RESUMEN
The DNA polymerase zeta (Polζ) plays a critical role in bypassing DNA damage. REV3L, the catalytic subunit of Polζ, is also essential in mouse embryonic development and cell proliferation for reasons that remain incompletely understood. In this study, we reveal that REV3L protein interacts with heterochromatin components including repressive histone marks and localizes in pericentromeric regions through direct interaction with HP1 dimer. We demonstrate that Polζ/REV3L ensures progression of replication forks through difficult-to-replicate pericentromeric heterochromatin, thereby preventing spontaneous chromosome break formation. We also find that Rev3l-deficient cells are compromised in the repair of heterochromatin-associated double-stranded breaks, eliciting deletions in late-replicating regions. Lack of REV3L leads to further consequences that may be ascribed to heterochromatin replication and repair-associated functions of Polζ, with a disruption of the temporal replication program at specific loci. This is correlated with changes in epigenetic landscape and transcriptional control of developmentally regulated genes. These results reveal a new function of Polζ in preventing chromosome instability during replication of heterochromatic regions.
Asunto(s)
Replicación del ADN , Proteínas de Unión al ADN/genética , ADN Polimerasa Dirigida por ADN/genética , ADN/genética , Células Madre Embrionarias/metabolismo , Epigénesis Genética , Heterocromatina/metabolismo , Animales , Línea Celular , Línea Celular Transformada , Proliferación Celular , Homólogo de la Proteína Chromobox 5/genética , Homólogo de la Proteína Chromobox 5/metabolismo , Inestabilidad Cromosómica , ADN/metabolismo , Roturas del ADN de Doble Cadena , Proteínas de Unión al ADN/metabolismo , ADN Polimerasa Dirigida por ADN/metabolismo , Embrión de Mamíferos , Células Madre Embrionarias/citología , Fibroblastos/citología , Fibroblastos/metabolismo , Regulación del Desarrollo de la Expresión Génica , Células HeLa , Heterocromatina/química , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Células 3T3 NIH , Transducción de SeñalRESUMEN
Polo-like kinase 1 (PLK1) plays key roles during mitosis, prompting the development of PLK1 inhibitors for anticancer therapy. We recently determined that PLK1 is crucially required for entry into mitosis. Hence, we discuss the potential and limitations of PLK1 inhibition strategies to promote mitotic arrest and death of cancer cells.
RESUMEN
The regulation of proteolysis is an efficient way to control protein function in cells. Here, we present a general strategy enabling to increase the spatiotemporal resolution of conditional proteolysis by using light activation as trigger. Our approach relies on the auxin-inducible degradation system obtained by transposing components of the plant auxin-dependent degradation pathway in mammalian cells. We developed a photoactivatable auxin that acts as a photoactivatable inducer of degradation. Upon local and short light illumination, auxin is released in cells and triggers the degradation of a protein of interest with spatiotemporal control.