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DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis

ABSTRACT: This SuperSeries is composed of the SubSeries listed below.

ORGANISM(S): Arabidopsis thaliana

PROVIDER: GSE203328 | GEO | 2022/10/09

REPOSITORIES: GEO

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DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis [RNA-seq]

Project description:Meiotic recombination is initiated from the SPORULATION 11 (SPO11)-triggered formation of double strand breaks (DSBs) that usually occur in open chromatin with active transcriptional features in many eukaryotes. However, gene transcription at meiotic DSB sites appears to be detrimental for repair, and the mechanisms for gene silencing at DSB sites are largely undefined, especially in plants. Here, we demonstrate that the largest DNA polymerase epsilon subunit POL2A interacts with SU(VAR)3-9 homologs SUVH2 and SUVH9. N-SIM (Structured Illumination Microscopy) observation shows that the colocalization of SUVH2 with meiotic DSB marker γ- H2AX is dependent on POL2A. RNA-seq of male meiocytes identifies that POL2A and SUVH2 jointly repress the expression of 865 genes, which have several known characteristics associated with meiotic DSB sites. Bisulfite-seq and small RNA-seq of male meiocytes support the idea that the silencing of these genes by POL2A and SUVH2/9 is likely independent of CHH methylation or 24-nt siRNA accumulation. Consequently, pol2a suvh2 suvh9 triple mutants have more severe defects in meiotic recombination and fertility compared with either pol2a or suvh2 suvh9. Taken together, our results not only identify a new epigenetic regulatory mechanism for gene silencing in male meiocytes, but also reveal new roles for DNA polymerase and SUVH2/9 beyond their classic functions in mitosis.

2022-10-08 | GSE203309 | GEO

DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis [miRNA-seq]

2022-10-09 | GSE203325 | GEO

DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis [BiSulfite-seq]

2022-10-09 | GSE203263 | GEO

DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis

Project description:DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis

| PRJNA839425 | ENA

DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis [miRNA-seq]

Project description:DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis [miRNA-seq]

| PRJNA839423 | ENA

DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis [RNA-seq]

Project description:DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis [RNA-seq]

| PRJNA839398 | ENA

DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis [BiSulfite-seq]

Project description:DNA polymerase epsilon interacts with SUVH2/9 to mediate meiotic DSB associated gene silencing beyond DNA methylation in Arabidopsis [BiSulfite-seq]

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MDM2 interacts with the C-terminus of the catalytic subunit of DNA polymerase epsilon.

Project description:MDM2 is induced by p53 in response to cellular insults such as DNA damage and can have effects upon the cell cycle that are independent or downstream of p53. We used a yeast two-hybrid screen to identify proteins that bind to MDM2 and which therefore might be involved in these effects. We found that MDM2 can bind to the C-terminus of the catalytic subunit of DNA polymerase epsilon (DNA pol epsilon), to a region that is known to be essential in yeast. In an in vitro system we confirmed that MDM2 could bind to the homologous regions of both mouse and human DNA pol epsilon and to full-length human DNA pol epsilon. DNA pol epsilon co-immunoprecipitated with MDM2 from transfected H1299 cells and also from a HeLa cell nuclear extract. We show here that the DNA pol epsilon-interacting domain of MDM2 is located between amino acids 50 and 166. Our studies provide evidence that MDM2 interacts with a region of DNA pol epsilon that plays a critical role in the function of DNA pol epsilon.

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The human Tim-Tipin complex interacts directly with DNA polymerase epsilon and stimulates its synthetic activity.

Project description:The Tim-Tipin complex plays an important role in the S phase checkpoint and replication fork stability in metazoans, but the molecular mechanism underlying its biological function is poorly understood. Here, we present evidence that the recombinant human Tim-Tipin complex (and Tim alone) markedly enhances the synthetic activity of DNA polymerase ε. In contrast, no significant effect on the synthetic ability of human DNA polymerase α and δ by Tim-Tipin was observed. Surface plasmon resonance measurements and co-immunoprecipitation experiments revealed that recombinant DNA polymerase ε directly interacts with either Tim or Tipin. In addition, the results of DNA band shift assays suggest that the Tim-Tipin complex (or Tim alone) is able to associate with DNA polymerase ε bound to a 40-/80-mer DNA ligand. Our results are discussed in view of the molecular dynamics at the human DNA replication fork.

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SAD-3, a Putative Helicase Required for Meiotic Silencing by Unpaired DNA, Interacts with Other Components of the Silencing Machinery.

Project description:In Neurospora crassa, genes lacking a pairing partner during meiosis are suppressed by a process known as meiotic silencing by unpaired DNA (MSUD). To identify novel MSUD components, we have developed a high-throughput reverse-genetic screen for use with the N. crassa knockout library. Here we describe the screening method and the characterization of a gene (sad-3) subsequently discovered. SAD-3 is a putative helicase required for MSUD and sexual spore production. It exists in a complex with other known MSUD proteins in the perinuclear region, a center for meiotic silencing activity. Orthologs of SAD-3 include Schizosaccharomyces pombe Hrr1, a helicase required for RNAi-induced heterochromatin formation. Both SAD-3 and Hrr1 interact with an RNA-directed RNA polymerase and an Argonaute, suggesting that certain aspects of silencing complex formation may be conserved between the two fungal species.

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