The viral factor E7 plays a key role in the well‐established association between “high‐risk” Human Papillomavirus (HPV) infection and the development of epithelial malignant tumors, as uterine cervix and ano‐genital cancer. To delve into the molecular mechanisms of HPV‐mediated cell transformation, we searched for novel potential cellular targets of the HPV‐16 E7 oncoprotein, by means of the yeast two‐hybrid technique, identifying a protein–protein interaction between HPV‐16 E7 and the pro‐apoptotic cellular factor Siva‐1. Using co‐precipitation assays and the “PepSets” technique, we confirmed this physical interaction and mapped accurately, for both proteins, the amino acid residues involved. Additionally, we found that HPV‐16 E7 competed in vitro with the binding of the Bcl‐X_L anti‐apoptotic factor to Siva‐1, an interaction that has a major inference in UV radiation‐induced apoptosis. In HaCaT immortalized human keratinocytes, forced HPV‐16 E7 expression by retroviral infection caused Siva‐1 transcript up‐regulation, detected by cDNA macroarray hybridization and real‐time quantitative PCR, paralleled by an increased amount of protein. Confirming the anti‐apoptotic role of HPV‐16 E7 in the HaCaT cellular model, evaluated by nuclear morphology, we also found that Siva‐1 expression produced a significant increase of the apoptotic rate in UV radiation‐exposed HaCaT cells, and that this effect resulted explicitly counteracted by HPV‐16 E7. Being apoptosis a key physiological process for the elimination of irreversibly injured cells, the anti‐apoptotic role of HPV‐16 E7, performed at least by its interference with Siva‐1, can be considered an additional mechanism for the survival of damaged, potentially transforming, cell clones. J. Cell. Physiol. 212: 118–125, 2007. © 2007 Wiley‐Liss, Inc.