Phosphorylation at Ser473 regulates heterochromatin protein 1 binding and corepressor function of TIF1β/KAP1
Date Issued
2008
Date
2008
Author(s)
Chang, Chiung-Wen
Abstract
The transcriptional intermediary factor TIF1beta(TIF1beta)/KAP1/TRIM28) is an epigenetic regulator, functions in gene expression and chromatin remodeling at specific loci by association with members of the heterochromatin protein 1 (HP1) family and various other chromatin factors. The interaction between TIF1beta and HP1 depend on the HP1-box of TIF1beta. This functional interaction is crucial for chromatin dynamics, maintenance and gene regulation during differentiation, development, viral restriction, and cellular physiological regulation. However, the underlying mechanism of how the interaction is regulated remains poorly understood. TIF1beta is a multiply modificated proteins. The modifications include phosphorylation and sumoylation. These modifications play important role(s) in regulating its activity at different events. n the thesis part one, these studies conclude that the PKCdelta pathway mediated phosphorylation on TIF1beta/S473 is important for activation of G1-S cell cycle gene. The Ser473 of TIF1beta locates near the HP1b-binding motif, PXVXL. Phosphorylation of TIF1beta at Ser473 is cell cycle regulated which is elevated during S phase and M-phase. The dynamical alteration of which is functionally associated with cell cycle genes expression. Phosphorylation of TIF1beta/Ser473 coincides with the induction of cyclin A, cdc2, and cdc25A, due to the interaction with HP1beta is compromised by TIF1beta/phospho-S473 itself. Non-phsophorylated form TIF1beta/S473A preferentially associated with HP1beta in G1-phase nuclear matrix and affect cell cycle progression with more cell population stall in G2/M-phase while it is over-expressed. The phosphorylation of TIF1beta/Ser473 in early S phase is mediated by the PKCdelta pathway and is closely related to cell proliferation. n the thesis part two, these studies conclude that TIF1beta is functionally important for mitotic progression through its dynamically regulated hyperphosphorylation during mitosis. The abnormal cell morphology and apoptosis phenotype in TIF1beta knockdown HeLa cells is due to the failure to complete the mitotic phase. The failure in cleavage furrow determination, mitotic delay, gradually having unevenly distributed chromatin, and finally apoptosis, are revealed by FACs, immunostaining, and Time Lapse Cinematography. TIF1beta exhibits a hyperphosphorylation state only in M-phase. At least three phosphorylation sites, S757, S752, and S473 are identified by LC/MS/MS. The exclusion of TIF1beta from M-phase chromosome is due to its hyperphosphorylation. The re-orchestration of TIF1beta with reorganized chromatin is tightly correlated with its gradually decreased phosphorylation level during mitosis. The functional TIF1beta in M-phase may coincide with the mitotic failure observed in TIF1beta knockdown HeLa cells.
Subjects
TIF1beta/KAP1
heterochromatin protein 1(HP1)
phosphorylation
cell cycle
E2F
PKC
differentiation
mitosis
histone code
chromatin
cdk1
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