Coexpression of the adenovirus 12 E1B 55 kDa oncoprotein and cellular tumor suppressor p53 is sufficient to induce metaphase fragility of the human RNU2 locus

Adenovirus 12 (Ad12), but not adenovirus 2 or 5, induces metaphase chromosome fragility at four specific loci in humans: RNU1, RNU2, PSU1, and RN5S. As each of these sites corresponds to a tandemly repeated multigene family encoding a small, abundant structural RNA, we proposed that Ad12 hinders metaphase chromatin condensation, interfering either directly or indirectly with transcriptional regulation or chromatin Racking of these small RNA genes. We and others subsequently found that Ad12-induced fragility of the RNU2 locus requires U2 promoter elements, viral early functions, and p53. We now show that RNU2 fragility can be induced by transfection with an expression vector encoding Ad12 E1B 55 kDa alone but not by an E1 vector encoding all E1 products (3 E1A proteins, as well as the E1B 19 kDa and 55 kDa proteins). Although Ad12 E1B 55 kDa efficiently induced fragility in transfected cells, Ad2 E1B 55 kDa did not. By swapping domains between the Ad12 and Ad2 E1B, we found that the aminoterminus of Ad12 E1B is required for induction of fragility and that the ability of the hybrid E1B proteins to induce fragility appears to correlate with nuclear localization. Furthermore, in Saos-2 cells lacking p53 function, RNU2 fragility could be induced by cotransfection with vectors encoding Ad12 E1B 55 kDa and either wild-type p53 or the R273H mutant with impaired DNA binding activity. We conclude that a functional land probably physical) interaction between Ad12 E1B 55 kDa and p53 within the nucleus is sufficient to induce metaphase fragility of the RNU2 locus. (C) 1999 Academic Press.