to facilitate biological interpretation. While most pipelines focus exclusively on transcriptional data, the ultimate activity of the resulting gene product also depends critically on its integrity. Although traditional hybridization-based transcriptomics methodologies (microarrays) miss this information, RNA-seq data also contains information on variants present in the transcripts that can affect the function of the gene product, which is systematically ignored by current RNA-seq pipelines. MIGNON is the first workflow able to perform an integrative analysis of transcriptomic and genomic data in the proper functional context, provided by a mechanistic model of signaling pathway activity, making thus the most of the information contained in RNA-Seq data. MIGNON is easy to use and to deploy and may become a valuable asset in fields such as personalized medicine.

Background

Sarcoidosis is a systemic inflammatory granulomatous disease, frequently affecting the lung. If left untreated, it may end in lung fibrosis. Proangiogenic and profibrotic vascular endothelial growth factor (VEGF), transforming growth factor (TGF)-β1, fibroblast growth factor (FGF)-2 and platelet-derived growth factor (PDGF)-AB are a known therapeutical target in pulmonary fibrosing diseases, e.g. IPF, but there is no targeted therapy option for pulmonary fibrosis in sarcoidosis.

Sensation, behavior, and cognition all depend on the proper function of neuronal connections called synapses. Synapses that use the neurotransmitter glutamate to signal between nerve cells are the most abundant type in our brain. Presynaptic neurons release glutamate, which activates glutamate receptors on postsynaptic neurons. Dysfunction of glutamate synapses leads to several neurological disorders, and changing their strength–in part by altering glutamate receptors numbers on the surface of the postsynaptic cell—provides the cellular basis of learning and memory. Much remains to be learned about how factors released from other cell types affects synaptic communication. We took advantage of light-activated molecular switches engineered into specific sensory neurons of C. elegans worms to trigger a behavioral reflex that depends on glutamate synapses. Using this behavior, we identified proteins called VER-1 and VER-4 as important for glutamate synapse function. We found that worms missing these VER proteins or their activator PVF-1 have reduced levels of glutamate receptors at the postsynaptic surface and defects in glutamate-dependent behaviors. Our results suggest that inter-tissue cross-talk between muscle PVF-1 and neuronal VERs is important for controlling the number of glutamate receptors at the cell surface, robust neuronal communication and behavioral responses.

In all organisms there are sets of “reserve” cells that have the capacity to self-renew and eventually differentiate to repair or provide new structures. Here we report that the Drosophila headcase gene provides such kinds of cells with a stress protective mechanism. Interestingly, headcase is also involved in providing such “reserve” cells with the signal required for initiating their differentiation into new structures. Thus, headcase establishes a fine equilibrium between differentiation and stress protection of these “reserve” cells. Of note headcase is the founding member of a group of homolog proteins identified in many animals. In humans, the headcase homolog is associated with different kinds of cancers but its function and role remains unknown. The finding that the Drosophila headcase confers stress protection opens up a new way to be explored regarding the role of the human Headcase as it could also act as a stress protector and its drop-off may induce stress conditions favouring the initiation and progression of tumorigenesis.

Cells use molecular signaling networks to translate dynamically changing stimuli into appropriate downstream responses. Specialized network structures, or motifs, allow cells to properly decode a variety of temporal input signals. In this paper, we present an algorithm that infers signaling motifs from multiple examples of an upstream signal paired with its downstream response in a population of single cells. We compare the predictive power of single-cell versus averaged time-series traces and the incremental benefit of adding more single-cell traces to the algorithm. We use this approach to understand how yeast respond to environmental stresses.

Fungal pathogens of crops and humans pose annual threats to our food and health. There are many steps to the host infection process, during which fungal pathogens display unique growth, and use specific genes to cause disease. Despite this knowledge, many aspects of how pathogens regulate their genome to enact this process remain unknown. Here, we demonstrate how chemical modification of lysine residues on the histone H3, which helps organize and control DNA usage, play an important regulatory role in the model fungal pathogen causing rice blast disease. Our analysis shows a significant association between genes important for host infection and H3 lysine 27 methylation. We show that by experimentally changing histone modifications, many fungal genes normally used during plant infection are turned on outside of the host. Furthermore, we detail how histone modifications can change naturally in the fungus during plant infection. These findings help broaden our knowledge of genome regulation for these pathogens, and advances the goal of a more comprehensive understanding of the infection process.

Molecular dating using phylogenetics allows us to estimate the date of an infection from time-stamped within-host sequences alone. There are large datasets of HIV-1 sequences, but genome and gene analyses are not often performed in parallel and rarely with the possibility to compare results against a known narrow window of infection. We showed that all but the longest genes are near-clonal in acute infection, with little information for dating purposes. For infections with single founders, we estimated the eclipse phase—the time between HIV-1 exposure and the first positive diagnostic test—to last between one and two weeks using env, gag, pol and near full-length genomes. This approach could be used to narrow the date of suspected infection in ongoing clinical trials for the prevention of HIV-1 infection.

In the eukaryotic cell nucleus, DNA loops bring together gene promoters and distal regulatory elements to tune gene expression. To adapt to the 24h periodic changes in the environment, such as light-dark cycles, many organisms on the planet adopted an internal circadian clock encoded genetically. Thus, animal physiology resonates with external cycles. In mouse liver, thousands of genes, including ones belonging to the central clock, lipid and glucose metabolism, as well as detoxification are expressed with a 24h period. Here, we monitored DNA contacts across the circadian cycle involving the promoters of genes playing key roles in the daily physiology of the mouse liver. While the dynamics of some loops oscillates in sync with gene expression along the 24h day, other loops connected daily active regulatory elements to target gene promoters stably over time. Using arrhythmic mutant animals, we demonstrated that the maintenance of such stable loops goes beyond the circadian clock function. Also, we discovered a set of frequently interacting genes having coherent temporal expression regulation in wild type and clock impaired animals. Thus, both dynamic and static DNA contacts underlie the circadian transcription of mouse liver physiology.

Developmental arrest pervades organismal development and physiology where it facilitates an enormous range of adaptive responses to stressful conditions. Many animals exhibit various forms of developmental arrest that ensures survival under the most adverse environments. Reproductive diapause occurs when adults temporarily suspend reproduction in response to environmental stress and has been documented for a variety of invertebrates, particularly insects. Endocrine signals play a central role in translating environmental cues such as photoperiod into reproductive diapause-related physiology and behavior. However, it has been an unresolved issue as to which endocrine factors can respond to photoperiodic inputs and regulate diapause outputs. In this study, we found that a decrease in ecdysone levels is critical for reproductive diapause to occur. Also, ecdysone could interact with juvenile hormone to regulate the occurrence of reproductive diapause in response to photoperiodic cues. Our findings provide new insight into endocrine mechanisms of photoperiodic reproductive diapause and an example of phenotypic plasticity in animals.

Vaccinia virus (VACV) is a large DNA virus with an acute and increasing demand for energy and macromolecules to build hundreds and thousands of viral particles in a single cell within hours of infection. The demand postulates reprogramming of the TCA cycle, as it is the central metabolic hub of a cell that generates metabolites for energy production and macromolecule synthesis. We show that VACV infection reprograms cellular metabolism globally, elevating the TCA cycle intermediate levels and modulating related cell metabolism. The elevation of the TCA cycle intermediates depends on the virus-encoded growth factor that stimulates non-canonical STAT3 signaling during VACV infection. Our results provide the metabolic foundation of viral growth factor to boost VACV infection.

We first gather information from literature and build a logical model summarizing the regulatory network of the mitogen-activated protein kinase (MAPK) pathway surrounding BRAF, with factors involved in the BRAF inhibition resistance mechanisms. The relevance of this model is verified by automatically assessing that it qualitatively reproduces response or resistance behaviors identified in the literature. Data from over 100 melanoma and colorectal cancer cell lines are then used to validate the model’s ability to explain differences in sensitivity. This generic model is transformed into personalized cell line-specific logical models by integrating the omics information of the cell lines as constraints of the model. The use of mutations alone allows personalized models to correlate significantly with experimental sensitivities to BRAF inhibition, both from drug and CRISPR targeting, and even better with the joint use of mutations and RNA, supporting multi-omics mechanistic models. A comparison of these untrained models with learning approaches highlights similarities in interpretation and complementarity depending on the size of the datasets.

This parsimonious pipeline, which can easily be extended to other biological questions, makes it possible to explore the mechanistic causes of the response to treatment, on an individualized basis.

We constructed a logical model to study, from a dynamical perspective, the differences between melanomas and colorectal cancers that share the same BRAF mutations but exhibit different sensitivities to anti-BRAF treatments. The model was built from the literature and completed from existing pathway databases. The model encompasses the key proteins of the MAPK pathway and was made specific to each cancer cell line (100 melanoma and colorectal cell lines from public database) using available omics data, including mutations and RNAseq data. It can simulate the effect of drugs and show high correlation with experimental results. Moreover, the structure of the network confirms both the importance of the reactivation of the MAPK pathway through CRAF and the involvement of PI3K/AKT pathway in the mechanisms of resistance to BRAF inhibition.

The study shows that, because of the low number of samples, the mechanistic approach that we propose provides different insights than powerful standard machine learning methodologies would, showing the complementarity between the two approaches. An important aspect to mention is that the mechanistic approach presented here does not rely on training datasets but directly interprets and maps data on the model to simulate drug responses.

Ebola virus (EBOV) and Marburg virus (MARV) are zoonotic pathogens that can cause severe hemorrhagic fevers in humans and non-human primates. They are members of the growing Filoviridae family that also includes three other species of Ebolaviruses known to be highly pathogenic in humans. While vaccines for EBOV are being deployed and showed high efficacy, pan-filoviral treatment is still lacking. To infect cells, EBOV requires the endosomal/lysosomal resident protein Niemann-Pick C1 (NPC1). Accordingly, viral particles require extensive trafficking within endosomal pathways for entry and delivery of the viral genome into the host cell cytoplasm. Here, we used chemical biology to reveal that EBOV triggers receptor tyrosine kinase (RTK)-dependent signaling to traffic to intracellular vesicles that contain the receptor and are conducive to entry. The characterization of host trafficking factors and signaling pathways that are potentially triggered by the virus are important as these could be targeted for antiviral therapies. In our study, we identified several RTK inhibitors, some of which are FDA-approved drugs, that potently block EBOV infection. Since all filoviruses known to date, even Měnglà virus that was recently discovered in bats in China, use NPC1 as their entry receptor, these inhibitors have the potential to be effective pan-filovirus antivirals.

Optogenetics is a novel technique involving the targeted delivery of light-sensitive ion channels like Channelrhodopsin-2 (ChR2) to cells. Recently, this technique has been expanded to non-neuronal cell types, e.g., astrocytes. Optogenetic control of astrocytes in vivo can aid in further understanding the intricacies of their debated roles in the brain. Here, using a mathematical model, we evaluate the effect of short and long-term light stimulation of ChR2-expressing astrocytes on their Ca²⁺ spiking activity and basal level. We further investigate how ChR2 gating dynamics, buffering, and coupling coefficient of Ca²⁺ influence astrocytic activity in a single cell and a network. Results of our study can serve as a tool for experimental design and engineering of new application-oriented optogenetic constructs targeting astrocytes.

Angiogenesis, the process by which new blood vessels are formed by sprouting from the pre-existing vascular bed, plays a key role in both physiological and pathological processes, including tumour growth. The structure of a growing vascular network is determined by the coordinated behaviour of endothelial cells in response to various signalling cues. Recent experimental studies have highlighted the importance of cell rearrangements as a driver for sprout elongation. However, the functional role of this phenomenon remains unclear. We formulate a new multiscale model of angiogenesis which, by accounting explicitly for the complex dynamics of endothelial cells within growing angiogenic sprouts, is able to reproduce generic features of angiogenic structures (branching, chemotactic sensitivity, cell mixing, etc.) as emergent properties of its dynamics. We validate our model against experimental data and then use it to quantify the phenomenon of cell mixing in vascular networks generated by endothelial cells of different lineages. Our results show that there is a direct correlation between the time evolution of cell mixing in a growing vascular network and its branching structure, thus paving the way for understanding the functional role of cell rearrangements in angiogenesis.

Rsp5, the Nedd4 family member in yeast, is an E3 ubiquitin ligase involved in numerous cellular processes, many of which require Rsp5 to interact with PY-motif containing adaptor proteins. Here, we show that two paralogous transmembrane Rsp5 adaptors, Rcr1 and Rcr2, are sorted to distinct cellular locations: Rcr1 is a plasma membrane (PM) protein, whereas Rcr2 is sorted to the vacuole. Rcr2 is delivered to the vacuole using ubiquitin as a sorting signal. Rcr1 is delivered to the PM by the exomer complex using a newly uncovered PM sorting motif. Further, we show that Rcr1, but not Rcr2, is up-regulated via the calcineurin/Crz1 signaling pathway. Upon exogenous calcium treatment, Rcr1 ubiquitinates and down-regulates the chitin synthase Chs3. We propose that the PM-anchored Rsp5/Rcr1 ubiquitin ligase-adaptor complex can provide an acute response to degrade unwanted proteins under stress conditions, thereby maintaining cell integrity.

Different environmental isolates (strains) of the same microbial species can vary greatly in their ability to cause disease, ranging from avirulent to hypervirulent. What makes some strains deadly pathogens, while others are relatively benign? This study describes the characterization of key genetic differences that underlie variation in traits thought to promote virulence in Cryptococcus deneoformans, a wide-spread opportunistic fungal pathogen. Using a combination of quantitative genetic and molecular genetic approaches we dissected the genetic architecture of virulence-related cellular traits (melanin production and the production of a polysaccharide capsule), physiological responses to stress (tolerance of thermal, oxidative, and osmotic stress), and sensitivity to multiple antifungal drugs. Strikingly we find that variation in most of these traits is governed by a small number of genetic differences that modify the function of two major cell signaling networks, cyclic AMP–Protein Kinase A (cAMP-PKA) signaling and a fungal specific MAP-kinase cascade called the high osmolarity glycerol (HOG) pathway. Similar to recent studies in a number of other fungal species, our findings point to an outsize role for a small number of highly pleiotropic signaling pathways in potentiating phenotypic variation both within and between fungal species.

This study tested three models of how a BMP morphogen gradient is translated into differential gene activation that specifies distinct cell fates, finding that BMP signal concentration thresholds, not gradient shape or signal duration, position three distinct gene activation domains.

Previous work has shown that substrates of Post-Translational Modification (PTM) cycles can have coupled responses if those substrates share enzymes. This implies that modifications leading to substrate degradation (e.g. ubiquitylation by an E3 ligase) could introduce coupling in concentrations of substrates sharing a ligase. Using mathematical models, we found adding protein turnover to a PTM cycle diminishes both sensitivity and ultrasensitivity, particularly in models admitting long ubiquitin chains. We also found that proteins sharing an E3 ligase can indeed have coupled changes in both expression and sensitivity to signals. These results imply that accounting for crosstalk in protein degradation networks is crucial for the interpretation of results from a wide variety of common experimental perturbations to living systems.

Human papillomavirus (HPV) infections occur in differentiating squamous epithelium and induce hyperplasia during the viral replicative cycle. Although HPV oncogene expression is necessary to promote cellular proliferation for viral genome amplification in the middle epithelial layers, oncogene levels are thereafter suppressed to permit differentiation-induced late gene expression in the uppermost epithelial cells. Yet, the mechanisms responsible for controlling HPV oncogene expression are not well understood. Here, we demonstrate that EGFR/MEK/ERK signaling, which is subject to the normal cellular cues of contact inhibition and epithelial tissue differentiation, is a critical regulator of hr-HPV oncogene expression. We found that extrinsic activation of ERK overrides cellular control to promote oncogene expression and the neoplastic phenotype. Many epidemiologically defined risk factors activate the EGFR/MEK/ERK pathway, suggesting a common mechanism whereby they may promote HPV persistence and disease progression. Lastly, we show that HPV oncogene transcription and protein expression remain susceptible to MEK/ERK control in early neoplastic tissues and tumor cells and that targeted inhibition of MEK/ERK signaling might be exploited therapeutically for HPV-induced infections and tumors.

Bone loss in osteoporosis and many other degenerative bone diseases is characterized by decreased bone formation and increased fat accumulation in the bone marrow, especially during aging, and many are suspected to have a genetic basis. Bone marrow mesenchymal stem cells (BMSCs) can differentiate into adipocytes, chondrocytes, and osteoblasts. BMSCs from aging or genetic defect subjects are less likely to differentiate into osteoblasts while more inclined to differentiate into adipocytes. However, the molecular mechanisms remain elusive. Here, we describe a new mutant mice model-Runx1 mesenchymal progenitor-specific and osteoblast-specific conditional knockout (Runx1 CKO) mice, which showed the severe bone formation defects and adipocytes accumulation. Here, we revealed that Runx1 enhances osteoblast lineage commitment promotes bone formation and inhibits adipogenesis by up-regulating the Bmp7/Alk3/Smad1/5/8/Runx2/ATF4 and WNT/β-catenin signaling pathways and orchestrating multiple signaling pathways involved in bone formation. These findings further elucidated the roles of Runx1 in bone homeostasis with implications into development of novel therapeutic strategies for osteoporosis as well as other degenerative bone diseases.

Tuberculosis is a disease caused by Mycobacterium tuberculosis (Mtb), a bacterium that infects over a third of the world’s population. The only available vaccine for TB has limited efficacy and drug treatment involves several antibiotics that are taken for several months. These drugs can have significant side-effects and a lack of compliance can lead to drug resistance in Mtb. A hallmark of Mtb infection is the development of clusters of cells that form around infected macrophages to contain the infection called granulomas. Older granulomas, or granulomas in patients treated with antibiotic, often become fibrotic and this can cause chronic lung problems long after the Mtb infection has cleared. The process that drives a fibrotic outcome has been difficult to assess in vivo. Herein we combined wet-lab and computational experimentation to identify a novel mechanism leading to peripheral fibrosis in granulomas. We find that macrophages transforming into myofibroblast-like cells, may be a key pathway to granuloma-associated fibrosis, a phenomena that has not been well characterized in vivo. Further we identified factors that can inhibit fibrosis development during TB that could be therapeutic targets during TB treatment to limit the risk of long-term tissue damage in TB.

Basal bodies (BBs) are macromolecular complexes required for the formation and cortical positioning of cilia. Both BB assembly and DNA replication are tightly coordinated with the cell cycle to ensure their accurate segregation and propagation to daughter cells, but the mechanisms ensuring coordination are unclear. The Tetrahymena Sas4/CPAP protein is enriched at assembling BBs, localizing to the core BB structure and to the base of BB-appendage microtubules and striated fiber. Sas4 is necessary for BB assembly and cortical microtubule organization, and Sas4 loss disrupts cell division furrow positioning and DNA segregation. The Hippo signaling pathway is known to regulate cell division furrow position, and Hippo molecules localize to BBs and BB-appendages. We find that Sas4 loss disrupts localization of the Hippo activator, Mob1, suggesting that Sas4 mediates Hippo activity by promoting scaffolds for Mob1 localization to the cell cortex. Thus, Sas4 links BBs with an ancient signaling pathway known to promote the accurate and symmetric segregation of the genome.

Type II DNA topoisomerases (TOP2) are a double-edged sword. They solve topological problems in the form of supercoiling, knots and tangles that inevitably accompany genome metabolism, but they do so at the cost of transiently cleaving DNA, with the risk that this entails for genome integrity, and the serious consequences for human health, such as neurodegeneration, developmental disorders or predisposition to cancer. A comprehensive analysis of TOP2 distribution throughout the genome is therefore essential for a deep understanding of its function and regulation, and how this can affect genome dynamics and stability. Here, we use machine learning to thoroughly explore genome-wide binding of TOP2B, a vertebrate TOP2 paralog that has been linked to genome organization and cancer-associated translocations. Our analysis shows that TOP2B-DNA binding can be accurately predicted exclusively using information on DNA accessibility and binding of genome-architecture factors. We show that such information is enough to generate virtual maps of TOP2B binding along the genome, which we validate with de novo experimental data. Our results highlight the importance of TOP2B for accessibility and 3D organization of chromatin, and show that computationally predicted TOP2 maps can be accurately obtained using minimal publicly available datasets, opening the door for their use in different organisms, cell types and conditions with experimental and/or clinical relevance.

Mitochondrial dysfunction has been implicated in life-threatening cardiac arrhythmias. Experimentalists have shown that mitochondrial depolarization could cause period-doubling behavior in cytosolic Ca²⁺ transient (i.e., Ca²⁺ alternans). However, the underlying mechanisms behind these experimental observations remain unclear. Here, we focus on revealing the underlying mechanisms of cellular Ca²⁺ alternans in the context of mitochondrial depolarization. Using our newly developed action potential model that incorporates a 3D network of Ca²⁺ release unit with a 3D network of mitochondria, we dissect out individual roles of mitochondrial Ca²⁺ and complex signaling pathways (e.g., reactive oxygen species, oxidized Ca²⁺/calmodulin-dependent protein kinase II activation, and ATP) in the genesis of cellular Ca²⁺ alternans. The key finding is that mitochondrial depolarization promotes Ca²⁺ alternans acutely via the redox effect of ROS and chronically by ATP reduction, and oxidized Ca²⁺/calmodulin-dependent protein kinase II activation may suppress Ca²⁺ alternans chronically. We believe that this study reveals insightful information for understanding the mechanisms of the genesis of Ca²⁺ alternans in the context of mitochondrial depolarization and provides valuable simulation results to guide future experiments.

The neural crest is a migratory stem cell population that plays a crucial role in the development of vertebrate embryos. These cells contribute to multiple tissues and organs, including the craniofacial skeleton, the peripheral nervous system, and the pigmentation of the skin. Neural crest formation requires sequential expression of sets of genes that gradually change the identity of progenitor cells. Here we characterize a gene circuit that plays an important role in these developmental transitions. We show that a pair of transcription factors downstream of the Wingless (Wnt) signaling pathway (SP5 and AXUD1) control the temporal activation of neural crest genes. While SP5 drives the expression of genes expressed at gastrula stages, AXUD1 promotes neural crest genes transcribed during neurulation. We found that these regulators are linked by inhibitory interactions that prevent overlap between transcriptional states. Our results shed light on how the architecture of gene regulatory circuits underlies the timing of developmental transitions during neural crest development.

When and where genes are expressed is controlled by regulatory DNA regions known as enhancers. Genes often have multiple enhancers that control expression in different cell types or embryonic regions, but there are also genes that have multiple enhancers that control near-identical expression patterns. These ‘shadow’ enhancers are common features of many animal genomes, but it is unclear to what extent they are truly identical in function. Here we studied a pair of shadow enhancers for the notochord-specific gene Brachyury in the simple model chordate Ciona that are both directly activated by the same signaling pathway. We titrated the activity of this pathway with graded doses of a pharmacological inhibitor and measured the effects in quantitative enhancer assays. We found that the two enhancers had significant differences in sensitivity and cooperativity to the same shared regulator and are thus not identical in function when assessed quantitatively. We also identified subtle differences in sensitivity to this upstream signal between different notochord precursor cells.

The symbiotic bacteria that colonize the human intestine are major regulators of the immune system. While the collective ability of these organisms to shape immune function is clear the individual impact of different symbionts on immunity is poorly delineated. To address this, we have systematically characterized the immunological properties of an extensive panel of commensals from the major taxa in the human intestinal microbiota. We have uncovered the unique complement of pattern recognition receptors, intracellular signaling components and transcription factors through which each symbiotic species exerts their effect on the innate immune system. This allowed us to identify, for the first time, taxonomic patterns of commensal regulation of immunity and uncover a unique role for canonical NF-κB signaling in acting as a rheostat calibrating the degree of immune stimulation symbionts elicit. We used the immunological profiles of these symbionts to guide the construction of therapeutic symbiont communities to combat both AMR infection and pathological intestinal inflammation. Previously, harnessing the power of symbiotic microbes to combat disease was largely a trial-and-error process, and predicting how changes in microbiota composition will impact immunity was almost impossible. Our study addresses both of these challenges and provides a unique scientific resource to guide the rational design of symbiotic microbial communities to treat myriad diseases and reveals how loss of different symbionts will impact host health.

Introduction

Interleukin-34 (IL-34) shares a receptor (cFMS) with colony stimulating factor-1 (CSF-1), and these two ligands mediate macrophage proliferation. However, in contrast to CSF-1, the influence of IL-34 on tubular epithelial cells (TECs) injury remains unclear. We investigated the physiological effects of IL-34 on TEC damage caused by cisplatin nephrotoxicity (CP-N).

Our tissues are subject to chronic physiological and environmental damage, yet little is known about how healthy human cells normally respond to stress. We examined the effect of damage on cells obtained from breast tissue of disease-free women. Unexpectedly, we identified a well-known protein regulator of chromosomal function, CTCF, as a robust target of stress signals. In normal mammary cells, a pool of CTCF is localized to large “depots” within the nucleus that regulate RNA processing. Upon cellular damage, CTCF rapidly disappears from nuclear “depots” by stress-inducible protein degradation while genome occupancy by CTCF is relatively unaffected. We observe the same phenomenon in neuronal progenitors differentiated from human pluripotent stem cells. We propose that in specific cell types, stress-sensitive forms of CTCF exist that have a unique function in RNA metabolism potentially by fine-tuning gene expression near nuclear speckles, which may maintain cells in a progenitor or adaptive state. Upon stress, this particular CTCF function is rapidly disabled, which may change the identity of cells most vulnerable to disease in order to safeguard them from becoming dysfunctional. Persistently stressed cells have lost this CTCF function, which may facilitate the genesis of damage-induced cancer initiation and neuro-degeneration.

Background

Radioiodide (¹³¹I) is commonly used to treat thyroid cancer and hyperthyroidis.¹³¹I released during nuclear accidents, have resulted in increased incidence of thyroid cancer in children. Therefore, a better understanding of underlying cellular mechanisms behind ¹³¹I exposure is of great clinical and radiation protection interest. The aim of this work was to study the long-term dose-related effects of ¹³¹I exposure in thyroid tissue and plasma in young rats and identify potential biomarkers.

Background

Sorghum bicolor (SB) is rich in protective phytoconstituents with health benefits and regarded as a promising source of natural anti-diabetic substance. However, its comprehensive bioactive compound(s) and mechanism(s) against type-2 diabetes mellitus (T2DM) have not been exposed. Hence, we implemented network pharmacology to identify its key compounds and mechanism(s) against T2DM.

An intracellular signaling pathway is often represented by a set of nonlinear ordinary differential equations, which translate our current knowledge about the signaling pathway into a testable mathematical model. However, predictions from such models are often subject to high uncertainty since many signaling pathways are only partially known beforehand. In this study, we propose a systematic approach to develop a hybrid model to improve model accuracy by combining machine learning and the first-principle modeling. Specifically, model correction terms are learned from discrepancy between model predictions and measurements, and these terms are added to the first-principle model to enhance the prediction accuracy. Once these correction terms are learned from the data, an artificial neural network (ANN) model is developed to find an empirical relation between the model and the correction terms so that the developed ANN can be used to posses improved predictive capabilities even in new operating conditions (i.e., generalizability). The final hybrid model is then constructed by coupling the first-principle model with the developed ANN.

A stem cell niche is the specialized micro-environment that provides the signal to the resident stem cells to support their undifferentiated, self-renewing state. This study shows that the cells that compose the niche do not only provide the signal, but also take up the receptor of stem cells for subsequent lysosomal degradation; this mechanism is essential for restriction of niche signal range.

Despite the known importance of mesenchymal cells for the homeostasis of the intestinal system, how these cells communicate with the associated epithelial layer remains unclear. A single cell atlas of matched colonic epithelium and mesenchyme identifies two fibroblast populations that orchestrate maintenance and differentiation of colonic epithelial stem cells.

Pathological cardiac hypertrophy is a disease in which the heart grows abnormally in response to different motivators such as high blood pressure or variations in hormones and growth factors. The shape of the heart after its growth depends on the context in which it grows. Since cell signaling in the cardiac cells plays a key role in the determination of heart shape, a thorough understanding of cardiac cells signaling in each context enlightens the mechanisms which control response of cardiac cells. However, cell signaling in cardiac hypertrophy comprises a complex web of pathways with numerous interactions, and predicting how these interactions control the hypertrophic signal in each context is not achievable by only experiments or general computational models. To address this need, we developed an approach to bring together the experimental data of each context with a signaling network curated from literature to identify the main players of cardiac cells response in each context and attain the context-specific models of cardiac hypertrophy. By utilizing our approach, we identified the main regulators of cardiac hypertrophy in four important contexts. We developed a network model of β-adrenergic cardiac hypertrophy, and predicted and validated new interactions that regulate cardiac cells response in this context.

Objective

Mature hair follicles represent an important stage of hair follicle development, which determines the stability of hair follicle structure and its ability to enter the hair cycle. Here, we used weighted gene co-expression network analysis (WGCNA) to identify hub genes of mature skin and hair follicles in Inner Mongolian cashmere goats.

At presynaptic active zones, arrays of large conserved scaffold proteins mediate fast and temporally precise release of synaptic vesicles (SVs). SV release sites could be identified by clusters of Munc13, which allow SVs to dock in defined nanoscale relation to Ca²⁺ channels. We here show in Drosophila that RIM-binding protein (RIM-BP) connects release sites physically and functionally to the ELKS family Bruchpilot (BRP)-based scaffold engaged in SV recruitment. The RIM-BP N-terminal domain, while dispensable for SV release site organization, was crucial for proper nanoscale patterning of the BRP scaffold and needed for SV recruitment of SVs under strong stimulation. Structural analysis further showed that the RIM-BP fibronectin domains form a “hinge” in the protein center, while the C-terminal SH3 domain tandem binds RIM, Munc13, and Ca²⁺ channels release machinery collectively. RIM-BPs’ conserved domain architecture seemingly provides a relay to guide SVs from membrane far scaffolds into membrane close release sites.

Cellular communication is critical in controlling the growth of organs and must be carefully regulated to prevent disease. The Ras signaling pathway is frequently used for cellular communication of tissue growth regulation but can operate at different signaling strengths. Here, we used a novel strategy to identify genes that specifically tune weak or strong Ras signaling states. We find that the gene RpS21 preferentially tunes weak Ras signaling states.

The Yin and Yang of TNF-α signaling in heart failure: this Primer explores the history of failure of TNF-α antagonistic therapy in heart failure and how a recent study suggests that inhibition of TNF-α converting enzyme TACE might be a way out of this dilemma.

Tissue damage triggers a rapid and robust inflammatory response in order to clear and repair a wound. Remarkably, many of the cell biology features that underlie the ability of leukocytes to home in to sites of injury and to fight infection—most of which are topics of intensive current research—were originally observed in various weird and wonderful translucent organisms over a century ago by Elie Metchnikoff, the “father of innate immunity,” who is credited with discovering phagocytes in 1882. In this review, we use Metchnikoff’s seminal lectures as a starting point to discuss the tremendous variety of cell biology features that underpin the function of these multitasking immune cells. Some of these are shared by other cell types (including aspects of motility, membrane trafficking, cell division, and death), but others are more unique features of innate immune cells, enabling them to fulfill their specialized functions, such as encapsulation of invading pathogens, cell–cell fusion in response to foreign bodies, and their self-sacrifice as occurs during NETosis.