bims-micpro 2024-04-07 papers

J Mol Biol. 2024 Apr 03. pii: S0022-2836(24)00154-2. [Epub ahead of print] 168559

An inner mitochondrial membrane microprotein from the SLC35A4 upstream ORF regulates cellular metabolism.

Andrea L Rocha, Victor Pai, Guy Perkins, Tina Chang, Jiao Ma, Qian Chu, Joan M Vaughan, Jolene K Diedrich, Mark H Ellisman, Alan Saghatelian.

Upstream open reading frames (uORFs) are cis-acting elements that can dynamically regulate the translation of downstream ORFs by suppressing downstream translation under basal conditions and, in some cases, increasing translation under stress conditions. Computational and empirical methods have identified uORFs in the 5'-UTRs of approximately half of all mouse and human transcripts, making uORFs one the largest regulatory elements known. Because the prevailing dogma was that eukaryotic mRNAs produce a single functional protein, the peptides and small proteins, or microproteins, encoded by uORFs are under studied. We hypothesized that a uORF in the SLC35A4 mRNA is producing a functionalmicroprotein (SLC35A4-MP) because of its conserved amino acid sequence. Through a series of biochemical and cellular experiments, we find that the 103-amino acid SLC35A4-MP is a single-pass transmembrane inner mitochondrial membrane (IMM) microprotein. The IMM contains the protein machinery crucial for cellular respiration and ATP generation, and loss of function studies with SLC35A4-MP significantly diminish maximal cellular respiration, indicating a vital role for this microprotein in cellular metabolism. The findings add to the growing list of functional microproteins and, more generally, indicate that uORFs that encode conserved microproteins are an untapped reservoir of functional microproteins.

Keywords: cellular metabolism; inner mitochondrial membrane; microprotein; mitochondria; upstream open reading frame (uORF)

DOI: https://doi.org/10.1016/j.jmb.2024.168559

bioRxiv. 2024 Apr 01. pii: 2024.03.22.586333. [Epub ahead of print]

Upstream open reading frames may contain hundreds of novel human exons.

Hyun Joo Ji, Steven L Salzberg.

Several recent studies have presented evidence that the human gene catalogue should be expanded to include thousands of short open reading frames (ORFs) appearing upstream or downstream of existing protein-coding genes, each of which would comprise an additional bicistronic transcript in humans. Here we explore an alternative hypothesis that would explain the translational and evolutionary evidence for these upstream ORFs without the need to create novel genes or bicistronic transcripts. We examined 2,199 upstream ORFs that have been proposed as high-quality candidates for novel genes, to determine if they could instead represent protein-coding exons that can be added to existing genes. We checked for the conservation of these ORFs in four recently sequenced, high-quality human genomes, and found a large majority (87.8%) to be conserved in all four as expected. We then looked for splicing evidence that would connect each upstream ORF to the downstream protein-coding gene at the same locus, thus creating a novel splicing variant using the upstream ORF as its first exon. These protein coding exon candidates were further evaluated using protein structure predictions of the protein sequences that included the proposed new exons. We determined that 582 out of 2,199 upstream ORFs have strong evidence that they can form protein coding exons that are part of an existing gene, and that the resulting protein is predicted to have similar or better structural quality than the currently annotated isoform.

DOI: https://doi.org/10.1101/2024.03.22.586333

Cancer Immunol Res. 2024 Apr 04.

The neo-open reading frame peptides that comprise the tumor framome are a rich source of neoantigens for cancer immunotherapy.

Identification of immunogenic cancer neoantigens as targets for therapy is challenging. Here, we integrate cancer whole genome and long-read transcript sequencing to identify the collection of novel open reading frame peptides (NOPs) expressed in tumors, termed the framome. NOPs represent tumor-specific peptides that are different from wild-type proteins and may be strongly immunogenic. We describe an uncharacterized class of hidden NOPs, which derive from structural genomic variants involving an upstream protein coding gene driving expression and translation of non-coding regions of the genome downstream of a rearrangement breakpoint. NOPs represent a vast amount of possible neoantigens particularly in tumors with many (complex) structural genomic variants and a low number of missense mutations. We show that NOPs are immunogenic and epitopes derived from NOPs can bind to MHC class I molecules. Finally, we provide evidence for the presence of memory T-cells specific for hidden NOPs in lung cancer patient peripheral blood.

DOI: https://doi.org/10.1158/2326-6066.CIR-23-0158