Although ectopic overexpression of miRNAs can influence mammary normal and cancer stem cells (SCs/CSCs), their physiological relevance remains uncertain. Here, we show that miR-146 is relevant for SC/CSC activity. MiR-146a/b expression is high in SCs/CSCs from human/mouse primary mammary tissues, correlates with the basal-like breast cancer subtype, which typically has a high CSC content, and specifically distinguishes cells with SC/CSC identity. Loss of miR-146 reduces SC/CSC self-renewal in vitro and compromises patient-derived xenograft tumor growth in vivo, decreasing the number of tumor-initiating cells, thus supporting its pro-oncogenic function. Transcriptional analysis in mammary SC-like cells revealed that miR-146 has pleiotropic effects, reducing adaptive response mechanisms and activating the exit from quiescent state, through a complex network of finely regulated miRNA targets related to quiescence, transcription, and one-carbon pool metabolism. Consistent with these findings, SCs/CSCs display innate resistance to anti-folate chemotherapies either in vitro or in vivo that can be reversed by miR-146 depletion, unmasking a “hidden vulnerability” exploitable for the development of anti-CSC therapies.

During translation initiation, 40S ribosomes scan the mRNA until they encounter the start codon, where conformational changes produce a translation-competent 80S complex. Destabilizing the scanning complex results in misinitiation at non-AUG codons, demonstrating its importance for fidelity. Here, we use a combination of biochemical and genetic analyses to demonstrate that the ability of the nascent subunit to adopt the scanning complex is tested during assembly via structural mimicry. Specifically, formation of the 80S-like assembly intermediate, which structurally resembles scanning complexes, requires the correct folding of two rRNA elements in the subunit head and the proper positioning of the universally conserved head proteins Rps3, Rps15, Rps20, and Rps29. rRNA misfolding impairs the formation of 80S-like ribosomes, and bypass of individual checkpoints using cancer-associated mutations produces ribosomes defective in accurate start-site selection. Thus, the formation of 80S-like assembly intermediates is a quality control step that ensures scanning competence of the nascent subunit.

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.

In mammals, argonaute (AGO) proteins have been characterized for their roles in small RNA–mediated posttranscriptional and also in transcriptional gene silencing. Here, we report a different role for AGO1 in estradiol-triggered transcriptional activation in human cells. We show that in MCF-7 mammary gland cells, AGO1 associates with transcriptional enhancers of estrogen receptor α (ERα) and that this association is up-regulated by treating the cells with estrogen (E2), displaying a positive correlation with the activation of these enhancers. Moreover, we show that AGO1 interacts with ERα and that this interaction is also increased by E2 treatment, but occurs in the absence of RNA. We show that AGO1 acts positively as a coactivator in estradiol-triggered transcription regulation by promoting ERα binding to its enhancers. Consistently, AGO1 depletion decreases long-range contacts between ERα enhancers and their target promoters. Our results point to a role of AGO1 in transcriptional regulation in human cells that is independent from small RNA binding.

Stress granules (SGs) are evolutionarily conserved condensates of ribonucleoproteins that assemble in response to metabolic stresses. Because aberrant SG formation is associated with amyotrophic lateral sclerosis (ALS), understanding the connection between metabolic activity and SG composition can provide therapeutic insights into neurodegeneration. Here, we identify 17 metabolic enzymes recruited to yeast SGs in response to physiological growth stress. Furthermore, the product of one of these enzymes, AdoMet, is a regulator of SG assembly and composition. Decreases in AdoMet levels increase SG formation, while chronic elevation of AdoMet produces SG remnants lacking proteins associated with the 5′ end of transcripts. Interestingly, acute elevation of AdoMet blocks SG formation in yeast and motor neurons. Treatment of ALS-derived motor neurons with AdoMet also suppresses the formation of TDP-43–positive SGs, a hallmark of ALS. Together, these results argue that AdoMet is an evolutionarily conserved regulator of SG composition and assembly with therapeutic potential in neurodegeneration.

Stress granules are dynamic assemblies of proteins and nontranslating RNAs that form when translation is inhibited in response to diverse stresses. Defects in ubiquitin–proteasome system factors including valosin-containing protein (VCP) and the proteasome impact the kinetics of stress granule induction and dissolution as well as being implicated in neuropathogenesis. However, the impacts of dysregulated proteostasis on mRNA regulation and stress granules are not well understood. Using single mRNA imaging, we discovered ribosomes stall on some mRNAs during arsenite stress, and the release of transcripts from stalled ribosomes for their partitioning into stress granules requires the activities of VCP, components of the ribosome-associated quality control (RQC) complex, and the proteasome. This is an unexpected contribution of the RQC system in releasing mRNAs from translation under stress, thus identifying a new type of stress-activated RQC (saRQC) distinct from canonical RQC pathways in mRNA substrates, cellular context, and mRNA fate.