Biology subjects

Black, J. C.

Publications and source records attributed to Black, J. C..

3 recordsLinked to original sources

bioRxiv2023

PKCδ regulates chromatin remodeling and DNA repair through SIRT6

Protein kinase C delta (PKC{delta}) is a ubiquitous kinase whose function is defined in part by localization to specific cellular compartments. Nuclear PKC{delta} is both necessary and sufficient for IR-induced apoptosis, while inhibition of PKC{delta} activity provides radioprotection in vivo. How nuclear PKC{delta} regulates DNA-damage induced cell death is poorly understood. Here we show that PKC{delta} regulates histone modification, chromatin accessibility, and double stranded break (DSB) repair through a mechanism that requires SIRT6. Overexpression of PKC{delta} promotes genomic instability and increases DNA damage and apoptosis. Conversely, depletion of PKC{delta} increases DNA repair via non-homologous end joining (NHEJ) and homologous recombination (HR) as evidenced by more rapid formation of NHEJ (DNA-PK) and HR (Rad51) DNA damage foci, increased expression of repair proteins, and increased repair of NHEJ and HR fluorescent reporter constructs. Nuclease sensitivity indicates that PKC{delta} depletion is associated with more open chromatin, while overexpression of PKC{delta} reduces chromatin accessibility. Epiproteome analysis revealed that PKC{delta} depletion increases chromatin associated H3K36me2, and reduces ribosylation of KDM2A and chromatin bound KDM2A. We identify SIRT6 as a downstream mediator of PKC{delta}. PKC{delta}-depleted cells have increased expression of SIRT6, and depletion of SIRT6 reverses the changes in chromatin accessibility, histone modification and NHEJ and HR DNA repair seen with PKC{delta}-depletion. Furthermore, depletion of SIRT6 reverses radioprotection in PKC{delta}-depleted cells. Our studies describe a novel pathway whereby PKC{delta} orchestrates SIRT6- dependent changes in chromatin accessibility to increase DNA repair, and define a mechanism for regulation of radiation-induced apoptosis by PKC{delta}. One Sentence SummaryProtein kinase C delta modifies chromatin structure via SIRT6 to regulate DNA repair.

molecular biology↗

bioRxiv2023

Transition from Transient DNA Rereplication to Inherited Gene Amplification Following Prolonged Environmental Stress

Cells require the ability to adapt to changing environmental conditions, however, it is unclear how these changes elicit stable permanent changes in genomes. We demonstrate that, in response to environmental metal exposure, the metallothionein (MT) locus undergoes DNA rereplication generating transient site-specific gene amplifications (TSSGs). Chronic metal exposure allows transition from MT TSSG to inherited MT gene amplification through homologous recombination within and outside of the MT locus. DNA rereplication of the MT locus is suppressed by H3K27me3 and EZH2. Long-term ablation of EZH2 activity eventually leads to integration and inheritance of MT gene amplifications without the selective pressure of metal exposure. The rereplication and inheritance of MT gene amplification is an evolutionarily conserved response to environmental metal from yeast to human. Our results describe a new paradigm for adaptation to environmental stress where targeted, transient DNA rereplication precedes stable inherited gene amplification.

cell biology↗

bioRxiv2021

SIX1 is a master regulator of the Rhabdomyosarcoma undifferentiated state

Rhabdomyosarcoma (RMS) is a pediatric skeletal muscle sarcoma characterized by the expression of the myogenic-lineage transcription factors (TF) MYOD1 and MYOG. Despite high expression of these TFs, RMS cells fail to terminally differentiate, suggesting the presence of factors that alter their function. Here, we demonstrate that the developmental TF, SIX1, is highly expressed in RMS and is critical to maintain a muscle progenitor-like state. SIX1 loss induces terminal differentiation of RMS cells into myotube-like cells and dramatically impedes tumor growth in vivo. We show that SIX1 maintains the RMS undifferentiated state by controlling enhancer activity and MYOD1 occupancy at loci more permissive to tumor growth over terminal muscle differentiation. Finally, we demonstrate that a gene signature derived from SIX1 loss correlates with differentiation status in RMS and predicts RMS progression in human disease. Our findings demonstrate a master regulatory role for SIX1 in the repression of RMS differentiation via genome-wide alterations in MYOD1-mediated transcription. HighlightsO_LISIX1 prevents differentiation in RMS while it promotes differentiation during normal development C_LIO_LIFN-RMS are highly dependent on SIX1 for growth in both zebrafish and mouse xenograft models C_LIO_LILoss of SIX1 alters the transcriptional landscape of RMS cells, inducing a growth to differentiation switch C_LIO_LISIX1 knockdown in FN-RMS causes reduced super enhancer-based activity at stem-related genes and enhanced MYOD1 binding to differentiation loci, resulting in the activation of a myogenic differentiation program C_LIO_LIA gene signature derived from SIX1 loss strongly correlates with myogenic differentiation status and is predictive of advanced RMS. C_LI

cancer biology↗