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RNA polymerase elongation along the gene body is tightly regulated to ensure proper transcription and alternative splicing events. Understanding the mechanism and factors critical in regulating the rate of RNA polymerase II elongation and processivity is clearly important. Recently we showed that PARP1, a well-known DNA repair protein, when bound to chromatin, regulates RNA polymerase II elongation. However, the mechanism by which it does so is not known. In the current study, we aimed to tease out how PARP1 regulates RNAPII elongation. We show, both in vivo and in vitro, that PARP1 binds directly to the Integrator subunit 3 (IntS3), a member of the elongation Integrator complex. The association between the two proteins is mediated via the C-terminal domain of PARP1 to the C-terminal domain of IntS3. Interestingly, the occupancy of IntS3 along two PARP1 target genes mimicked that of PARP1, suggesting a role in its recruitment/assembly of elongation factors. Indeed, the knockdown of PARP1 resulted in differential chromatin association and gene occupancy of IntS3 and other key elongation factors. Most of these PARP1-mediated effects were due to the physical presence of PARP1 rather than its PARylation activity. These studies argue that PARP1 controls the progressive RNAPII elongation complexes. In summary, we present a platform to begin to decipher PARP1′s role in recruiting/scaffolding elongation factors along the gene body regions during RNA polymerase II elongation and gene regulation.
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PARP1 Regulates Circular RNA Biogenesis though Control of Transcriptional Dynamics
Circular RNAs (circRNAs) are a recently discovered class of RNAs derived from protein-coding genes that have important biological and pathological roles. They are formed through backsplicing during co-transcriptional alternative splicing; however, the unified mechanism that accounts for backsplicing decisions remains unclear. Factors that regulate the transcriptional timing and spatial organization of pre-mRNA, including RNAPII kinetics, the availability of splicing factors, and features of gene architecture, have been shown to influence backsplicing decisions. Poly (ADP-ribose) polymerase I (PARP1) regulates alternative splicing through both its presence on chromatin as well as its PARylation activity. However, no studies have investigated PARP1’s possible role in regulating circRNA biogenesis. Here, we hypothesized that PARP1’s role in splicing extends to circRNA biogenesis. Our results identify many unique circRNAs in PARP1 depletion and PARylation-inhibited conditions compared to the wild type. We found that while all genes producing circRNAs share gene architecture features common to circRNA host genes, genes producing circRNAs in PARP1 knockdown conditions had longer upstream introns than downstream introns, whereas flanking introns in wild type host genes were symmetrical. Interestingly, we found that the behavior of PARP1 in regulating RNAPII pausing is distinct between these two classes of host genes. We conclude that the PARP1 pausing of RNAPII works within the context of gene architecture to regulate transcriptional kinetics, and therefore circRNA biogenesis. Furthermore, this regulation of PARP1 within host genes acts to fine tune their transcriptional output with implications in gene function.
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- Award ID(s):
- 2230470
- PAR ID:
- 10429142
- Date Published:
- Journal Name:
- Cells
- Volume:
- 12
- Issue:
- 8
- ISSN:
- 2073-4409
- Page Range / eLocation ID:
- 1160
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3390/cells12081160
