A polyadenylation factor subunit implicated in regulating oxidative stress responses in Arabidopsis thaliana
Ontology highlight
ABSTRACT:
ORGANISM(S): Arabidopsis thaliana
PROVIDER: GSE7211 | GEO | 2007/03/07
SECONDARY ACCESSION(S): PRJNA98305
REPOSITORIES: GEO
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Project description:The oxt6 mutant is an oxidative stress-tolerant Arabidopsis mutant that is deficient in a polyadenylation factor subunit. Expression analysis suggests that impaired poly(A) site choice is responsible for the stress-tolerant phenotype. We used microarrays to understand the link between the polyadenylation defect and stress tolerance. Experiment Overall Design: Arabidopsis plants (the wild-type, oxt6 mutant, and two independent oxt6 transformants each carrying a gene encoding just the smaller of the two At1g30460-encoded proteins) were seeded and grown in soil. Plants were harvested for RNA isolation after 30 days; for this, six samples derived from 10-15 plants were prepared for each line. Ten samples were selected for study â three wild-type, three oxt6, and two from each of the complemented lines. Analysis of independent complemented plants minimizes the possible effects that may arise due to T-DNA insertion.
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Project description:Alternative polyadenylation has been explored in multiple native and disease transitions. The prevailing hypothesis being that differentiated cells use longer 3’UTRs with more scope for regulation, whereas undifferentiated cells use shorter, less regulated 3’UTRs. Here we describe gene-expression and alternative polyadenylation of human primary myocytes over a time course differentiation. Contrary to expectation, only minor changes to 3’-end choice were detected. To reconcile this finding with published differentiation data in the immortalized C2C12 myocyte cell line, a systematic comparison was undertaken. Less than half the genes differentially expressed in the immortalized model were recapitulated in primary cells, and of these, important metabolic states were either absent, underrepresented or regulated in the opposite direction. A new bioinformatic approach, developed to quantitate the degree of alternative polyadenylation between unrelated experiments demonstrated that alternative polyadenylation was reduced by ~50% with less than 1/10 of the genes that underwent alternative polyadenylation in C2C12 differentiation showing alternative processing in primary muscle differentiation. A possible explanation for this difference was a less pronounced down regulation of the cleavage and polyadenylation factors in the differentiation primary cell. In sum, the data promote the use of primary human myocytes to model muscle biogenesis over immortalized models that may not fully recapitulate human muscle development.
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Project description:Alternative cleavage and polyadenylation (APA) is a mechanism generating multiple mRNA isoforms with different 3'UTRs and/or coding sequences from a single gene. Here we used 3' Region Extraction And Deep Sequencing (3'READS) to systematically map cleavage and polyadenylation sites (PAS) in Drosophila and analyzsed APA in the RpII215 C4 mutant strain, which harbors a mutant RNA polymerase II (RNAPII) with a slower elongation rate. We found that about two thirds of mRNA genes in fly undergo APA and cis element analysis indicates that fly PASs are associated with downstream UGUG elements to a lesser degree than mammalian PASs. The head tissue tends to use distal PASs compared to the body, leading to preferential expression of long 3'UTR isoforms and transcripts using distal terminal exons. Gene Ontology analysis correlates brain-specific APA events with neuronal functions, indicating their functional relevance. We also found that the distance between the proximal and distal PASs and intron locations might play important roles for brain APA. Analysis of the RpII215 C4 mutant strain, reveals that a 50% decrease in transcriptional elongation rate leads to shortened 3'UTRs in the fly body, but not in the head. This correlates with an upregulation of Ssu72 specifically in RpII215 C4 bodies, suggesting that the mechanism underlying those observations involves another layer of regulation by RNAPII termination factors. By contrast, more significant activation of intronic PASs was observed in the head compared to the body when RNAPII speed is slow. Together, our results reveal that in Drosophila RNAPII elongation impacts on PAS choice in a context-specific manner.
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