ISSN:
0021-9541
Schlagwort(e):
Life and Medical Sciences
;
Cell & Developmental Biology
Quelle:
Wiley InterScience Backfile Collection 1832-2000
Thema:
Biologie
,
Medizin
Notizen:
Basic fibroblast growth factor (bFGF, FGF-2) is progressively lost from mammary epithelial cells as they become malignant. To investigate the effects of restoring the expression of bFGF in breast cancer cells, we constructed MCF-7 cells that permanently overexpress 18-kD cytoplasm-localizing bFGF (MCF-7/ΔAFGF(18) cells) and cells that express both the 18-kD along with the 22- and 24-kD nucleus-localizing bFGF peptides (MCF-7/NCFFGF(18,22,24) cells), using retroviral transduction. These stable cell constructs grew more slowly and had a larger fraction of their populations in the G0/G1 phase of the cell cycle than control cells. All forms of bFGF were eluted from MCF-7/NCFFGF(18,22,24) cell monolayers with 2 M NaCl, in contrast to fibroblasts that were demonstrated to secrete only the 18-kD bFGF isoform. High-affinity binding of 18-kD 125I-bFGF to these cells was significantly decreased, probably because of competitive binding by the autocrine-secreted bFGF. Recombinant 18-kD bFGF that was previously demonstrated in our laboratory to inhibit proliferation, activate MAP kinase, and induce the cyclin-dependent kinase inhibitor p21WAF1/CIP1 in MCF-7 cells, further inhibited MCF-7/ΔAFGF(18) cells but had no effect on MCF-7/NCFFGF(18,22,24) cells. The total cellular content of the high-affinity FGF receptors 1-3 was unchanged, but FGF receptor 4 was decreased in MCF-7/NCFFGF(18,22,24) cells. Both cell types overexpressing bFGF isoforms had elevated levels of the cyclin-dependent kinase inhibitor p27Kip1 but not that of p21WAF1/CIP1. In MCF-7/ΔAFGF(18) cells, FGFR1 and MAP kinase were constitutively phosphorylated. Exogenous recombinant 18-kD bFGF did not accentuate these effects but did induce an increase in the levels of p21WAF1/CIP1 corresponding to the further inhibition induced by exogenous bFGF in these cells. In MCF-7/NCFFGF(18,22,24) cells, FGFR1 and MAP kinase were not phosphorylated at baseline nor upon stimulation with recombinant bFGF, and exogenous bFGF only had a minimal effect on low steady-state p21WAF1/CIP1 levels. However, stimulation of these cells with phorbol ester or insulin did result in MAP kinase phosphorylation. While growth-inhibited in the G1 phase of the cell cycle, MCF-7/NCFFGF(18,22,24) cells retained active isoforms of cdk2 and the hyperphosphorylated form of Rb. These data suggest that high molecular weight forms of bFGF overexpressed in MCF-7 cells do not activate the receptor-mediated MAP kinase pathway, and do not induce p21WAF1/CIP1 in an autocrine manner, but inhibit proliferation through other, possibly direct nuclear signalling mechanisms. J. Cell. Physiol. 177:411-425, 1998. © 1998 Wiley-Liss, Inc.
Zusätzliches Material:
7 Ill.
Materialart:
Digitale Medien
Permalink
