The 14-subunit metazoan-specific Integrator contains an endonuclease that cleaves nascent RNA transcripts. Here, we identified a complex containing Integrator and protein phosphatase 2A core enzyme (PP2A-AC), termed INTAC. The 3.5-angstrom-resolution structure reveals that nine human Integrator subunits and PP2A-AC assemble into a cruciform-shaped central scaffold formed by the backbone and shoulder modules, with the phosphatase and endonuclease modules flanking the opposite sides. As a noncanonical PP2A holoenzyme, the INTAC complex dephosphorylates the carboxy-terminal repeat domain of RNA polymerase II at serine-2, -5, and -7 and thus regulates transcription. Our study extends the function of PP2A to transcriptional regulation and reveals how dual enzymatic activities—RNA cleavage and RNA polymerase II dephosphorylation—are structurally and functionally integrated into the INTAC complex.

UHRF1 (Ubiquitin-like, with PHD and RING finger domains 1) plays an important role in DNA CpG methylation, heterochromatin function and gene expression. Overexpression of UHRF1 has been suggested to contribute to tumorigenesis. However, regulation of UHRF1 is largely unknown. Here we show that the deubiquitylase USP7 interacts with UHRF1. Using interaction-defective and catalytic mutants of USP7 for complementation experiments, we demonstrate that both physical interaction and catalytic activity of USP7 are necessary for UHRF1 ubiquitylation and stability regulation. Mass spectrometry analysis identified phosphorylation of serine (S) 652 within the USP7-interacting domain of UHRF1, which was further confirmed by a UHRF1 S652 phosphor (S652ph)-specific antibody. Importantly, the S652ph antibody identifies phosphorylated UHRF1 in mitotic cells and consistently S652 can be phosphorylated by the M phase-specific kinase CDK1-cyclin B in vitro. UHRF1 S652 phosphorylation significantly reduces UHRF1 interaction with USP7 in vitro and in vivo, which is correlated with a decreased UHRF1 stability in the M phase of the cell cycle. In contrast, UHRF1 carrying the S652A mutation, which renders UHRF1 resistant to phosphorylation at S652, is more stable. Importantly, cells carrying the S652A mutant grow more slowly suggesting that maintaining an appropriate level of UHRF1 is important for cell proliferation regulation. Taken together, our findings uncovered a cell cycle-specific signaling event that relieves UHRF1 from its interaction with USP7, thus exposing UHRF1 to proteasome-mediated degradation. These findings identify a molecular mechanism by which cellular UHRF1 level is regulated, which may impact cell proliferation.deubiquitination | phosphorylation UHRF1 S652 E pigenetic regulation has emerged as an important mechanism that regulates many chromatin template-based processes, including transcription, DNA replication, and repair. An important component of epigenetic regulation is DNA CpG methylation, which is mediated by DNA methyltransferases such as DMNT1 and DNMT3a/b and an accessory factor DNMT3L (1, 2). Recent studies demonstrate that maintenance of DNA methylation patterns requires UHRF1 (Ubiquitin-like, with PHD and RING finger domains 1) (also called Np95 and ICBP90). UHRF1 binds hemimethylated CpG and recruits DNMT1 to ensure faithful propagation of the DNA methylation patterns through DNA replication (3, 4). UHRF1 is also localized to euchromatic regions where it regulates transcription possibly by impacting DNA methylation and histone modifications (5, 6). UHRF1 has been shown to regulate cell proliferation, and its loss has been implicated in the mis-regulation of both G1 and G2/M phases of the cell cycle, respectively (7). However, very little is known how this important epigenetic regulator itself is regulated. To address this question, we have recently undertaken a proteomics approach and identified a cell cycle signaling-regulated physical interaction of UHRF1 with the deubiquitylase USP7 (HAUSP) (8,9) ...

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Background Nuclei of eukaryotes contain various higher-order chromatin architectures and nuclear bodies (NBs), which are critical for proper nuclear functions. Recent studies showed that active chromatin regions are associated with nuclear speckles (NSs), a type of NBs involved in RNA processing. However, the functional roles of NSs in 3D genome organization remain unclear. Results Using mouse hepatocytes as the model, we knocked down SRRM2, a core protein component scaffolding NSs, and performed Hi-C experiments to examine genome-wide chromatin interactions. We found that Srrm2 depletion disrupted the NSs and changed the expression of 1282 genes. The intra-chromosomal interactions were decreased in type A (active) compartments and increased in type B (repressive) compartments. Furthermore, upon Srrm2 knockdown, the insulation of TADs was decreased specifically in active compartments, and the most significant reduction occurred in A1 sub-compartments. Interestingly, the change of intra-TAD chromatin interactions upon Srrm2 depletion was not associated with the alteration of gene expression. Conclusions We show that disruption of NSs by Srrm2 knockdown causes a global decrease in chromatin interactions in active compartments, indicating critical functions of NSs in the organization of the 3D genome. Electronic supplementary material The online version of this article (10.1186/s13072-019-0289-2) contains supplementary material, which is available to authorized users.