A regulatory transcription module of ZBRK1/KAP1 complex and its signaling network in regulating DNA damage-responsive genes expression.

摘要

In pursuit of new SUMOylation targets that regulate the activities of cell cycle progression and mediate cellular apoptosis, a proteomic screening that combined affinity chromatography and tandem mass spectrometry was launched to identify novel targets from SUMO-1 stably-expressed HEK293 cells. With a series of discreet screens and analyses, this effort yields twenty-three SUMOylation candidates, which are found to carry out distinct cellular functions. KAP1 was verified as a bona fide SUMO-1 substrate and a following literature research pointed KAP1 to a novel role in regulating the transcription of a cluster of cell cycle regulator genes and pro-apoptotic genes via its interaction with a transcriptional factor ZBRK1 that bound a 15-bp DNA sequence motif.;Subsequently, the lysines 554, 779 and 804 in KAP1 were identified as the major SUMOylation sites. Using two KAP1 mutants---one deficient in SUMOylation and the other mimics constitutive SUMOylation, the Dox-mediated induction of cell cycle regulator p21WAF1/CIP1 transcription is differentially regulated by KAP1 SUMOylation status. The SUMOylation-dependent modulation in the p21 transcription was achieved through changes in the lysine acetylation and methylation of histone 3 at the p21 promoter. Furthermore, the KAP1 SUMOylation level was transiently decreased upon Dox-exposure, and the introduction of constitutively SUMOylated KAP1 desensitized breast cancer MCF-7 cells to Dox-elicited cell death. Taken together, I provide a novel mechanistic basis underlying the Dox-induced de-repression of p21 transcription, and my results suggest that Dox-induced decrease in KAP1 SUMOylation is essential for Dox-induced p21 expression and subsequent cell growth inhibition in MCF-7 cells.;Further tracking the upstream signaling cascade that leads to the presumed Dox-induced KAP1 de-SUMOylation, my preliminary results indicated that SUMO-specific protease SENP1 and DNA damage-responsive protein kinase ATM might have served for this purpose. Lastly, a proposed future studies and the rationale underlying it was described herein.