NDF/GLYR1 Promotes RNA Polymerase II Processivity via Pol II Binding and Nucleosome Destabilization
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ABSTRACT:
PROVIDER: PRJNA1248046 | ENA |
REPOSITORIES: ENA
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Project description:The Nucleosome Destabilizing Factor (NDF) facilitates transcription through chromatin, but its precise mechanism remains incompletely understood. Here, we identify a critical region (amino acids 140-160) within NDF that specifically interacts with phosphorylated RPB1, the largest subu-nit of elongating RNA Polymerase II (Pol II). Mutations in this region disrupt Pol II interaction and impair Pol II elongation both in vitro and in cells, yet do not affect NDF's ability to destabilize nu-cleosomes, establishing a functional separation between these two activities. Cellular studies re-veal that NDF knockout cells display faster Pol II elongation rates but produce fewer nascent tran-scripts, demonstrating NDF's primary role in maintaining transcriptional processivity through-out gene bodies. Our findings demonstrate that NDF uses distinct mechanisms to ensure productive transcription elongation rather than simply enhancing elongation speed, offering new insights in-to how transcription efficiency is maintained in chromatin.
2025-05-28 | GSE294088 | GEO
Project description:The Nucleosome Destabilizing Factor (NDF) facilitates transcription through chromatin, but its precise mechanism remains incompletely understood. Here, we identify a critical region (amino acids 140-160) within NDF that specifically interacts with phosphorylated RPB1, the largest subu-nit of elongating RNA Polymerase II (Pol II). Mutations in this region disrupt Pol II interaction and impair Pol II elongation both in vitro and in cells, yet do not affect NDF's ability to destabilize nu-cleosomes, establishing a functional separation between these two activities. Cellular studies re-veal that NDF knockout cells display faster Pol II elongation rates but produce fewer nascent tran-scripts, demonstrating NDF's primary role in maintaining transcriptional processivity through-out gene bodies. Our findings demonstrate that NDF uses distinct mechanisms to ensure productive transcription elongation rather than simply enhancing elongation speed, offering new insights in-to how transcription efficiency is maintained in chromatin.
2025-05-28 | GSE294087 | GEO
Project description:NDF/GLYR1 Promotes RNA Polymerase II Processivity via Pol II Binding and Nucleosome Destabilization
| PRJNA1248047 | ENA
Project description:Our understanding of transcription by RNA polymerase II (Pol II) is limited by our knowledge of the factors that mediate this critically important process. Here we describe the identification of NDF, a nucleosome destabilizing factor that facilitates Pol II transcription in chromatin. NDF has a PWWP motif, interacts with nucleosomes near the dyad, partially disassembles nucleosomes in an ATP-independent manner, and enhances transcription by Pol II through nucleosomes in vitro and in cell nuclei. NDF affects gene expression and co-localizes with H3K36me3 over transcribed regions of a subset of active genes with a preference for longer genes relative to shorter genes. Moreover, induction of transcription leads to the recruitment of NDF over gene bodies. In humans, NDF is present in all tested tissue types, is essential in stem cells, and is frequently overexpressed in breast cancer. Thus, NDF is a factor of primary importance for Pol II transcription in chromatin.
2019-01-25 | E-MTAB-6466 | biostudies-arrayexpress
Project description:Our understanding of transcription by RNA polymerase II (Pol II) is limited by our knowledge of the factors that mediate this critically important process. Here we describe the identification of NDF, a nucleosome destabilizing factor that facilitates Pol II transcription in chromatin. NDF has a PWWP motif, interacts with nucleosomes near the dyad, destabilizes nucleosomes in an ATP-independent manner, and facilitates transcription by Pol II through nucleosomes in a purified and defined transcription system as well as in cell nuclei. Upon transcriptional induction, NDF is recruited to the transcribed regions of thousands of genes and co-localizes with a subset of H3K36me3-enriched regions. Notably, the recruitment of NDF to gene bodies is accompanied by an increase in the transcript levels of many of the NDF-enriched genes. In addition, the global loss of NDF results in a decrease in the RNA levels of many genes. In humans, NDF is present at high levels in all tested tissue types, is essential in stem cells, and is frequently overexpressed in breast cancer. These findings indicate that NDF is a nucleosome destabilizing factor that is recruited to gene bodies during transcriptional activation and facilitates Pol II transcription through nucleosomes.
2018-04-16 | GSE109690 | GEO
Project description:The mechanisms governing the termination and subsequent reinitiation of RNA polymerase II (Pol II) remain poorly understood. Here we found that depletion of RPB7 leads to the destabilization of Pol II’s largest subunit, RPB1. This destabilization is influenced by the loop regions of RPB7, CDK9, the C-terminal domain (CTD) of RPB1, and its linker region. The stabilization process of RPB1 is regulated by the E3 ubiquitin ligase Cullin 3. Additionally, RPB7 interacts with the phosphatase CTDP1, which is crucial for maintaining RPB1 stability. RPB7 is also vital for the reinitiation of Pol II, engages with RNA processing factors, and is localized to the RNA exit channel of the Pol II complex. The absence of RPB7 compromises RNA processing. We propose that RPB7 recruits CTDP1 to dephosphorylate Pol II, enhancing its stability and facilitating efficient reinitiation, adding a new dimension to transcriptional regulation.
2025-02-05 | PXD048909 | Pride
Project description:Spt5 is a highly conserved RNA polymerase II (Pol II)-associated pausing and elongation factor. However, its impact on global elongation and Pol II processivity in mammalian cells has not been clarified. Here, we show that depleting Spt5 in mouse embryonic fibroblasts (MEFs) does not cause global elongation defects or decreased elongation rates. Instead, in the absence of Spt5, a fraction of Pol II molecules are dislodged during elongation thus decreasing the number of Pol II complexes that complete the transcription cycle. Most strikingly, this decrease is restricted to a narrow window between 15-20 kb from the promoter, a distance which coincides with the stage where accelerating Pol II attains maximum elongation speed and processivity. Consequently, long genes show a greater dependency on Spt5 for optimal elongation efficiency and overall gene expression than short genes. We propose that an important role of Spt5 in mammalian elongation is to promote the processivity of those Pol II complexes that are transitioning towards maximum elongation speed 15-20 kb from the promoter.
2018-03-06 | GSE106313 | GEO
Project description:RNA polymerase II (Pol II) elongation is a critical step in gene expression. Here we find that NDF, which was identified as a bilaterian nucleosome-destabilizing factor, is also a Pol II transcription factor that stimulates elongation with plain DNA templates in the absence of nucleosomes. NDF binds directly to Pol II and enhances elongation by a different mechanism than does transcription factor TFIIS. Moreover, yeast Pdp3, which is related to NDF, binds to Pol II and stimulates elongation. Thus, NDF is a Pol II-binding transcription elongation factor that is localized over gene bodies and is conserved from yeast to humans.
2022-03-10 | GSE185464 | GEO
Project description:The goal of this study was to determine the effect of Spt4 on transcription by Pol I in vivo. Therefore, Native Elongating Transcript Sequencing (NET-seq) was used to evaluate the occupancy of Pol I in wild-type and spt4△ strains. We determined that in spt4△ yeast, there was a processivity defect and Pol I accumulated at the 5' end of the 35S gene. Additionally, we observed that there was an enrichment for G-rich sequences just downstream of a high A content in the spt4△ strain, indicating that perhaps Spt4 plays a role in helping to propel Pol I through these regions of the rDNA. Altogether, this study suggests that Spt4 is an important transcription factor for Pol I by promoting polymerase processivity.
2021-02-18 | GSE166983 | GEO
Project description:Spt4 promotes Pol I processivity and transcription elongation
| PRJNA702517 | ENA