P21 impairs Pol eta recruitment to stalling sites after UV irradiation though its PCNA binding domain

SORIA G AND GOTTIFREDI V.

Ventura, California, USA

Conferencia; Gordon Research Conference_Mammalian DNA Repair; 2007

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. En la sección de texto completo adjunto la aceptación de mi participación por parte de los organizadores  y l a página de la lista de participantes donde se detalla mi presentación con formato de póster. P21 impairs Pol eta recruitment to stalling sites after UV irradiation though its PCNA binding domain Gaston Soria and Vanesa Gottifredi Fundación Instituto Leloir. CONICET. Av Patricias Argentinas 435. Buenos Aires. 1405. Argentina. While p21 up-regulation is required for the sustained inhibition of DNA synthesis after many genotoxic insults, UV irradiation triggers p21 proteolysis. It was previously proposed that p21 down-regulation during UV irradiation might promote PCNA dependent DNA repair. PCNA is a processivity factor for many DNA polymerases such as pol delta and it is required both for un-stressed DNA replication and for the re-synthesis step of NER. Another polymerase that interacts with PCNA is pol eta. Pol eta is central for translesion DNA synthesis after UV irradiation, an essential process that avoids replication blockage during S phase. We have recently shown that a 6 myc tag fused to p21 N-terminus inhibits its degradation after UV. This non-degradable p21 mutant inhibits PCNA ubiquitination, a modification of PCNA that promotes pol eta/PCNA interaction. Therefore, we explore the effect of stable p21 expression on its two potential DNA repair related processes: NER and TLS. We found that p21 does not block DNA synthesis in G1 arrested cells after UV irradiation therefore suggesting that high levels of p21 do not efficiently inhibit NER in vivo. On the contrary, single cell analysis of PCNA and pol eta foci formation after UV irradiation indicates that p21 interferes with pol eta recruitment to PCNA foci. Similar experiments performed with p21 mutants unables to bind PCNA and CDKs showed that the inhibition of pol eta foci formation by p21 depends on PCNA/p21 interaction. Moreover, stable expression of a p21 mutant that interferes with PCNA/pol eta interaction promotes the accumulation of p-gamma H2AX, a well established marker of DNA damage. Taken togheter, these data suggest that increased p21 proteolysis after UV irradiation might be relevant for efficient by-pass of lesions by translesion polymerases.