EFFECTS OF EXOGENOUS P16(INK4A) ON GROWTH OF CELLS WITH VARIOUS STATUS OF CELL-CYCLE REGULATORS

p16(INK4a), a protein that inhibits cyclin-dependent kinase 4 (Cdk4) a nd Cdk6, is deficient in many human cancers and in established lines o f tumor cells. It has been reported that transfection with cDNA for p1 6(INK4a) inhibits the growth of cell lines that express retinoblastoma protein (pRB). However, it is unclear whether the introduction of cDN A for p16(INK4a) affects the growth of cells that express p16(INK4a) p rotein. Moreover, the effects of other cell-cycle regulators on the in hibition of cell growth by p16(INK4a) remain unknown. In this study, c DNA for p16(INK4a) was used to transfect human cell lines that had var ious status of expression of RE pathway-related proteins, such as memb ers of the RE family proteins and Cdk-inhibitory proteins. We found th at status of p107, p130, p15(INK4b), p18(INK4c) p21(Cip1), p27(Kip1) c yclin D1, and Cdk4 were not correlated with the growth-inhibitory acti vity of exogenous p16(INK4a). BY contrast, transfection with cDNA for p16(INK4a) had a significant effect on the growth of cells depended on the status not only of pRB but also of p16(INK4a). Although exogenous p16(INK4a) inhibited the growth of cells that expressed pRB but did n ot express p16(INK4a) (pRB(+)/p16(-) cells), it had little affect on e ither pRB(-)/p16(+) cells or pRB(-)/p16(+) cells. Moreover, transfecti on with cDNA for p16(INK4a) also inhibited the activity of the E2 prom oter of the dehydrofolate reductase gene in the same manner that depen ded on the absence of p16(INK4a), as well as on the presence of pRB. T hese results suggest that deregulation of the RE pathway by p16(INK4a) deficiency plays a very important role in the proliferation of cells that lack p16(INK4a) protein. (C) 1997 Academic Press.