Chk2 is a DNA damage-responsive checkpoint kinase implicated in repair events, cell cycle arrest and apoptosis, activated by phosphorylation in an ATM-dependent manner. To better understand the role and regulation of this kinase, we analyzed both Chk2 phosphorylations sites and interacting proteins. We identified three novel serine residues (S19, S33 and S35) on Chk2 that became phosphorylated in vivo rapidly after ionizing radiation (IR), in an ATM- and Nbs1-dependent, but ATR-independent manner. These phosphorylations were induced by a dose of IR higher than 1Gy, and declined by 45-90 min concomitant with a rise in Chk2 autophosphorylation at T387. Mutation of S19, S33 and S35 to alanine impaired Chk2 homodimerization and activities, reduced Hdmx degradation and failed to suppress cell growth. These findings underline a critical role for S19, S33 and S35, in the ATM-dependent activation and modulation of Chk2 response in relation to the amount of double strand breaks. Furthermore, using yeast two hybrid assay, we identified the proteasome activator PSME3 as a novel Chk2 binding protein. We confirmed a DNA damage independent association between Chk2 and PSME3 in human cells by reciprocal co-immunoprecipitation experiments and we additionally found that over-expressed PSME3 induced the accumulation of ectopic Chk2 by increasing its stability, while either PSME3 overexpression or depletion had no effect on the phosphorylation and catalytic activity of Chk2. The functional role of this interaction is currently under investigation.
|Titolo:||Identification of three novel phosphoresidues and a new interactor for Chk2 protein kinase|
|Data di pubblicazione:||2007|
|Settore Scientifico Disciplinare:||Settore BIO/11 - Biologia Molecolare|
|Citazione:||Identification of three novel phosphoresidues and a new interactor for Chk2 protein kinase / L. Zannini, G. Buscemi, L. Carlessi, S. Lisanti, E. Fontanella, D. Delia. ((Intervento presentato al convegno Gordon Research Conference : Cancer Genetics and Epigenetics tenutosi a Lucca nel 2007.|
|Appare nelle tipologie:||14 - Intervento a convegno non pubblicato|