bims-micpro 2024-10-13 papers

bims-micpro

Biomed News

on Discovery and characterization of microproteins

Issue of 2024–10–13
eight papers selected by
Thomas Farid Martínez, University of California, Irvine

The cryptic immunopeptidome in health and disease.
Multi-omics integration analysis reveals the role of N6-methyladenosine in lncRNA translation during glioma stem cell differentiation.
Uncovering the small proteome of Methanosarcina mazei using Ribo-seq and peptidomics under different nitrogen conditions.
Zika viruses encode 5' upstream open reading frames affecting infection of human brain cells.
Identification and characterization of a novel small viral peptide (VSP59) encoded by Bombyx mori cypovirus (BmCPV) that negatively regulates viral replication.
Microproteins in cancer: identification, biological functions, and clinical implications.
CEAM is a mitochondrial-localized, amyloid-like motif-containing microprotein expressed in human cardiomyocytes.
Peptidomics and machine-learning-based evaluation of ncRNA-derived micropeptides in breast cancer: Expression patterns and functional/therapeutic insights.

Trends Genet. 2024 Oct 09. pii: S0168-9525(24)00210-5. [Epub ahead of print]

The cryptic immunopeptidome in health and disease.

Eralda Kina, Jean-David Larouche, Pierre Thibault, Claude Perreault.

  Peptides presented by MHC proteins regulate all aspects of T cell biology. These MHC-associated peptides (MAPs) form what is known as the immunopeptidome and their comprehensive analysis has catalyzed the burgeoning field of immunopeptidomics. Advances in mass spectrometry (MS) and next-generation sequencing have facilitated significant breakthroughs in this area, some of which are highlighted in this article on the cryptic immunopeptidome. Here, 'cryptic' refers to peptides and proteins encoded by noncanonical open reading frames (ORFs). Cryptic MAPs derive mainly from short unstable proteins found in normal, infected, and neoplastic cells. Cryptic MAPs show minimal overlap with cryptic proteins found in whole-cell extracts. In many cancer types, most cancer-specific MAPs are cryptic.

Keywords:  MHC; T lymphocytes; autoimmunity; immunopeptidome; noncanonical translation; tumor antigens

DOI:  https://doi.org/10.1016/j.tig.2024.09.003
Brief Funct Genomics. 2024 Oct 08. pii: elae037. [Epub ahead of print]

Multi-omics integration analysis reveals the role of N6-methyladenosine in lncRNA translation during glioma stem cell differentiation.

Meng Zhang, Runqiu Cai, Jingjing Liu, Yulan Wang, Shan He, Quan Wang, Xiaofeng Song, Jing Wu, Jian Zhao.

  Glioblastoma is one of the most lethal brain diseases in humans. Although recent studies have shown reciprocal interactions between N6-methyladenosine (m6A) modifications and long noncoding RNAs (lncRNAs) in gliomagenesis and malignant progression, the mechanism of m6A-mediated lncRNA translational regulation in glioblastoma remains unclear. Herein, we profiled the transcriptomes, translatomes, and epitranscriptomics of glioma stem cells and differentiated glioma cells to investigate the role of m6A in lncRNA translation comprehensively. We found that lncRNAs with numerous m6A peaks exhibit reduced translation efficiency. Transcript-level expression analysis demonstrates an enrichment of m6A around short open reading frames (sORFs) of translatable lncRNA transcripts. Further comparison analysis of m6A modifications in different RNA regions indicates that m6A peaks downstream of sORFs inhibit lncRNA translation more than those upstream. Observations in glioma-associated lncRNAs H19, LINC00467, and GAS5 further confirm the negative effect of m6A methylation on lncRNA translation. Overall, these findings elucidate the dynamic profiles of the m6A methylome and enhance the understanding of the complexity of lncRNA translational regulation.

Keywords:  glioma; m6A methylation; noncoding RNA; sORF; translational regulation

DOI:  https://doi.org/10.1093/bfgp/elae037
Nat Commun. 2024 Oct 06. 15(1): 8659

Uncovering the small proteome of Methanosarcina mazei using Ribo-seq and peptidomics under different nitrogen conditions.

Muhammad Aammar Tufail, Britta Jordan, Lydia Hadjeras, Rick Gelhausen, Liam Cassidy, Tim Habenicht, Miriam Gutt, Lisa Hellwig, Rolf Backofen, Andreas Tholey, Cynthia M Sharma, Ruth A Schmitz.

The mesophilic methanogenic archaeal model organism Methanosarcina mazei strain Gö1 is crucial for climate and environmental research due to its ability to produce methane. Here, we establish a Ribo-seq protocol for M. mazei strain Gö1 under two growth conditions (nitrogen sufficiency and limitation). The translation of 93 previously annotated and 314 unannotated small ORFs, coding for proteins ≤ 70 amino acids, is predicted with high confidence based on Ribo-seq data. LC-MS analysis validates the translation for 62 annotated small ORFs and 26 unannotated small ORFs. Epitope tagging followed by immunoblotting analysis confirms the translation of 13 out of 16 selected unannotated small ORFs. A comprehensive differential transcription and translation analysis reveals that 29 of 314 unannotated small ORFs are differentially regulated in response to nitrogen availability at the transcriptional and 49 at the translational level. A high number of reported small RNAs are emerging as dual-function RNAs, including sRNA154, the central regulatory small RNA of nitrogen metabolism. Several unannotated small ORFs are conserved in Methanosarcina species and overproducing several (small ORF encoded) small proteins suggests key physiological functions. Overall, the comprehensive analysis opens an avenue to elucidate the function(s) of multitudinous small proteins and dual-function RNAs in M. mazei.

DOI: https://doi.org/10.1038/s41467-024-53008-8
Nat Commun. 2024 Oct 12. 15(1): 8822

Zika viruses encode 5' upstream open reading frames affecting infection of human brain cells.

Charlotte Lefèvre, Georgia M Cook, Adam M Dinan, Shiho Torii, Hazel Stewart, George Gibbons, Alex S Nicholson, Liliana Echavarría-Consuegra, Luke W Meredith, Valeria Lulla, Naomi McGovern, Julia C Kenyon, Ian Goodfellow, Janet E Deane, Stephen C Graham, András Lakatos, Louis Lambrechts, Ian Brierley, Nerea Irigoyen.

Zika virus (ZIKV), an emerging mosquito-borne flavivirus, is associated with congenital neurological complications. Here, we investigate potential pathological correlates of virus gene expression in representative ZIKV strains through RNA sequencing and ribosome profiling. In addition to the single long polyprotein found in all flaviviruses, we identify the translation of unrecognised upstream open reading frames (uORFs) in the genomic 5' region. In Asian/American strains, ribosomes translate uORF1 and uORF2, whereas in African strains, the two uORFs are fused into one (African uORF). We use reverse genetics to examine the impact on ZIKV fitness of different uORFs mutant viruses. We find that expression of the African uORF and the Asian/American uORF1 modulates virus growth and tropism in human cortical neurons and cerebral organoids, suggesting a potential role in neurotropism. Although the uORFs are expressed in mosquito cells, we do not see a measurable effect on transmission by the mosquito vector in vivo. The discovery of ZIKV uORFs sheds new light on the infection of the human brain cells by this virus and raises the question of their existence in other neurotropic flaviviruses.

DOI: https://doi.org/10.1038/s41467-024-53085-9
Microbiol Spectr. 2024 Oct 09. e0082624

Identification and characterization of a novel small viral peptide (VSP59) encoded by Bombyx mori cypovirus (BmCPV) that negatively regulates viral replication.

Manman Cao, Qunnan Qiu, Xing Zhang, Wenxue Zhang, Zeen Shen, Chang Ma, Min Zhu, Jun Pan, Xingyu Tong, Guangli Cao, Chengliang Gong, Xiaolong Hu.

  Bombyx mori cypovirus (BmCPV), a member of the Reoviridae family, is a well-established research model for double-stranded RNA (dsRNA) viruses with segmented genomes. Despite its small genome size, the coding potential of BmCPV remains largely unexplored. In this study, we identified a novel small open reading frame within the S10 dsRNA genome, encoding a small viral peptide (VSP59) with 59 amino acid residues. Functional characterization revealed that VSP59 acts as a negative regulator of viral replication. VSP59 predominantly localizes to the cytoplasm, where it interacts with prohibitin 2 (PHB2), an inner membrane mitophagy receptor. This interaction targets mitochondria and triggers caspase 3-dependent apoptosis. Transient expression of vsp59 in BmN cells suppressed viral replication, an effect that was reversed by silencing PHB2 expression. Moreover, recombinant BmCPV with a mutated vsp59 exhibited reduced replication. Our findings demonstrate that VSP59 interacts with PHB2 on mitochondria, inducing apoptosis and thereby diminishing viral replication. This study expands our understanding of the genetic information encoded by the BmCPV genome and highlights the role of novel small peptides in host-virus interactions.
IMPORTANCE: A novel small open reading frame (sORF) from the viral genome was identified and characterized. The sORF could encode a small viral peptide (VSP59) that targeted mitochondria and induced prohibitin 2-related apoptosis, further attenuating Bombyx mori cypovirus replication.

Keywords:  BmCPV; VSP59; apoptosis; sORF; viral replication

DOI:  https://doi.org/10.1128/spectrum.00826-24
Trends Genet. 2024 Oct 07. pii: S0168-9525(24)00211-7. [Epub ahead of print]

Microproteins in cancer: identification, biological functions, and clinical implications.

Damon A Hofman, John R Prensner, Sebastiaan van Heesch.

  Cancer continues to be a major global health challenge, accounting for 10 million deaths annually worldwide. Since the inception of genome-wide cancer sequencing studies 20 years ago, a core set of ~700 oncogenes and tumor suppressor genes has become the basis for cancer research. However, this research has been based largely on an understanding that the human genome encodes ~19 500 protein-coding genes. Complementing this genomic landscape, recent advances have described numerous microproteins which are now poised to redefine our understanding of oncogenic processes and open new avenues for therapeutic intervention. This review explores the emerging evidence for microprotein involvement in cancer mechanisms and discusses potential therapeutic applications, with an emphasis on highlighting recent advances in the field.

Keywords:  cancer biology; microproteins; proteomics; small open reading frames

DOI:  https://doi.org/10.1016/j.tig.2024.09.002
Biochem Biophys Res Commun. 2024 Oct 05. pii: S0006-291X(24)01273-7. [Epub ahead of print]734 150737

CEAM is a mitochondrial-localized, amyloid-like motif-containing microprotein expressed in human cardiomyocytes.

Ruobing Li, Ti Qin, Yabo Guo, Shan Zhang, Xiaogang Guo.

  Microproteins synthesized through non-canonical translation pathways are frequently found within mitochondria. However, the functional significance of these mitochondria-localized microproteins in energy-intensive organs such as the heart remains largely unexplored. In this study, we demonstrate that the long non-coding RNA CD63-AS1 encodes a mitochondrial microprotein. Notably, in ribosome profiling data of human hearts, there is a positive correlation between the expression of CD63-AS1 and genes associated with cardiomyopathy. We have termed this microprotein CEAM (CD63-AS1 encoded amyloid-like motif containing microprotein), reflecting its sequence characteristics. Our biochemical assays show that CEAM forms protease-resistant aggregates within mitochondria, whereas deletion of the amyloid-like motif transforms CEAM into a soluble cytosolic protein. Overexpression of CEAM triggers mitochondrial stress responses and adversely affect mitochondrial bioenergetics in cultured cardiomyocytes. In turn, the expression of CEAM is reciprocally inhibited by the activation of mitochondrial stresses induced by oligomycin. When expressed in mouse hearts via adeno-associated virus, CEAM impairs cardiac function. However, under conditions of pressure overload-induced cardiac hypertrophy, CEAM expression appears to offer a protective benefit and mitigates the expression of genes associated with cardiac remodeling, presumably through a mechanism that suppresses stress-induced translation reprogramming. Collectively, our study uncovers a hitherto unexplored amyloid-like microprotein expressed in the human cardiomyocytes, offering novel insights into myocardial hypertrophy pathophysiology.

Keywords:  CD63-AS1; Microprotein; Mitochondrial unfolded protein response; Pressure overload-induced cardiac hypertrophy; Transverse aortic constriction

DOI:  https://doi.org/10.1016/j.bbrc.2024.150737
Lab Invest. 2024 Oct 09. pii: S0023-6837(24)01828-2. [Epub ahead of print] 102150

Peptidomics and machine-learning-based evaluation of ncRNA-derived micropeptides in breast cancer: Expression patterns and functional/therapeutic insights.

Alexandre Luiz Korte de Azevedo, Talita Helen Bombardelli Gomig, Michel Batista, Jaqueline Carvalho de Oliveira, Iglenir João Cavalli, Daniela Fiori Gradia, Enilze Maria de Souza Fonseca Ribeiro.

  Breast cancer is a highly heterogeneous disease characterized by different subtypes arising from molecular alterations that give the disease different phenotypes, clinical behaviors, and prognostic. The ncRNA-derived micropeptides (MPs) represent a novel layer of complexity in cancer study once they can be biologically active and can present potential as biomarkers and also in therapeutics. However, few large-scale studies address the expression of these peptides at the peptidomics level or evaluate their functions and potential in peptide-based therapeutics for breast cancer. In this study, we propose deepening the landscape of ncRNA-derived MPs in breast cancer subtypes and advance the comprehension of the relevance of these molecules to the disease. Firstly, we constructed a 16,349 unique putative MP sequence dataset by integrating two previously published lists of predicted ncRNA-derived MPs. We evaluated its expression on high-throughput mass spectrometry data of breast tumor samples from different subtypes. Next, we applied several machine and deep learning tools, such as AntiCP 2.0, MULocDeep, PEPstrMOD, Peptipedia, and PreAIP, to predict its functions, cellular localization, tertiary structure, physicochemical features, and other properties related to therapeutics. We identified 58 peptides expressed on breast tissue, including 27 differentially expressed MPs in tumor compared to non-tumor samples and MPs exhibiting tumor or subtype specificity. These peptides presented physicochemical features compatible with the canonical proteome and were predicted to influence the tumor immune environment and participate in cell communication, metabolism, and signaling processes. Also, some MPs presented potential as anti-cancer, anti-inflammatory, and anti-angiogenic molecules. Our data demonstrate that MPs derived from ncRNAs have expression patterns associated with specific breast cancer subtypes and tumor specificity, thus highlighting their potential as biomarkers for molecular classification. We also reinforce the relevance of MPs as biologically active molecules that play a role in breast tumorigenesis, besides their potential in peptide-based therapeutics.

Keywords:  Micropeptides; machine-learning; non-coding RNAs; subtypes; tumorigenesis

DOI:  https://doi.org/10.1016/j.labinv.2024.102150