OTUD5 promotes end-joining of deprotected telomeres by promoting ATM-dependent phosphorylation of KAP1.

Appropriate repair of damaged DNA and the suppression of DNA damage responses at telomeres are essential to preserve genome stability. DNA damage response (DDR) signaling consists of cascades of kinase-driven phosphorylation events, fine-tuned by proteolytic and regulatory ubiquitination. It is not fully understood how crosstalk between these two major classes of post-translational modifications impact DNA repair at deprotected telomeres.

Resolving the subtle details of human DNA alkyltransferase lesion search and repair mechanism by single-molecule studies.

SignificanceWe directly visualize DNA translocation and lesion recognition by the O-alkylguanine DNA alkyltransferase (AGT). Our data show bidirectional movement of AGT monomers and clusters on undamaged DNA that depended on Zn occupancy of AGT. A role of cooperative AGT clusters in enhancing lesion search efficiencies by AGT has previously been proposed.

Shelterin and subtelomeric DNA sequences control nucleosome maintenance and genome stability.

Telomeres and the shelterin complex cap and protect the ends of chromosomes. Telomeres are flanked by the subtelomeric sequences that have also been implicated in telomere regulation, although their role is not well defined. Here, we show that, in , the telomere-associated sequences (TAS) present on most subtelomeres are hyper-recombinogenic, have metastable nucleosomes, and unusual low levels of H3K9 methylation.

H4K20me2 distinguishes pre-replicative from post-replicative chromatin to appropriately direct DNA repair pathway choice by 53BP1-RIF1-MAD2L2.

The main pathways for the repair of DNA double strand breaks (DSBs) are non-homologous end-joining (NHEJ) and homologous recombination directed repair (HDR). These operate mutually exclusive and are activated by 53BP1 and BRCA1, respectively. As HDR can only succeed in the presence of an intact copy of replicated DNA, cells employ several mechanisms to inactivate HDR in the G1 phase of cell cycle.

Ubiquitin ligase trapping identifies an SCF(Saf1) pathway targeting unprocessed vacuolar/lysosomal proteins.

We have developed a technique, called Ubiquitin Ligase Substrate Trapping, for the isolation of ubiquitinated substrates in complex with their ubiquitin ligase (E3). By fusing a ubiquitin-associated (UBA) domain to an E3 ligase, we were able to selectively purify the polyubiquitinated forms of E3 substrates. Using ligase traps of eight different F box proteins (SCF specificity factors) coupled with mass spectrometry, we identified known, as well as previously unreported, substrates.

The influence of catalysis on mad2 activation dynamics.

Mad2 is a key component of the spindle assembly checkpoint, a safety device ensuring faithful sister chromatid separation in mitosis. The target of Mad2 is Cdc20, an activator of the anaphase-promoting complex/cyclosome (APC/C). Mad2 binding to Cdc20 is a complex reaction that entails the conformational conversion of Mad2 from an open (O-Mad2) to a closed (C-Mad2) conformer.

Physical and functional characterization of the genetic locus of IBtk, an inhibitor of Bruton's tyrosine kinase: evidence for three protein isoforms of IBtk.

Bruton's tyrosine kinase (Btk) is required for B-cell development. Btk deficiency causes X-linked agammaglobulinemia (XLA) in humans and X-linked immunodeficiency (Xid) in mice. Btk lacks a negative regulatory domain and may rely on cytoplasmic proteins to regulate its activity.

In vitro FRAP identifies the minimal requirements for Mad2 kinetochore dynamics.

Background: Mad1 and Mad2 are constituents of the spindle-assembly checkpoint, a device coupling the loss of sister-chromatid cohesion at anaphase to the completion of microtubule attachment of the sister chromatids at metaphase. Fluorescence recovery after photobleaching (FRAP) revealed that the interaction of cytosolic Mad2 with kinetochores is highly dynamic, suggesting a mechanism of catalytic activation of Mad2 at kinetochores followed by its release in a complex with Cdc20. The recruitment of cytosolic Mad2 to kinetochores has been attributed to a stable receptor composed of a distinct pool of Mad2 tightly bound to Mad1.