53BP1 regulates heterochromatin through liquid phase separation.

Human 53BP1 is primarily known as a key player in regulating DNA double strand break (DSB) repair choice; however, its involvement in other biological process is less well understood. Here, we report a previously uncharacterized function of 53BP1 at heterochromatin, where it undergoes liquid-liquid phase separation (LLPS) with the heterochromatin protein HP1α in a mutually dependent manner. Deletion of 53BP1 results in a reduction in heterochromatin centers and the de-repression of heterochromatic tandem repetitive DNA. We identify domains and residues of 53BP1 required for its LLPS, which overlap with, but are distinct from, those involved in DSB repair. Further, 53BP1 mutants deficient in DSB repair, but proficient in LLPS, rescue heterochromatin de-repression and protect cells from stress-induced DNA damage and senescence. Our study suggests that in addition to DSB repair modulation, 53BP1 contributes to the maintenance of heterochromatin integrity and genome stability through LLPS.

The 'New (Nu)-clear' evidence for the tumor-driving role of PI3K.

The classical phosphatidylinositol 3-kinases (PI3Ks) are heterodimers of p110 and p85. PIK3CA, the gene encoding the catalytic p110α subunit, is one of the most frequently mutated oncogenes in human cancers with hot spot mutations occurring in the helical domain or in the kinase domain. Tumors with these two types of PIK3CA mutations show overlapping yet distinct phenotypes; however, the underlying mechanisms remain unclear. In a recent publication [1], Hao et al revealed exciting findings about the PI3K p85ß regulatory subunit in promoting PIK3CA helical domain mutation-driven cancer progression. The authors found that p85ß disassociated from the PI3K complex and translocated into the nucleus only in cancer cells harboring PIK3CA helical domain mutations. Disrupting nuclear localization of p85ß suppressed mouse tumor growth of cancer cells with PIK3CA helical domain mutation. Mechanistically, they elegantly showed that nuclear p85ß recruited the deubiquitinase USP7 to stabilize the histone methyltransferases EZH1/2, leading to enhanced H3K27 trimethylation and gene transcription. Combining an EZH inhibitor with a PI3K inhibitor specifically resulted in regression of mouse xenograft tumors with PIK3CA helical domain mutations. These findings illustrate a previously uncharacterized function of p85ß in tumor development and suggest an effective approach to target tumors with PIK3CA helical mutations.

Molecular chaperone Hsp90 regulates REV1-mediated mutagenesis.

REV1 is a Y-family polymerase that plays a central role in mutagenic translesion DNA synthesis (TLS), contributing to tumor initiation and progression. In a current model, a monoubiquitinated form of the replication accessory protein, proliferating cell nuclear antigen (PCNA), serves as a platform to recruit REV1 to damaged sites on the DNA template. Emerging evidence indicates that posttranslational mechanisms regulate REV1 in yeast; however, the regulation of REV1 in higher eukaryotes is poorly understood. Here we show that the molecular chaperone Hsp90 is a critical regulator of REV1 in human cells. Hsp90 specifically binds REV1 in vivo and in vitro. Treatment with a specific inhibitor of Hsp90 reduces REV1 protein levels in several cell types through proteasomal degradation. This is associated with suppression of UV-induced mutagenesis. Furthermore, Hsp90 inhibition disrupts the interaction between REV1 and monoubiquitinated PCNA and suppresses UV-induced focus formation. These results indicate that Hsp90 promotes folding of REV1 into a stable and/or functional form(s) to bind to monoubiquitinated PCNA. The present findings reveal a novel role of Hsp90 in the regulation of TLS-mediated mutagenesis.

The molecular chaperone Hsp90 regulates accumulation of DNA polymerase eta at replication stalling sites in UV-irradiated cells.

DNA polymerase eta (Pol eta) is a member of the mammalian Y family polymerases and performs error-free translesion synthesis across UV-damaged DNA. For this function, Pol eta accumulates in nuclear foci at replication stalling sites via its interaction with monoubiquitinated PCNA. However, little is known about the posttranslational control mechanisms of Pol eta, which regulate its accumulation in replication foci. Here, we report that the molecular chaperone Hsp90 promotes UV irradiation-induced nuclear focus formation of Pol eta through control of its stability and binding to monoubiquitinated PCNA. Our data indicate that Hsp90 facilitates the folding of Pol eta into an active form in which PCNA- and ubiquitin-binding regions are functional. Furthermore, Hsp90 inhibition potentiates UV-induced cytotoxicity and mutagenesis in a Pol eta-dependent manner. Our studies identify Hsp90 as an essential regulator of Pol eta-mediated translesion synthesis.