Comprehensive characterization of somatic variants associated with intronic polyadenylation in human cancers

Somatic single nucleotide variants (SNVs) in cancer genome affect gene expression through various mechanisms depending on their genomic location. While somatic SNVs near canonical splice sites have been reported to cause abnormal splicing of cancer-related genes, whether these SNVs can affect gene expression through other mechanisms remains an open question. Here, we analyzed RNA sequencing and exome data from 4,998 cancer patients covering ten cancer types and identified 152 somatic SNVs near splice sites that were associated with abnormal intronic polyadenylation (IPA). IPA-associated somatic variants favored the localization near the donor splice sites compared to the acceptor splice sites. A proportion of SNV-associated IPA events overlapped with premature cleavage and polyadenylation events triggered by U1 small nuclear ribonucleoproteins (snRNP) inhibition. GC content, intron length and polyadenylation signal were three genomic features that differentiated between SNV-associated IPA and intron retention. Notably, IPA-associated SNVs were enriched in tumor suppressor genes (TSGs), including the well-known TSGs such as PTEN and CDH1 with recurrent SNV-associated IPA events. Minigene assay confirmed that SNVs from PTEN, CDH1, VEGFA, GRHL2, CUL3 and WWC2 could lead to IPA. This work reveals that IPA acts as a novel mechanism explaining the functional consequence of somatic SNVs in human cancer.     

A model for random genetic damage directing selection of diploid or aneuploid tumours

Objectives: To test whether genetic instability may determine whether tumours become aneuploid or diploid. Materials and methods: We have identified genes needed for cell survival or replication by combining Affymetrix gene expression array data from 12 experimental cell lines with in silico GEO+GNF and expO databases. Specific loss of heterozygosis (LOHs), chromosomal abnormalities (called derivative chromosomes) and numbers of normal homologues were identified by SNP and SKY analyses. Random gene losses were calculated under the assumption that bi‐allelic MMR gene inactivation causes a 20‐fold increase in rate of gene loss. Results: There were ～1.23 × 10⁴ genes widely dispersed throughout the genome and possibly expressed by all cells for survival or proliferation, many of these genes performed housekeeping functions. Conservation of the genes may explain the complete haploid genomes found for 15 different cell types and derivative chromosomes selectively retained in aneuploid cancer cell lines after LOH formations, and normal homologue losses. Loss of cell survival/replication genes was calculated to be higher in colon stem cells of carriers of MMR gene mutations than carriers of APC gene mutations. Conclusion: Random loss of cell survival/replication genes was calculated to be low enough for colon stem cells with APC gene mutations to ‘select’ LOH and derivative chromosome combinations favouring tumour cell proliferation. However, cell survival/replication gene loss was calculated to be too high for colonic stem cells lacking MMR genes to survive chromosomal instability, explaining why MMR mutations only produce tumours with diploid chromosome cells.     

Human phospho‐signaling networks of SARS‐CoV‐2 infection are rewired by population genetic variants

SARS‐CoV‐2 infection hijacks signaling pathways and induces protein–protein interactions between human and viral proteins. Human genetic variation may impact SARS‐CoV‐2 infection and COVID‐19 pathology; however, the genetic variation in these signaling networks remains uncharacterized. Here, we studied human missense single nucleotide variants (SNVs) altering phosphorylation sites modulated by SARS‐CoV‐2 infection, using machine learning to identify amino acid substitutions altering kinase‐bound sequence motifs. We found 2,033 infrequent phosphorylation‐associated SNVs (pSNVs) that are enriched in sequence motif alterations, potentially reflecting the evolution of signaling networks regulating host defenses. Proteins with pSNVs are involved in viral life cycle and host responses, including RNA splicing, interferon response (TRIM28), and glucose homeostasis (TBC1D4) with potential associations with COVID‐19 comorbidities. pSNVs disrupt CDK and MAPK substrate motifs and replace these with motifs of Tank Binding Kinase 1 (TBK1) involved in innate immune responses, indicating consistent rewiring of signaling networks. Several pSNVs associate with severe COVID‐19 and hospitalization (STARD13, ARFGEF2). Our analysis highlights potential genetic factors contributing to inter‐individual variation of SARS‐CoV‐2 infection and COVID‐19 and suggests leads for mechanistic and translational studies.Subject Categories: Computational Biology, Microbiology, Virology & Host Pathogen Interaction, Post-translational Modifications & Proteolysis

An integrative proteogenomic study reveals that human phospho‐signaling networks responding to SARS‐CoV‐2 infection are enriched in genetic variants that modify kinase binding motifs, suggesting that genetic variation may impact infection and COVID‐19 pathology.     

The AMP-dependent kinase pathway is upregulated in BAP1 mutant uveal melanoma

Metastatic uveal melanoma (UM) responds poorly to targeted therapies and immune checkpoint inhibitors. Loss of BRCA1-associated protein 1 (BAP1) via inactivating mutations in the BAP1 gene is associated with UM progression. Thus, molecular alterations caused by BAP1 dysfunction may be novel therapeutic targets for metastatic UM. Here, we found that phosphorylation of AMP-dependent kinase (AMPK) was elevated in BAP1-altered (or mutant) compared to BAP1-unaltered (or wild-type [WT]) UM tumors. As a readout of AMPK pathway activation, phosphorylation of an AMPK downstream effector, acetyl-CoA-carboxylase (ACC), was also elevated. BAP1 re-expression in BAP1-null UM cell lines decreased phospho-AMPK (pAMPK) and phospho-ACC (pACC) levels. AMPK phosphorylation is mediated by calcium/calmodulin dependent protein kinase kinase 2 (CaMKK2) and potentially liver kinase B1 (LKB1) in BAP1 mutant UM cells. Knockdown of AMPKα1/2 reduced the viability of BAP1 mutant UM cells, indicating a survival function of AMPK in BAP1 mutant UM. Our data suggest that the AMPK pathway is an important mechanism mediating the survival of BAP1 mutant UM. Targeting the AMPK pathway may be a novel therapeutic strategy for metastatic UM.     

Comprehensive landscape and interference of clonal haematopoiesis mutations for liquid biopsy: A Chinese pan-cancer cohort

Tumour-derived DNA found in the plasma of cancer patients provides the probability to detect somatic mutations from circulating cell-free DNA (cfDNA) in plasma samples. However, clonal hematopoiesis (CH) mutations affect the accuracy of liquid biopsy for cancer diagnosis and treatment. Here, we integrated landscape of CH mutations in 11,725 pan-cancer patients of Chinese and explored effects of CH on liquid biopsies in real-world. We first identified 5933 CHs based on panel sequencing of matched DNA of white blood cell and cfDNA on 301 genes for 5100 patients, in which CH number of patients had positive correlation with their diagnosis age. We observed that canonical genes related to CH, including DNMT3A, TET2, ASXL1, TP53, ATM, CHEK2 and SF3B1, were dominant in the Chinese cohort and 13.29% of CH mutations only appeared in the Chinese cohort compared with the Western cohort. Analysis of CH gene distribution bias indicated that CH tended to appear in genes with functions of tyrosine kinase regulation, PI3K-Akt signalling and TP53 activity, suggesting unfavourable effects of CH mutations in cancer patients. We further confirmed effect of driver genes carried by CH on somatic mutations in liquid biopsy of cancer patients. Forty-eight actionable somatic mutations in 17 driver genes were considered CH genes in 92 patients (1.80%) of the Chinese cohort, implying potential impacts of CH on clinical decision-making. Taken together, this study exhibits strong evidence that gene mutations from CH interfere accuracy of liquid biopsies using cfDNA in cancer diagnosis and treatment in real-world.     

Genome‐wide characterization and expression analysis of genetic variants in sweet orange

Heterozyosity is an important feature of many plant genomes, and is related to heterosis. Sweet orange, a highly heterozygous species, is thought to have originated from an inter‐species hybrid between pummelo and mandarin. To investigate the heterozygosity of the sweet orange genome and examine how this heterozygosity affects gene expression, we characterized the genome of Valencia orange for single nucleotide variations (SNVs), small insertions and deletions (InDels) and structural variations (SVs), and determined their functional effects on protein‐coding genes and non‐coding sequences. Almost half of the genes containing large‐effect SNVs and InDels were expressed in a tissue‐specific manner. We identified 3542 large SVs (>50 bp), including deletions, insertions and inversions. Most of the 296 genes located in large‐deletion regions showed low expression levels. RNA‐Seq reads and DNA sequencing reads revealed that the alleles of 1062 genes were differentially expressed. In addition, we detected approximately 42 Mb of contigs that were not found in the reference genome of a haploid sweet orange by de novo assembly of unmapped reads, and annotated 134 protein‐coding genes within these contigs. We discuss how this heterozygosity affects the quality of genome assembly. This study advances our understanding of the genome architecture of sweet orange, and provides a global view of gene expression at heterozygous loci.     

Pseudogenization in pathogenic fungi with different host plants and lifestyles might reflect their evolutionary past

Pseudogenes are genes with significant homology to functional genes, but contain disruptive mutations (DMs) leading to the production of non‐ or partially functional proteins. Little is known about pseudogenization in pathogenic fungi with different lifestyles. Here, we report the identification of DMs causing pseudogenes in the genomes of the fungal plant pathogens Botrytis cinerea, Cladosporium fulvum, Dothistroma septosporum, Mycosphaerella fijiensis, Verticillium dahliae and Zymoseptoria tritici. In these fungi, we identified 1740 gene models containing 2795 DMs obtained by an alignment‐based gene prediction method. The contribution of sequencing errors to DMs was minimized by analyses of resequenced genomes to obtain a refined dataset of 924 gene models containing 1666 true DMs. The frequency of pseudogenes varied from 1% to 5% in the gene catalogues of these fungi, being the highest in the asexually reproducing fungus C. fulvum (4.9%), followed by D. septosporum (2.4%) and V. dahliae (2.1%). The majority of pseudogenes do not represent recent gene duplications, but members of multi‐gene families and unitary genes. In general, there was no bias for pseudogenization of specific genes in the six fungi. Single exceptions were those encoding secreted proteins, including proteases, which appeared more frequently pseudogenized in C. fulvum than in D. septosporum. Most pseudogenes present in these two phylogenetically closely related fungi are not shared, suggesting that they are related to adaptation to a different host (tomato versus pine) and lifestyle (biotroph versus hemibiotroph).     

Genomic analysis of oceanic cyanobacterial myoviruses compared with T4‐like myoviruses from diverse hosts and environments

T4‐like myoviruses are ubiquitous, and their genes are among the most abundant documented in ocean systems. Here we compare 26 T4‐like genomes, including 10 from non‐cyanobacterial myoviruses, and 16 from marine cyanobacterial myoviruses (cyanophages) isolated on diverse Prochlorococcus or Synechococcus hosts. A core genome of 38 virion construction and DNA replication genes was observed in all 26 genomes, with 32 and 25 additional genes shared among the non‐cyanophage and cyanophage subsets, respectively. These hierarchical cores are highly syntenic across the genomes, and sampled to saturation. The 25 cyanophage core genes include six previously described genes with putative functions (psbA, mazG, phoH, hsp20, hli03, cobS), a hypothetical protein with a potential phytanoyl‐CoA dioxygenase domain, two virion structural genes, and 16 hypothetical genes. Beyond previously described cyanophage‐encoded photosynthesis and phosphate stress genes, we observed core genes that may play a role in nitrogen metabolism during infection through modulation of 2‐oxoglutarate. Patterns among non‐core genes that may drive niche diversification revealed that phosphorus‐related gene content reflects source waters rather than host strain used for isolation, and that carbon metabolism genes appear associated with putative mobile elements. As well, phages isolated on Synechococcus had higher genome‐wide %G+C and often contained different gene subsets (e.g. petE, zwf, gnd, prnA, cpeT) than those isolated on Prochlorococcus. However, no clear diagnostic genes emerged to distinguish these phage groups, suggesting blurred boundaries possibly due to cross‐infection. Finally, genome‐wide comparisons of both diverse and closely related, co‐isolated genomes provide a locus‐to‐locus variability metric that will prove valuable for interpreting metagenomic data sets.     

Identification of castration‐resistant prostate cancer‐related hub genes using weighted gene co‐expression network analysis

Prostate cancer is the most common malignancy in urinary system and brings heavy burdens in men. We downloaded gene expression profile of mRNA and related clinical data of GSE70768 data set from public database. Weighted gene co‐expression network analysis (WGCNA) was used to identify the relationships between gene modules and clinical features, as well as the candidate genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses were developed to investigate the potential functions of related hub genes. Importantly, basic experiments were performed to verify the relationship between hub genes and the phenotype previously identified. Lastly, copy number variation (CNV) analysis was conducted to explore the genetical alteration. WGCNA identified that black module was the most relevant module which was tightly related to castration‐resistant prostate cancer (CRPC) phenotype. KEGG and GO analysis results revealed genes in black module were mainly related to RNA splicing. Additionally, 9 genes were chosen as hub genes and heterogeneous nuclear ribonucleoprotein A2/B1 (HNRNPA2B1), golgin A8 family member B (GOLGA8B) and mitogen‐activated protein kinase 8 interacting protein 3 (MAPK8IP3) were identified to be associated with PCa progression and prognosis. Moreover, all above three genes were highly expressed in CRPC‐like cells and their suppression led to hindered cell proliferation in vitro. Finally, CNV analysis found that amplification was the main type of alteration of the 3 hub genes. Our study found that HNRNPA2B1, GOLGA8B and MAPK8IP3 were identified to be tightly associated with tumour progression and prognosis, and further researches are needed before clinical application.     

High‐resolution array‐CGH in patients with oculocutaneous albinism identifies new deletions of the TYR,OCA2, and SLC45A2 genes and a complex rearrangement of the OCA2 gene

Oculocutaneous albinism (OCA) is caused by mutations in six different genes, and their molecular diagnosis encompasses the search for point mutations and intragenic rearrangements. Here, we used high‐resolution array‐comparative genome hybridization (CGH) to search for rearrangements across exons, introns and regulatory sequences of four OCA genes: TYR, OCA2, TYRP1, and SLC45A2. We identified a total of ten new deletions in TYR, OCA2, and SLC45A2. A complex rearrangement of OCA2 was found in two unrelated patients. Whole‐genome sequencing showed deletion of a 184‐kb fragment (identical to a deletion previously found in Polish patients), whereby a large portion of the deleted sequence was re‐inserted after severe reshuffling into intron 1 of OCA2. The high‐resolution array‐CGH presented here is a powerful tool to detect gene rearrangements. Finally, we review all known deletions of the OCA1–4 genes reported so far in the literature and show that deletions or duplications account for 5.6% of all mutations identified in the OCA1–4 genes.