Biology subjects

Yates, N. A.

Publications and source records attributed to Yates, N. A..

3 recordsLinked to original sources

bioRxiv2024

TRIP12 governs DNA Polymerase β involvement in DNA damage response and repair

The multitude of DNA lesion types, and the nuclear dynamic context in which they occur, present a challenge for genome integrity maintenance as this requires the engagement of different DNA repair pathways. Specific repair controllers that facilitate DNA repair pathway crosstalk between double strand break (DSB) repair and base excision repair (BER), and regulate BER protein trafficking at lesion sites, have yet to be identified. We find that DNA polymerase {beta} (Pol{beta}), crucial for BER, is ubiquitylated in a BER complex-dependent manner by TRIP12, an E3 ligase that partners with UBR5 and restrains DSB repair signaling. Here we find that, TRIP12, but not UBR5, controls cellular levels and chromatin loading of Pol{beta}. Required for Pol{beta} foci formation, TRIP12 regulates Pol{beta} involvement after DNA damage. Notably, excessive TRIP12-mediated shuttling of Pol{beta} affects DSB formation and radiation sensitivity, underscoring its precedence for BER. We conclude that the herein discovered trafficking function at the nexus of DNA repair signaling pathways, towards Pol{beta}-directed BER, optimizes DNA repair pathway choice at complex lesion sites. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=165 SRC="FIGDIR/small/588474v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@70c5bdorg.highwire.dtl.DTLVardef@1c60dfborg.highwire.dtl.DTLVardef@1c00ef6org.highwire.dtl.DTLVardef@1677f2_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2022

Mechanism by which PF-3758309, a Pan Isoform Inhibitor of p21-Activated Kinases, Blocks Reactivation of HIV-1 Latency

The "block and lock" strategy is one approach that might elicit a sterilizing cure for HIV-1 infection. The "block" refers to a compounds ability to inhibit latent HIV-1 proviral transcription, while the "lock" refers to its capacity to induce permanent proviral silencing. We identified PF-3758309, a pan-isoform inhibitor of p21-activated kinases (PAKs), as a potent inhibitor of HIV-1 latency reversal (Vargas et al., Antimicrob Agents Chemother. 2019;63(2):e01744-18). The goal of this study was to define the mechanism(s) involved. We found that both 24ST1NLESG cells (a cell line model of HIV-1 latency) and purified CD4+ naive and central memory T cells express high levels of PAK2, and lower levels of PAK1 and PAK4. Knockdown of PAK1 or PAK2, but not PAK4, in 24ST1NLESG cells resulted in a modest, but statistically significant decrease in the magnitude of HIV-1 latency reversal. Overexpression of PAK1 significantly increased the magnitude of latency reversal. A phospho-protein array analysis revealed that PF-3758309 down-regulates the NF-{kappa}B signaling pathway, which provides the most likely mechanism by which PF-3758309 inhibits latency reversal. Finally, we used cellular thermal shift assays combined with liquid chromatography and mass spectrometry to ascertain whether PF-3758309 off-target binding contributed to its activity. In 24ST1NLESG cells and in peripheral blood mononuclear cells, PF-3758309 bound to mitogen-activated protein kinase 1 and protein kinase A; however, knockdown of either of these kinases did not impact HIV-1 latency reversal. Collectively, our study suggests that PAK1 and PAK2 play a key role in the maintenance of HIV-1 latency. IMPORTANCEThe persistence of latent, replication-competent HIV-1 proviruses in resting CD4+ T cells, and other cellular reservoirs, represents a major barrier to a cure. The "block and lock" strategy is one approach proposed to elicit a sterilizing cure for HIV-1 infection. In this study, we define the mechanism by which PF-3758309, a pan-isoform p21-activated kinase (PAK) inhibitor, blocks the reversal of HIV-1 latency. Our data show that PAK1 and PAK2 play a role in the maintenance of HIV-1 latency, and further suggest that PAK inhibitors, such as PF-3758309, could form part of "block and lock" therapeutic strategies.

microbiology↗

bioRxiv2018

The N-cadherin interactome in primary cardiomyocytes as defined by quantitative proximity proteomics.

The junctional complexes that couple cardiomyocytes must transmit the mechanical forces of contraction while maintaining adhesive homeostasis. The adherens junction (AJ) connects the actomyosin networks of neighboring cardiomyocytes and is required for proper heart function. Yet little is known about the molecular composition of the cardiomyocyte AJ or how it is organized to function under mechanical load. Here we define the architecture, dynamics and proteome of the cardiomyocyte AJ. Mouse neonatal cardiomyocytes assemble stable AJs along intercellular contacts with organizational and structural hallmarks similar to mature contacts. We combine quantitative mass spectrometry with proximity labeling to identify the N-cadherin (CDH2) interactome. We define over 350 proteins in this interactome, nearly 200 of which are unique to CDH2 and not part of the E-cadherin (CDH1) interactome. CDH2-specific interactors are comprised primarily of adaptor and adhesion proteins that promote junction specialization. Finally, we find evidence of dynamic interplay between AJ and Z-disc proteins. Together, our results provide novel insight into the cardiomyocyte AJ and provide a proteomic atlas for defining the molecular complexes that regulate cardiomyocyte intercellular adhesion.\n\nSummary StatementProximity proteomics reveals a specific and specialized N-cadherin (CDH2) interactome along the cell-cell contacts of primary cardiomyocytes.

cell biology↗