Polyomaviruses including human JCV and BKV, and their simian counterpart, SV40, are small non-enveloped viruses with a single copy of double-stranded DNA. After viral infection, the cells are forced to reenter the S-phase of the cell cycle, so that the DNA replication machinery of the cell becomes available to the virus. Only the early region is active at this time of infection. It transcribes a common precursor RNA, which is differentially spliced yielding several viral products among which the tumor antigens (large and small T-antigens) predominate (Khalili and Stoner, 2001). Polyomaviruses infect humans, monkeys, rodents, and birds with a restricted host and tissue specificity. The infection of tissues in which the virus does not replicate efficiently may lead to partial activation of the virus, expression of an early viral genome (Imperiale, 2001), dysregulation of cell growth mechanisms, and possibly transformation (Reiss and Khalili, 2003). The transforming properties of SV40 and JCV T-antigens are well established in the literature (Khalili and Stoner, 2001). The major known cellular targets for T-antigen-mediated cell cycle dysregulation are p53 and pRb (Chen et al., 1992; Kao et al., 1993; Saenz-Robles et al., 2001). Although T-antigen-mediated inactivation of these nuclear proteins is very important for the fate of the affected cell, these interactions are well documented in the literature and will not be discussed further in this review. Instead, we will attempt to explore the molecular and cellular mechanisms by which T-antigens affect integrity of the genome. As previously reported, T-antigens are strongly suspected to play a role in the development of genomic instability. However, the molecular mechanisms involved in this process are not well characterized. Several studies have documented a substantial chromosomal instability with no consistent patterns and often with many new karyotypes emerging at each consecutive passage of T-antigen positive cells (Hunter and Gurney, 1994; Woods et al., 1994; Ramel et al., 1995; Kappler et al., 1999; Ricciardiello et al., 2003). The diversity in chromosomal defects found in cells carrying T-antigen suggests a random nature for its action. This could also suggest that T-antigens affect stability of the genome at a very basic level. One possibility, which could explain the appearance of random mutations in T-antigen expressing cells, is an obstruction of DNA repair of double strand breaks (DSBs). As previously mentioned, DSBs must be repaired to ensure cell survival, especially when cells progress through the cell cycle. To guarantee uninterrupted DNA replication and to avoid apoptosis, which is readily induced by the stalled replication forks, at least one of the two major DNA repair mechanisms: homologous recombination directed DNA repair (HRR) or NHEJ, has to be active. The first documentation of a possible interaction between T-antigen and DSBs repair was furnished by experiments in which the formation of MRE11 nuclear foci was significantly disrupted by the presence in SV40 T-antigen (Digweed et al., 2002). Since the attenuation of the foci formation was found in both T-antigen immortalized cells and cells transiently expressing this viral oncoprotein, it was concluded that the effect on DNA repair was direct, and did not rely on secondary mutations. Similarly, another DNA repair protein, Nbs1, which forms an early DNA repair complex with MRE11 and Rad50, was shown to interact with SV40 T-antigen, disrupting DNA replication control (Wu et al., 2004). In addition, our results demonstrate that large T-antigen from JC virus inhibits HRR, resulting in an accumulation of mutations during DNA repair (JCP 2005, in press)1. In this process, JCV T-antigen does not operate directly but utilizes a cytosolic molecule, insulin receptor substrate 1 (IRS-1), which is the major cytosolic substrate for the IGF-I receptor (Sun et al., 1991; Myers et al., 1994). Following T-antigen-mediated nuclear translocation (Lassak et al., 2002), IRS-1 binds Rad51 at the site of damaged DNA. This T-antigen-mediated inhibition of HRR does not function in cells lacking IRS-1, and can be reproduced in the absence of T-antigen by a mutant of IRS-1, which contains artificial nuclear localization signal. These observations could define a new strategy by which T-antigens interfere with the insulin-like growth factor I receptor (IGF-IR) signaling system compromising the fidelity of DNA repair. This could be even more relevant since the activated IGF-I receptor was shown to support faithful DNA repair, possibly contributing to the maintenance of the genome during cell proliferation (Trojanek et al., 2003).