The role of p21 on Y polymerases organization and function

SABRINA MANSILLA, GASTON SORIA, JULIANA SPERONI, PAOLA CASSANO AND VANESA GOTTIFREDI

Ventura, California

Congreso; Gordon Conference: Mammalian DNA Repair; 2010

Gordon research Conferences

En esta conferencia de alto prestigio internacional la presentación de datos tiene como condición ineludible que dichas observaciones no hayan sido publicadas. Para proteger la confidencialidad de dichos datos la política de estos congresos es la de  no generar libros de resúmenes. El abstract presentado por nuestro grupo se detalla a continuación. The role of p21 on Y polymerases organization and function Sabrina Mansilla, Gaston Soria, Juliana Speroni, Paola Cassano and Vanesa Gottifredi Cell Cycle and Genomic Stability Laboratory, Fundación Instituto Leloir - CONICET, Universidad de Buenos Aires, Av. Patricias Argentinas 435 (C1405BWE), Buenos Aires, Argentina While many genotoxic signals provoke p53 dependent p21 upregulation, UV irradiation does not result in strong p21 induction. Moreover, many groups have reported increasing evidences that support the activation of p21 degradation signals after UV exposure. We have previously reported a direct correlation between p21 degradation and PCNA ubiquitination. This suggested that p21 degradation after UV irradiation could be linked to the upregulation of TLS (translesion DNA synthesis). Strengthening this notion, the utilization of non-degradable p21 mutants showed that stable p21 can impair PCNA ubiquitination, pol eta/PCNA interaction and pol eta focal organization. Here we show that the interaction between p21 and PCNA impairs the focal organization, not only of pol eta but also of other Y family members such as pol iota, pol kappa and Rev1. This was observed by transiently expressing stable p21 mutants or by comparing isogenic HCT116 p21-/- and p21+/+ cells. Interestingly as well, the percentage of cells with detectable pol eta, pol iota and pol kappa focal organization in unstressed conditions was higher when p21 was absent. This suggested that p21 could control the loading of Y polymerases to DNA in the absence of stress. We also evaluated the effect of p21 on the rate of DNA replication using DNA combing assays. We observed a clear reduction in the rates of DNA synthesis after UV irradiation when p21 was present. This suggests that p21 association with stalled replication forks at DNA lesions might interfere with the TLS events across those lesions. Moreover, this is in line with the excessive cell death of UV irradiated cells expressing stable p21 that we have reported in the past. Taking our results together we can propose that p21 might be in charge of blocking unnecessary recruitment of Y polymerases to DNA during unstressed replication. In turn, when TLS events are required after UV exposure, p21 degradation could promote gradual recruitment of TLS polymerases to DNA to prevent the irreversible stalling of replication forks.