Induction of apoptosis in HeLa cells by MSSP, c-myc binding proteins.

MSSP (c-myc gene single strand binding proteins) were identified as protein factors binding to a putative replication origin/transcriptional enhancer sequence present upstream from the human c-myc gene, and two cDNAs encoding highly homologous proteins, MSSP-1 and MSSP-2, have been cloned. Scr2, independently cloned as a factor which complements the cdc2 defective mutant of Schizosaccharomyces pombe, has turned out to be identical to MSSP-1. MSSP-1/Scr2 and MSSP-2 similarly stimulated the initiation of SV40 DNA replication, and thus were suggested to be involved in regulation of cell cycle movement, especially from the G1 to S phase. Here, we examined the functions of MSSP in apoptosis. MSSP expression plasmids were transfected to human HeLa cells together with a beta-galactosidase expression vector. After incubation in the presence of 2% calf serum, cells were stained with X-gal and morphologically apoptotic cells among the beta-galactosidase-positive cells were counted. Both MSSP-1 and 2 induced apoptosis in a dose-dependent manner as in the control experiments with c-myc or adenovirus E1A. DNA fragmentation, a hallmark of apoptosis, was also observed in cells transfected with MSSP expression plasmids. The results of experiments using various deletion mutants of MSSP indicated that the region containing one of the two RNP consensus motifs, RNP1-B, is required for induction of apoptosis as well as specific DNA binding activity.