MUTATION OF ING1 TUMOR SUPRESSOR DETECTED IN HUMAN NEOPLASIA ABROGATES DNA REPAIR.

CERUTI, JULIETA; MARTÍN-GARRIDO, E; PALMERO, IGNACIO; CÁNEPA, EDUARDO

Iguazú, Argentina

Congreso; XL Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB); 2004

SAIB

CB-P39. MUTATION OF ING1 TUMOR SUPPRESSOR DETECTED IN HUMAN NEOPLASIA ABROGATES DNA REPAIRING1 TUMOR SUPPRESSOR DETECTED IN HUMAN NEOPLASIA ABROGATES DNA REPAIR Ceruti J1, Martín-Garrido E2, Palmero I2, Cánepa E1.1, Martín-Garrido E2, Palmero I2, Cánepa E1. 1Depto Química Biológica, FCEyN-UBA, Buenos Aires, Argentina and 2Instituto de Investigaciones Biomédicas, UAM, Madrid, España. E-mail: ecanepa@qb.fcen.uba.arDepto Química Biológica, FCEyN-UBA, Buenos Aires, Argentina and 2Instituto de Investigaciones Biomédicas, UAM, Madrid, España. E-mail: ecanepa@qb.fcen.uba.ar2Instituto de Investigaciones Biomédicas, UAM, Madrid, España. E-mail: ecanepa@qb.fcen.uba.ar The tumor suppressor ING1 inhibits cell growth in G1 phase by transactivating the CDK inhibitor p21waf1. Its biological functions have been studied in the last years and have been reported to mediate senescence, apoptosis and DNA repair. In this study, we investigated the involvement of p33ING1 (one of ING1 isoforms) in the modulation of DNA repair and the effect of point mutations on this p33 activity. We first examined whether p33ING1 responds to UVC in HeLa cell line. By northern blot, we found that UV light induces expression of p33 in a dose- and time-dependent manner. Western blots reveal a concomitant protein induction. To determine if p33 mediates DNA repair in HeLa cells, we used the host-cell-reactivation assay (HCR) where a UV-damaged plasmid is contransfected with p33 expression vector. Our data show that cells overexpressing p33 increase the rate of repair of the UVdamaged plasmid. To investigate whether ING1 mutation affects DNA repair, we assessed HCR assays using expression vectors codifying point mutations C215S or K270N. While C215S did not affect p33 ability to improve DNA repair, K270N reduced CAT activity. Similar results were obtained by means of unscheduled DNA synthesis assay, that evaluates the extent of repair of genomic DNA in UV irradiated cells. These results demonstrate that p33ING1 mutation, detected in human neoplasia, reduces the NER capacity, leading to an increased genomic instability, a condition that favors tumor progressionING1 mutation affects DNA repair, we assessed HCR assays using expression vectors codifying point mutations C215S or K270N. While C215S did not affect p33 ability to improve DNA repair, K270N reduced CAT activity. Similar results were obtained by means of unscheduled DNA synthesis assay, that evaluates the extent of repair of genomic DNA in UV irradiated cells. These results demonstrate that p33ING1 mutation, detected in human neoplasia, reduces the NER capacity, leading to an increased genomic instability, a condition that favors tumor progression