Browse
Submit Data
Databases
API
Help

Dataset Information

29 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Dual RNA 3'-end processing of H2A.X messenger RNA maintains DNA damage repair throughout the cell cycle.

ABSTRACT: Phosphorylated H2A.X is a critical chromatin marker of DNA damage repair (DDR) in higher eukaryotes. However, H2A.X gene expression remains relatively uncharacterised. Replication-dependent (RD) histone genes generate poly(A)- mRNA encoding new histones to package DNA during replication. In contrast, replication-independent (RI) histone genes synthesise poly(A)+ mRNA throughout the cell cycle, translated into histone variants that confer specific epigenetic patterns on chromatin. Remarkably H2AFX, encoding H2A.X, is a hybrid histone gene, generating both poly(A)+ and poly(A)- mRNA isoforms. Here we report that the selective removal of either mRNA isoform reveals different effects in different cell types. In some cells, RD H2A.X poly(A)- mRNA generates sufficient histone for deposition onto DDR associated chromatin. In contrast, cells making predominantly poly(A)+ mRNA require this isoform for de novo H2A.X synthesis, required for efficient DDR. This highlights the importance of differential H2A.X mRNA 3'-end processing in the maintenance of effective DDR.

SUBMITTER: Griesbach E

PROVIDER: S-EPMC7807067 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Dual RNA 3’ end processing of H2A.X messenger RNA maintains DNA damage repair throughout the cell cycle

Project description:Replication-dependent (RD) histone mRNA are the only non-polyadenylated protein coding transcripts in humans. However, a subset of RD histone genes also expresses polyadenylated mRNA upon terminal differentiation to maintain histone proteins. We now show that even in cycling cells, RD histone genes express some poly(A) mRNA. The H2AFX gene, encoding the DNA damage signalling histone variant H2A.X, expresses both an S-phase-specific SL mRNA ending in the typical stem-loop (SL) structure and substantial amounts of an extended poly(A) mRNA. We show, by the selective removal of either mRNA isoform, that the S-phase-specific H2A.X SL mRNA can generate sufficient histone for deposition onto DNA damage responsive (DDR) chromatin in some cell types. In contrast, when cells have evolved to make predominantly H2A.X poly(A) mRNA they then require de novo H2A.X for efficient DDR. This highlights the importance of differential H2AFX mRNA 3’ end processing for the maintenance of effective DDR

2020-11-23 | GSE144323 | GEO

Dual RNA 3’ end processing of H2A.X messenger RNA maintains DNA damage repair throughout the cell cycle

Project description:Dual RNA 3’ end processing of H2A.X messenger RNA maintains DNA damage repair throughout the cell cycle

| PRJNA603372 | ENA

Post-transcriptional polyadenylation site cleavage maintains 3'-end processing upon DNA damage.

Project description:The recognition of polyadenylation signals (PAS) in eukaryotic pre-mRNAs is usually coupled to transcription termination, occurring while pre-mRNA is chromatin-bound. However, for some pre-mRNAs, this 3'-end processing occurs post-transcriptionally, i.e., through a co-transcriptional cleavage (CoTC) event downstream of the PAS, leading to chromatin release and subsequent PAS cleavage in the nucleoplasm. While DNA-damaging agents trigger the shutdown of co-transcriptional chromatin-associated 3'-end processing, specific compensatory mechanisms exist to ensure efficient 3'-end processing for certain pre-mRNAs, including those that encode proteins involved in the DNA damage response, such as the tumor suppressor p53. We show that cleavage at the p53 polyadenylation site occurs in part post-transcriptionally following a co-transcriptional cleavage event. Cells with an engineered deletion of the p53 CoTC site exhibit impaired p53 3'-end processing, decreased mRNA and protein levels of p53 and its transcriptional target p21, and altered cell cycle progression upon UV-induced DNA damage. Using a transcriptome-wide analysis of PAS cleavage, we identify additional pre-mRNAs whose PAS cleavage is maintained in response to UV irradiation and occurring post-transcriptionally. These findings indicate that CoTC-type cleavage of pre-mRNAs, followed by PAS cleavage in the nucleoplasm, allows certain pre-mRNAs to escape 3'-end processing inhibition in response to UV-induced DNA damage.

| S-EPMC10068322 | biostudies-literature

A dual reporter approach to quantify defects in messenger RNA processing.

Project description:Splicing and nuclear export are vital components of eukaryotic gene expression. Defects in splicing due to cis mutations are known to cause a number of human diseases. Here we present a dual reporter system that can be used to look at splicing or export deficiencies resulting from an insufficiency in components of the cotranscriptional machinery. The constructs use a bidirectional promoter to coexpress a test reporter and a control reporter. In the splicing construct, maximal expression of the test reporter is dependent on efficient splicing and splicing-related nuclear export, whereas the control reporter is an intronless complementary DNA expression cassette. The dual reporters allow a robust ratiometric output that is independent of cell number or transfection efficiency. Therefore, our construct is internally controlled and amenable to high-throughput analysis. As a counterscreen, we have a nonsplicing control construct in which neither reporter bears an intron. We demonstrate the sensitivity of our construct to defects in nuclear export by depleting UAP56 and NXF1, essential components of the cotranscriptional machinery.

| S-EPMC2760683 | biostudies-literature

Post-transcriptional polyadenylation site cleavage maintains 3'-end processing upon DNA damage

Project description:The recognition of polyadenylation signals (PAS) in eukaryotic pre-mRNAs is usually coupled to transcription termination, occurring while pre-mRNA is chromatin-bound. However, for some pre-mRNAs, this 3'-end processing occurs post-transcriptionally, i.e. through a co-transcriptional cleavage (CoTC) event downstream of the PAS, leading to chromatin release and subsequent PAS cleavage in the nucleoplasm. While DNA-damaging agents trigger shutdown of co-transcriptional chromatin-associated 3ʹ-end processing, specific compensatory mechanisms exist to ensure efficient 3'-end processing for certain pre-mRNAs, including those that encode proteins involved in the DNA damage response, such as the tumor suppressor p53. We show that cleavage at the p53 polyadenylation site occurs in part post-transcriptionally following a co-transcriptional cleavage event. Cells with an engineered deletion of the p53 CoTC site exhibit impaired p53 3'-end processing, decreased mRNA and protein levels of p53 and its transcriptional target p21, and altered cell cycle progression upon UV-induced DNA damage. Using a transcriptome-wide analysis of PAS cleavage, we identify additional pre-mRNAs whose PAS cleavage is maintained in response to UV irradiation and occurring post-transcriptionally. These findings indicate that CoTC-type cleavage of pre-mRNAs, followed by PAS cleavage in the nucleoplasm, allows specific certain pre-mRNAs to escape 3'-end processing inhibition in response to UV-induced DNA damage.

| S-SCDT-10_15252-EMBJ_2022112358 | biostudies-other

RNA helicase-dependent gene looping impacts messenger RNA processing.

Project description:DDX5 and DDX17 are DEAD-box RNA helicase paralogs which regulate several aspects of gene expression, especially transcription and splicing, through incompletely understood mechanisms. A transcriptome analysis of DDX5/DDX17-depleted human cells confirmed the large impact of these RNA helicases on splicing and revealed a widespread deregulation of 3' end processing. In silico analyses and experiments in cultured cells showed the binding and functional contribution of the genome organizing factor CTCF to chromatin sites at or near a subset of DDX5/DDX17-dependent exons that are characterized by a high GC content and a high density of RNA Polymerase II. We propose the existence of an RNA helicase-dependent relationship between CTCF and the dynamics of transcription across DNA and/or RNA structured regions, that contributes to the processing of internal and terminal exons. Moreover, local DDX5/DDX17-dependent chromatin loops spatially connect RNA helicase-regulated exons with their cognate promoter, and we provide the first direct evidence that de novo gene looping modifies alternative splicing and polyadenylation. Overall our findings uncover the impact of DDX5/DDX17-dependent chromatin folding on pre-messenger RNA processing.

| S-EPMC9458439 | biostudies-literature

Messenger RNA processing in Methanocaldococcus (Methanococcus) jannaschii.

Project description:Messenger RNA (mRNA) processing plays important roles in gene expression in all domains of life. A number of cases of mRNA cleavage have been documented in Archaea, but available data are fragmentary. We have examined RNAs present in Methanocaldococcus (Methanococcus) jannaschii for evidence of RNA processing upstream of protein-coding genes. Of 123 regions covered by the data, 31 were found to be processed, with 30 including a cleavage site 12-16 nucleotides upstream of the corresponding translation start site. Analyses with 3'-RACE (rapid amplification of cDNA ends) and 5'-RACE indicate that the processing is endonucleolytic. Analyses of the sequences surrounding the processing sites for functional sites, sequence motifs, or potential RNA secondary structure elements did not reveal any recurring features except for an AUG translation start codon and (in most cases) a ribosome binding site. These properties differ from those of all previously described mRNA processing systems. Our data suggest that the processing alters the representation of various genes in the RNA pool and therefore, may play a significant role in defining the balance of proteins in the cell.

| S-EPMC2743043 | biostudies-literature

Dual recognition of phosphoserine and phosphotyrosine in histone variant H2A.X by DNA damage response protein MCPH1.

Project description:Tyr142, the C-terminal amino acid of histone variant H2A.X is phosphorylated by WSTF (Williams-Beuren syndrome transcription factor), a component of the WICH complex (WSTF-ISWI chromatin-remodeling complex), under basal conditions in the cell. In response to DNA double-strand breaks (DSBs), H2A.X is instantaneously phosphorylated at Ser139 by the kinases ATM and ATR and is progressively dephosphorylated at Tyr142 by the Eya1 and Eya3 tyrosine phosphatases, resulting in a temporal switch from a postulated diphosphorylated (pSer139, pTyr142) to monophosphorylated (pSer139) H2A.X state. How mediator proteins interpret these two signals remains a question of fundamental interest. We provide structural, biochemical, and cellular evidence that Microcephalin (MCPH1), an early DNA damage response protein, can read both modifications via its tandem BRCA1 C-terminal (BRCT) domains, thereby emerging as a versatile sensor of H2A.X phosphorylation marks. We show that MCPH1 recruitment to sites of DNA damage is linked to both states of H2A.X.

| S-EPMC3437875 | biostudies-literature

Messenger RNA processing is altered in autosomal dominant leukodystrophy.

Project description:Adult-onset autosomal dominant leukodystrophy (ADLD) is a slowly progressive neurological disorder characterized by autonomic dysfunction, followed by cerebellar and pyramidal features. ADLD is caused by duplication of the lamin B1 gene (LMNB1), which leads to its increased expression. The molecular pathways involved in the disease are still poorly understood. Hence, we analyzed global gene expression in fibroblasts and whole blood of LMNB1 duplication carriers and used Gene Set Enrichment Analysis to explore their gene signatures. We found that LMNB1 duplication is associated with dysregulation of genes involved in the immune system, neuronal and skeletal development. Genes with an altered transcriptional profile clustered in specific genomic regions. Among the dysregulated genes, we further studied the role of RAVER2, which we found to be overexpressed at mRNA and protein level. RAVER2 encodes a putative trans regulator of the splicing repressor polypyrimidine tract binding protein (PTB) and is likely implicated in alternative splicing regulation. Functional studies demonstrated an abnormal splicing pattern of several PTB-target genes and of the myelin protein gene PLP1, previously demonstrated to be involved in ADLD. Mutant mice with different lamin B1 expression levels confirmed that Raver2 expression is dependent on lamin B1 in neural tissue and determines an altered splicing pattern of PTB-target genes and Plp1. Overall our results demonstrate that deregulation of lamin B1 expression induces modified splicing of several genes, likely driven by raver-2 overexpression, and suggest that an alteration of mRNA processing could be a pathogenic mechanism in ADLD.

| S-EPMC4406291 | biostudies-literature

Pre-messenger RNA processing factors in the Drosophila genome.

Project description: Not available

| S-EPMC2180228 | biostudies-literature