MUTATIONS IN CR-1 OF E1A 12S YIELD DOMINANT-NEGATIVE SUPPRESSORS OF IMMORTALIZATION AND THE LYTIC CYCLE

The Adenovirus 5 E1A 12S gene is responsible for the establishment of immortalization or primary cells by Adenovirus. We have identified two mutants of 12S (30K and NTdI814), which, when coexpressed with wild-t ype 12S in primary baby rat kidney cells, were capable of suppressing the immortalizing function of the wild-type 12S gene, even when the mu tant proteins were expressed at levels lower than wild type. 30K and N TdI814 did not affect the ability of the coexpressed 12s to activate t he cell cycle, but have a suppressive effect on 12S-induced DNA synthe sis and proliferation at late times in the immortalization pathway. Bo th the dominant negative mutants have a deletion in conserved region ( CR)1 in the first exon of E1A, which encompasses one of the pRb-family binding regions. However, the mutants did not affect the binding of c ellular proteins to full-length 12S. A suppressive effect an wild-type 12S was not observed with mutants that have lost any other region or function. In addition, expression of 30K, which is equivalent to the p rotein encoded by the 10S mRNA of E1A, inhibited E1A function in lytic cycle. Thus, loss of the CR1 seems to be a prerequisite for a mutant to have a dominant negative effect on E1A functions. (C) 1997 Academic Press.