bims-micpro Biomed News
on Discovery and characterization of microproteins
Issue of 2022‒12‒11
five papers selected by
Thomas Farid Martínez
University of California, Irvine

  1. Mol Oncol. 2022 Dec 10.
      The biological functions of short open reading frame (sORF)-encoded micropeptides remain largely unknown. Here, we report that LINC00998, a previously annotated lncRNA, was upregulated in multiple cancer types and the sORF on LINC00998 encoded a micropeptide named SMIM30. SMIM30 was localized in the membranes of the endoplasmic reticulum (ER) and mitochondria. Silencing SMIM30 inhibited the proliferation of hepatoma cells in vitro and suppressed the growth of tumor xenografts and N-nitrosodiethylamine-induced hepatoma. Overexpression of the 5'UTR-sORF sequence of LINC00998, encoding wild-type SMIM30, enhanced tumor cell growth, but this was abolished when a premature stop codon was introduced into the sORF via single-base deletion. Gain- and loss-of-function studies revealed that SMIM30 peptide but not LINC00998 reduced cytosolic calcium level, increased CDK4, cyclin E2, phosphorylated-Rb and E2F1, and promoted the G1/S phase transition and cell proliferation. The effect of SMIM30 silencing was attenuated by a calcium chelator or the agonist of sarco/endoplasmic reticulum calcium ATPase (SERCA) pump. These findings suggest a novel function of micropeptide SMIM30 in promoting G1/S transition and cell proliferation by enhancing SERCA activity and reducing cytosolic calcium level.
  2. Nat Commun. 2022 Dec 06. 13(1): 7510
      Half of mammalian transcripts contain short upstream open reading frames (uORFs) that potentially regulate translation of the downstream coding sequence (CDS). The molecular mechanisms governing these events remain poorly understood. Here, we find that the non-canonical initiation factor Death-associated protein 5 (DAP5 or eIF4G2) is required for translation initiation on select transcripts. Using ribosome profiling and luciferase-based reporters coupled with mutational analysis we show that DAP5-mediated translation occurs on messenger RNAs (mRNAs) with long, structure-prone 5' leader sequences and persistent uORF translation. These mRNAs preferentially code for signalling factors such as kinases and phosphatases. We also report that cap/eIF4F- and eIF4A-dependent recruitment of DAP5 to the mRNA facilitates main CDS, but not uORF, translation suggesting a role for DAP5 in translation re-initiation. Our study reveals important mechanistic insights into how a non-canonical translation initiation factor involved in stem cell fate shapes the synthesis of specific signalling factors.
  3. Nucleic Acids Res. 2022 Dec 08. pii: gkac1140. [Epub ahead of print]
      Control of mRNA translation is key for stress responses. Translation initiation is usually rate-limiting and, in eukaryotes, involves mRNA scanning by the small ribosomal subunit. Despite its importance, many aspects of translation in vivo have not been explored fully, especially at the transcriptome-wide level. A recent method termed translation-complex profiling (TCP-seq) allows transcriptome-wide views of scanning ribosomal subunits. We applied TCP-seq to nutritional stress in the fission yeast Schizosaccharomyces pombe. At initiation sites, we observed multiple complexes resembling those of mammals, and consistent with queuing of scanning subunits. In 5' UTRs, small subunit accumulations were common and may reflect impediments to scanning. A key mediator of stress responses in S. pombe is the Fil1 transcription factor, which is regulated translationally by a poorly-understood mechanism involving upstream Open Reading Frames (uORFs). TCP-seq data of fil1 shows that stress allows scanning subunits to by-pass specific uORFs and reach the fil1 coding sequence. The integration of these observations with reporter assays revealed that fil1 translational control is mediated by a combination of scanning reinitiation-repressive and permissive uORFs, and establishes fil1 as a model for uORF-mediated translational control. Altogether, our transcriptome-wide study reveals general and gene-specific features of translation in a model eukaryote.
  4. Mol Cell Proteomics. 2022 Dec 06. pii: S1535-9476(22)00288-2. [Epub ahead of print] 100480
      Alternative open reading frames (AltORFs) are unannotated sequences in genome that encode novel peptides or proteins named alternative proteins (AltProts). Although ribosome profiling and bioinformatics predict a large number of AltProts, mass spectrometry (MS) as the only direct way of identification, is hampered by the short lengths and relative low abundance of AltProts. There is an urgent need for improvement of MS methodologies for AltProt identification. Here we report an approach based on size-exclusion chromatography (SEC) for simultaneous enrichment and fractionation of AltProts from complex proteome. This method greatly simplifies the variance of AltProts discovery by enriching small proteins smaller than 30 kDa. In a systematic comparison between ten methods, the approach we reported enabled discovery of more AltProts with overall higher intensities, with less cost of time and effort compared to other workflows. We applied this approach to identify 89 novel AltProts from mouse liver, 39 of which were differentially expressed between embryonic and adult mice. During embryonic development, the up-regulated AltProts were mainly involved in biological pathways on RNA splicing and processing, whereas the AltProts involved in metabolisms were more active in adult livers. Our study not only provides an effective approach for identifying AltProts, but also novel AltProts that are potentially important in developmental biology.
  5. Front Cell Neurosci. 2022 ;16 1019680
      Translation involves the biosynthesis of a protein sequence following the decoding of the genetic information embedded in a messenger RNA (mRNA). Typically, the eukaryotic mRNA was considered to be inherently monocistronic, but this paradigm is not in agreement with the translational landscape of cells, tissues, and organs. Recent ribosome sequencing (Ribo-seq) and proteomics studies show that, in addition to currently annotated reference proteins (RefProt), other proteins termed alternative proteins (AltProts), and microproteins are encoded in regions of mRNAs thought to be untranslated or in transcripts annotated as non-coding. This experimental evidence expands the repertoire of functional proteins within a cell and potentially provides important information on biological processes. This review explores the hitherto overlooked alternative proteome in neurobiology and considers the role of AltProts in pathological and healthy neuromolecular processes.
    Keywords:  mRNA; neurobiology; non-coding RNA (ncRNA); polycistronic; proteome