Protein phosphatase 2A and phosphatidylinositol 3-kinase regulate the activity of Sp1-responsive promoters

The transcription factor Spl regulates the activity of a large number of eukaryotic gene promoters, including early SV40 and human immunodeficiency virus type 1 (HIV-1), Here, we report that expression of SV40 small tumor antigen (small t) in quiescent CV-1 cells transactivates two Spl-responsive promoters, including a deletion mutant of HIV-1 LTR, through specific inhibition of endogenous AC and AB alpha C forms of protein phosphatase 2A (PP2A). Expression of a small t mutant, lacking the PP2A-binding domain, failed to transactivate Spl. Overexpression of the B56 alpha, B56 beta, and B56 gamma 1 regulatory PP2A subunits strongly inhibited the ability of small t, but not the phosphatase inhibitor, okadaic acid, to enhance Spl-driven gene expression. Using inhibitors and co-expression of kinase-deficient mutants, we also show that functional phosphatidylinositol 3-kinase (PI 3-kinase) and atypical protein kinase C zeta are required for small t-induced Spl-dependent promoter transcriptional activation. Moreover, two inhibitors of PI 3-kinase, wortmannin and LY294002, inhibit the initiation of SV40 DNA replication in quiescent CV-1 cells. Taken together, these results suggest that PP2A and PI 3-kinase contribute to the ability of small t to regulate Spl activity, stimulate early SV40 DNA replication, and enhance the transformation of resting cells during SV40 infection.