A. Ishida et al., Tumor suppressor p53 but not cGMP mediates NO- induced expression of p21(Waf1/Cip1/Sdi1) in vascular smooth muscle cells, MOLEC PHARM, 56(5), 1999, pp. 938-946
Cyclin-dependent kinase inhibitor p21(Waf1/Cip1/Sdi1) has been suggested to
be involved in the antiproliferative effect of nitric oxide (NO) in vascul
ar smooth muscle cells (VSMCs). To elucidate the mechanism underlying NO-in
duced p21 expression, we investigated the roles of tumor suppressor p53 and
the guanylate cyclase-cGMP pathway. The induction of p21 by the NO donor S
-nitroso-N-acetylpenicillamine (SNAP) seemed to be due to transactivation b
ecause SNAP elevated the activity of p21 promoter but did not stabilize p21
mRNA and protein. Because SNAP did not stimulate the deletion mutant of p2
1 promoter that lacked p53 binding sites, we tested the involvement of p53.
The expression level of p53 was down-regulated after mitogenic stimulation
, whereas it was sustained in the presence of SNAP. SNAP markedly stimulate
d DNA binding activity of p53. Furthermore, SNAP failed to induce p21 in VS
MCs obtained from p53-knock out mice and in A431 cells that contained mutat
ed p53. The antiproliferative effect of SNAP also was attenuated in these c
ells. NO stimulates guanylate cyclase and its product cGMP has been shown t
o inhibit VSMC proliferation. However, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxali
n-1-one, a guanylate cyclase inhibitor, did not prevent SNAP-induced p21 ex
pression. 8-Bromo-cGMP, 3-isobutyl-1-methylxanthine, and their combination
did not induce p21. Although 8-bromo-cGMP had a small antiproliferative eff
ect, the elevation of cGMP concentration induced by SNAP was little through
out the G(1) phase. The antiproliferative effect of SNAP was not attenuated
by Rp-8-bromoguanosine-3',5'-monophosphorothioate, an inhibitor of cGMP-de
pendent protein kinase. These results suggested that NO induces p21 through
a p53-dependent but cGMP-independent pathway.