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RNA Biology Jan 2024Although circular RNAs (circRNAs) play important roles in regulating gene expression, the understanding of circRNAs in livestock animals is scarce due to the significant...
Although circular RNAs (circRNAs) play important roles in regulating gene expression, the understanding of circRNAs in livestock animals is scarce due to the significant challenge to characterize them from a biological sample. In this study, we assessed the outcomes of bovine circRNA identification using six enrichment approaches with the combination of ribosomal RNAs removal (); linear RNAs degradation (); linear RNAs and RNAs with structured 3' ends degradation (); ribosomal RNAs coupled with linear RNAs elimination (); ribosomal RNA, linear RNAs and RNAs with poly (A) tailing elimination (); and ribosomal RNA, linear RNAs and RNAs with structured 3' ends elimination (), respectively. RNA-sequencing analysis revealed that different approaches led to varied ratio of uniquely mapped reads, false-positive rate of identifying circRNAs, and the number of circRNAs per million clean reads ( <0.05). Out of 2,285 and 2,939 highly confident circRNAs identified in liver and rumen tissues, respectively, 308 and 260 were commonly identified from five methods, with Ribo-RTP method identified the highest number of circRNAs. Besides, 507 of 4,051 identified bovine highly confident circRNAs had shared splicing sites with human circRNAs. The findings from this work provide optimized methods to identify bovine circRNAs from cattle tissues for downstream research of their biological roles in cattle.
Topics: Cattle; RNA, Circular; Animals; RNA, Ribosomal; Sequence Analysis, RNA; Liver; Rumen; Computational Biology; Gene Expression Profiling; Humans
PubMed: 38797889
DOI: 10.1080/15476286.2024.2356334 -
Viruses May 2024Papillomavirus gene regulation is largely post-transcriptional due to overlapping open reading frames and the use of alternative polyadenylation and alternative splicing... (Review)
Review
Papillomavirus gene regulation is largely post-transcriptional due to overlapping open reading frames and the use of alternative polyadenylation and alternative splicing to produce the full suite of viral mRNAs. These processes are controlled by a wide range of cellular RNA binding proteins (RPBs), including constitutive splicing factors and cleavage and polyadenylation machinery, but also factors that regulate these processes, for example, SR and hnRNP proteins. Like cellular RNAs, papillomavirus RNAs have been shown to bind many such proteins. The life cycle of papillomaviruses is intimately linked to differentiation of the epithelial tissues the virus infects. For example, viral late mRNAs and proteins are expressed only in the most differentiated epithelial layers to avoid recognition by the host immune response. Papillomavirus genome replication is linked to the DNA damage response and viral chromatin conformation, processes which also link to RNA processing. Challenges with respect to elucidating how RBPs regulate the viral life cycle include consideration of the orchestrated spatial aspect of viral gene expression in an infected epithelium and the epigenetic nature of the viral episomal genome. This review discusses RBPs that control viral gene expression, and how the connectivity of various nuclear processes might contribute to viral mRNA production.
Topics: RNA-Binding Proteins; Humans; Gene Expression Regulation, Viral; RNA, Viral; Papillomaviridae; Virus Replication; Viral Proteins; Papillomavirus Infections; Genome, Viral; Host-Pathogen Interactions; RNA, Messenger
PubMed: 38793664
DOI: 10.3390/v16050783 -
Viruses Apr 2024The HIV-1 capsid (CA) protein forms the outer shell of the viral core that is released into the cytoplasm upon infection. CA binds various cellular proteins, including...
The HIV-1 capsid (CA) protein forms the outer shell of the viral core that is released into the cytoplasm upon infection. CA binds various cellular proteins, including CPSF6, that direct HIV-1 integration into speckle-associated domains in host chromatin. Upon HIV-1 infection, CPSF6 forms puncta in the nucleus. Here, we characterised these CPSF6 puncta further in HeLa cells, T-cells and macrophages and confirmed that integration and reverse transcription are not required for puncta formation. Indeed, we found that puncta formed very rapidly after infection, correlating with the time that CA entered the nucleus. In aphidicolin-treated HeLa cells and macrophages, puncta were detected for the length of the experiment, suggesting that puncta are only lost upon cell division. CA still co-localised with CPSF6 puncta at the latest time points, considerably after the peak of reverse transcription and integration. Intriguingly, the number of puncta induced in macrophages did not correlate with the MOI or the total number of nuclear speckles present in each cell, suggesting that CA/CPSF6 is only directed to a few nuclear speckles. Furthermore, we found that CPSF6 already co-localised with nuclear speckles in uninfected T-cells, suggesting that HIV-1 promotes a natural behaviour of CPSF6.
Topics: HIV-1; Humans; mRNA Cleavage and Polyadenylation Factors; T-Lymphocytes; HeLa Cells; Macrophages; Virus Integration; Cell Nucleus; Capsid Proteins; HIV Infections; Capsid
PubMed: 38793552
DOI: 10.3390/v16050670 -
NPJ Parkinson's Disease May 2024A biallelic (AAGGG) expansion in the poly(A) tail of an AluSx3 transposable element within the gene RFC1 is a frequent cause of cerebellar ataxia, neuropathy, vestibular...
A biallelic (AAGGG) expansion in the poly(A) tail of an AluSx3 transposable element within the gene RFC1 is a frequent cause of cerebellar ataxia, neuropathy, vestibular areflexia syndrome (CANVAS), and more recently, has been reported as a rare cause of Parkinson's disease (PD) in the Finnish population. Here, we investigate the prevalence of RFC1 (AAGGG) expansions in PD patients of non-Finnish European ancestry in 1609 individuals from the Parkinson's Progression Markers Initiative study. We identified four PD patients carrying the biallelic RFC1 (AAGGG) expansion and did not identify any carriers in controls.
PubMed: 38789445
DOI: 10.1038/s41531-024-00723-0 -
Cells May 2024Mammalian oocyte development depends on the temporally controlled translation of maternal transcripts, particularly in the coordination of meiotic and early embryonic...
Mammalian oocyte development depends on the temporally controlled translation of maternal transcripts, particularly in the coordination of meiotic and early embryonic development when transcription has ceased. The translation of mRNA is regulated by various RNA-binding proteins. We show that the absence of cytoplasmic polyadenylation element-binding protein 3 (CPEB3) negatively affects female reproductive fitness. CPEB3-depleted oocytes undergo meiosis normally but experience early embryonic arrest due to a disrupted transcriptome, leading to aberrant protein expression and the subsequent failure of embryonic transcription initiation. We found that CPEB3 stabilizes a subset of mRNAs with a significantly longer 3'UTR that is enriched in its distal region with cytoplasmic polyadenylation elements. Overall, our results suggest that CPEB3 is an important maternal factor that regulates the stability and translation of a subclass of mRNAs that are essential for the initiation of embryonic transcription and thus for embryonic development.
Topics: Oocytes; Animals; RNA-Binding Proteins; Female; Mice; Meiosis; RNA, Messenger; Embryonic Development; Gene Expression Regulation, Developmental; 3' Untranslated Regions; Polyadenylation; RNA Stability
PubMed: 38786074
DOI: 10.3390/cells13100850 -
Development (Cambridge, England) Jun 2024The RNA-binding protein cytoplasmic polyadenylation element binding 1 (CPEB1) plays a fundamental role in regulating mRNA translation in oocytes. However, the specifics...
The RNA-binding protein cytoplasmic polyadenylation element binding 1 (CPEB1) plays a fundamental role in regulating mRNA translation in oocytes. However, the specifics of how and which protein kinase cascades modulate CPEB1 activity are still controversial. Using genetic and pharmacological tools, and detailed time courses, we have re-evaluated the relationship between CPEB1 phosphorylation and translation activation during mouse oocyte maturation. We show that both the CDK1/MAPK and AURKA/PLK1 pathways converge on CPEB1 phosphorylation during prometaphase of meiosis I. Only inactivation of the CDK1/MAPK pathway disrupts translation, whereas inactivation of either pathway alone leads to CPEB1 stabilization. However, CPEB1 stabilization induced by inactivation of the AURKA/PLK1 pathway does not affect translation, indicating that destabilization and/or degradation is not linked to translational activation. The accumulation of endogenous CCNB1 protein closely recapitulates the translation data that use an exogenous template. These findings support the overarching hypothesis that the activation of translation during prometaphase in mouse oocytes relies on a CDK1/MAPK-dependent CPEB1 phosphorylation, and that translational activation precedes CPEB1 destabilization.
Topics: Animals; Oocytes; Meiosis; mRNA Cleavage and Polyadenylation Factors; Phosphorylation; Mice; Protein Biosynthesis; Female; CDC2 Protein Kinase; Transcription Factors; Aurora Kinase A; Cyclin B1; Cell Cycle Proteins; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Signal Transduction
PubMed: 38785133
DOI: 10.1242/dev.202712 -
Microbial Cell (Graz, Austria) 2024In , polyadenylated forms of mature (and not precursor) small non-coding RNAs (sncRNAs) those fail to undergo proper 3'-end maturation are subject to an active...
In , polyadenylated forms of mature (and not precursor) small non-coding RNAs (sncRNAs) those fail to undergo proper 3'-end maturation are subject to an active degradation by Rrp6p and Rrp47p, which does not require the involvement of core exosome and TRAMP components. In agreement with this finding, Rrp6p/Rrp47p is demonstrated to exist as an exosome-independent complex, which preferentially associates with mature polyadenylated forms of these sncRNAs. Consistent with this observation, a C-terminally truncated version of Rrp6p (Rrp6p-ΔC2) lacking physical association with the core nuclear exosome supports their decay just like its full-length version. Polyadenylation is catalyzed by both the canonical and non-canonical poly(A) polymerases, Pap1p and Trf4p. Analysis of the polyadenylation profiles in WT and -Δ strains revealed that the majority of the polyadenylation sites correspond to either one to three nucleotides upstream or downstream of their mature ends and their poly(A) tails ranges from 10-15 adenylate residues. Most interestingly, the accumulated polyadenylated snRNAs are functional in the -Δ strain and are assembled into spliceosomes. Thus, Rrp6p-Rrp47p defines a core nuclear exosome-independent novel RNA turnover system in baker's yeast targeting imperfectly processed polyadenylated sncRNAs that accumulate in the absence of Rrp6p.
PubMed: 38783922
DOI: 10.15698/mic2024.05.823 -
DIPAN: Detecting personalized intronic polyadenylation derived neoantigens from RNA sequencing data.Computational and Structural... Dec 2024Intronic polyadenylation (IPA) refers to a particular type of alternative polyadenylation where a gene makes use of a polyadenylation site located within its introns....
Intronic polyadenylation (IPA) refers to a particular type of alternative polyadenylation where a gene makes use of a polyadenylation site located within its introns. Aberrant IPA events have been observed in various types of cancer. IPA can produce noncoding transcripts or truncated protein-coding transcripts with altered coding sequences in the resulting protein product. Therefore, IPA events hold the potential to act as a reservoir of tumor neoantigens. Here, we developed a computational method termed DIPAN, which incorporates IPA detection, protein fragmentation, and MHC binding prediction to predict IPA-derived neoantigens. Utilizing RNA-seq from breast cancer cell lines and ovarian cancer clinical samples, we demonstrated the significant contribution of IPA events to the neoantigen repertoire. Through mass spectrometry immunopeptidome analysis, we further illustrated the processing and presentation of IPA-derived neoantigens on the surface of cancer cells. While most IPA-derived neoantigens are sample-specific, shared neoantigens were identified in both cancer cell lines and clinical samples. Furthermore, we demonstrated an association between IPA-derived neoantigen burden and overall survival in cancer patients.
PubMed: 38783901
DOI: 10.1016/j.csbj.2024.05.008 -
Journal of Hepatology May 2024Transcription termination fine-tunes gene expression and contributes to the specification of RNA function in eukaryotic cells. Transcription termination of HBV is...
BACKGROUND & AIMS
Transcription termination fine-tunes gene expression and contributes to the specification of RNA function in eukaryotic cells. Transcription termination of HBV is subject to the recognition of the canonical polyadenylation signal (cPAS) common to all viral transcripts. However, the regulation of this cPAS and its impact on viral gene expression and replication is currently unknown.
METHODS
To unravel the regulation of HBV transcript termination, we implemented a 3' RACE (rapid amplification of cDNA ends)-PCR assay coupled to single molecule sequencing both in in vitro-infected hepatocytes and in chronically infected patients.
RESULTS
The detection of a previously unidentified transcriptional readthrough indicated that the cPAS was not systematically recognized during HBV replication in vitro and in vivo. Gene expression downregulation experiments demonstrated a role for the RNA helicases DDX5 and DDX17 in promoting viral transcriptional readthrough, which was, in turn, associated with HBV RNA destabilization and decreased HBx protein expression. RNA and chromatin immunoprecipitation, together with mutation of the cPAS sequence, suggested a direct role of DDX5 and DDX17 in functionally linking cPAS recognition to transcriptional readthrough, HBV RNA stability and replication.
CONCLUSIONS
Our findings identify DDX5 and DDX17 as crucial determinants of HBV transcriptional fidelity and as host restriction factors for HBV replication.
IMPACT AND IMPLICATIONS
HBV covalently closed circular (ccc)DNA degradation or functional inactivation remains the holy grail for the achievement of HBV cure. Transcriptional fidelity is a cornerstone in the regulation of gene expression. Here, we demonstrate that two helicases, DDX5 and DDX17, inhibit recognition of the HBV polyadenylation signal and thereby transcriptional termination, thus decreasing HBV RNA stability and acting as restriction factors for efficient cccDNA transcription and viral replication. The observation that DDX5 and DDX17 are downregulated in patients chronically infected with HBV suggests a role for these helicases in HBV persistence in vivo. These results open new perspectives for researchers aiming at identifying new targets to neutralise cccDNA transcription.
PubMed: 38782119
DOI: 10.1016/j.jhep.2024.05.016 -
Cellular & Molecular Biology Letters May 2024Alternative splicing of pre-mRNAs is a fundamental step in RNA processing required for gene expression in most metazoans. Serine and arginine-rich proteins (SR proteins)... (Review)
Review
Alternative splicing of pre-mRNAs is a fundamental step in RNA processing required for gene expression in most metazoans. Serine and arginine-rich proteins (SR proteins) comprise a family of multifunctional proteins that contain an RNA recognition motif (RRM) and the ultra-conserved arginine/serine-rich (RS) domain, and play an important role in precise alternative splicing. Increasing research supports SR proteins as also functioning in other RNA-processing-related mechanisms, such as polyadenylation, degradation, and translation. In addition, SR proteins interact with N-methyladenosine (mA) regulators to modulate the methylation of ncRNA and mRNA. Dysregulation of SR proteins causes the disruption of cell differentiation and contributes to cancer progression. Here, we review the distinct biological characteristics of SR proteins and their known functional mechanisms during carcinogenesis. We also summarize the current inhibitors that directly target SR proteins and could ultimately turn SR proteins into actionable therapeutic targets in cancer therapy.
Topics: Humans; Neoplasms; Animals; Serine-Arginine Splicing Factors; Alternative Splicing; RNA-Binding Proteins
PubMed: 38778254
DOI: 10.1186/s11658-024-00594-6