Highlights d Global profiling of mRNA decay pathways and aberrant translation events in mESCs d XRN1 mediates mRNA turnover, whereas SKIV2L acts widely in translation surveillance d AVEN interacts with ribosomes and the Ski complex and counteracts ribosome stalling d Histone mRNAs, uORFs, and small ORFs are key targets of SKIV2L and AVEN

Acid-sensing ion channels (ASICs) are key receptors for extracellular protons. These neuronal nonvoltage-gated Na ؉ channels are involved in learning, the expression of fear, neurodegeneration after ischemia, and pain sensation. We have applied a systematic approach to identify potential pH sensors in ASIC1a and to elucidate the mechanisms by which pH variations govern ASIC gating. We first calculated the pK a value of all extracellular His, Glu, and Asp residues using a Poisson-Boltzmann continuum approach, based on the ASIC three-dimensional structure, to identify candidate pH-sensing residues. The role of these residues was then assessed by site-directed mutagenesis and chemical modification, combined with functional analysis. The localization of putative pH-sensing residues suggests that pH changes control ASIC gating by protonation/ deprotonation of many residues per subunit in different channel domains. Analysis of the function of residues in the palm domain close to the central vertical axis of the channel allowed for prediction of conformational changes of this region during gating. Our study provides a basis for the intrinsic ASIC pH dependence and describes an approach that can also be applied to the investigation of the mechanisms of the pH dependence of other proteins.Acid-sensing ion channels (ASICs) 3 are neuronal nonvoltage-gated Na ϩ channels that are activated by a rapid drop in extracellular pH (1, 2). They are members of the epithelial Na ϩ channel/Degenerin family of ion channel proteins (3). Expression in nociceptive neurons and activation by protons suggest that ASICs may act as pain receptors (4), and evidence for such a role has been provided in several animal pain models (5-7). ASIC1a in the central nervous system plays a role in memory formation and the expression of fear (8, 9). ASIC1a is also an important mediator of cell injury induced by conditions associated with acidosis in the mammalian nervous system (10). Functional ASICs are formed by homo-or heterotrimeric assembly of ASIC subunits 1a, 1b, 2a, 2b, and 3. Each subunit has short intracellular N and C termini and two transmembrane domains that are separated by a large extracellular domain.Extracellular acidification opens ASICs. The ASIC activity is terminated in the continued presence of the acidic stimulus within hundreds of milliseconds to seconds by open channel inactivation, which is also called desensitization (11). Only in some ASIC isoforms and under certain pH conditions can acidification induce a sustained current, which has in most cases a much smaller amplitude than the peak current (12-14). At pH values slightly below the physiological pH, ASICs inactivate without apparent channel opening in a process that is called steady-state inactivation (SSIN) (15). By these two ways ASICs enter the inactivated state, which is a nonconducting, absorbing state. Experimental protocols have been applied to determine the kinetics of the recovery from the inactivated state, showing that channels need exposure for a certain duration ...

MicroRNA (miRNA) loaded Argonaute (AGO) complexes regulate gene expression via direct base-pairing with their mRNA targets. Current prediction approaches identified that between 20 to 60% of mammalian transcriptomes are regulated by miRNAs, but it remains largely unknown which fraction of these interactions are functional in a specific cellular context. Here, we integrated transcriptome data from a set of miRNA-depleted mouse embryonic stem cell (mESC) lines with published miRNA interaction predictions and AGO-binding profiles. This integrative approach, combined with molecular validation data, identified that only 6% of expressed genes are functionally and directly regulated by miRNAs in mESCs. In addition, analyses of the stem cell-specific miR-290-295 cluster target genes identified TFAP4 as an important transcription factor for early development. The extensive datasets developed in this study will support the development of improved predictive models for miRNA-mRNA functional interactions.

MicroRNA (miRNA) loaded Argonaute (AGO) complexes regulate gene expression via direct base pairing with their mRNA targets. Previous works suggest that up to 60% of mammalian transcripts might be subject to miRNA-mediated regulation, but it remains largely unknown which fraction of these interactions are functional in a specific cellular context. Here, we integrate transcriptome data from a set of miRNA-depleted mouse embryonic stem cell (mESC) lines with published miRNA interaction predictions and AGO-binding profiles. Using this integrative approach, combined with molecular validation data, we present evidence that < 10% of expressed genes are functionally and directly regulated by miRNAs in mESCs. In addition, analyses of the stem cell-specific miR-290-295 cluster target genes identify TFAP4 as an important transcription factor for early development. The extensive datasets developed in this study will support the development of improved predictive models for miRNA-mRNA functional interactions.