Phosphorylation of hRad17 by ATR is Required for Cell Cycle Checkpoint Activation
Annual summary rept. 1 Apr 2003-31 Mar 2004
TEXAS UNIV HEALTH SCIENCE CENTER AT SANANTONIO
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Targeted protein phosphorylaytion is a key event that serves to transduce DNA damage induced cell signals from upstream sensors to downstream effectors. ATR kinase phosphorylates BRCA1 and Rad17 upon DNA damage. Furthermore, the phosphorylation of BRCA1 can also influence the phosphorylation of other substrates by ATR. As a first approach to evaluate the functional consequence of IR-induced site-specifically phosphorylated residues on BRCA1, we have established a BRCA1-dependent transcription based assay which assesses the BRCA1-dependent repression function of ZBRK1 in mammalian cell from a reporter template bearing ZBRK1 DNA-binding sites. Functional dissection of ZBRK1 led to identification and characterization of a novel BRCA1-dependent repression domain encompassing ZBRK1 zinc fingers 5-8 and the unique C-terminus. This C-terminal repression domain functions in a BRCA1-dependent, histone deactylase-dependent and promoter-specific. Significantly, we also found that the BRCA1-dependent transcriptional repression domain on ZBRK1 includes elements that modulate its sequence-specific DNA- binding activity. This study revealed an unanticipated dual function for the ZBRK1 zinc fingers in DNA-binding and BRCA1-dependent transcriptional repression. Our BRCA1- dependent ZBRK1 repression assay may now be exploited to evaluate the influence of IR- induced site-specific phosphorylation of BRCA1 on its sequence-specific co-repressor function.
- Medicine and Medical Research