Growth-inhibitory functions of a mutated gelsolin (His321) in NIH/3T3 mouse fibroblasts.
We have previously established a murine flat revertant cell line R1 from an activated H-ras transformant EJ-NIH/3T3 by subjecting it to ethyl methanesulfonate. From the R1 cells, we cloned a mutated gelsolin gene His321 and have shown the inhibitory activity of His321 against EJ-NIH/3T3 tumors. Our present experiments were conducted to find out whether the His321 gene has any effects on untransformed NIH/3T3 fibroblasts. Rhodamine-phalloidin staining revealed that two NIH/3T3 clones expressing His321 (NIH/lambda 2S-3 and NIH/lambda 2S-6) form organized actin stress fibers as two clones transfected with the vector alone (NIH/neo-3 and NIH/neo-5). We also found that in a liquid medium, NIH/lambda 2S-3 and NIH/lambda 2S-6 grew more slowly than NIH/neo-3 and NIH/neo-5 and that the doubling times of the former were about 10 h slower than those of the latter. To investigate the effects of His321 on the signal transduction pathway necessary for cell growth, we stimulated the cell lines by prostaglandin E1 (PGE1), a platelet-derived growth factor (PDGF), or the epidermal growth factor (EGF). Although stimulation by PGE1 increased intercellular cyclic AMP in R1 cells, it did not do so in NIH/lambda 2S-3 and NIH/lambda 2S-6 cells. On the other hand, stimulation by PDGF or EGF induced far less DNA synthesis in NIH/lambda 2S-3 and NIH/lambda 2S-6 than in NIH/neo-3 and NIH/neo5. These results suggest that through the effects on the signal transduction pathway of PDGF and/or EGF His321-mutated gelsolin inhibits the growth of NIH/3T3.
Ishizaki
A
A
Second Department of Surgery, Asahikawa Medical College, Japan.
Fujita
H
H
Kuzumaki
N
N
D016475:3T3 Cells; D000199:Actins / Q000502:physiology; D000527:Alprostadil / Q000494:pharmacology; D000818:Animals; D002455:Cell Division / Q000502:physiology; D000242:Cyclic AMP / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis; D018260:Gelsolin / Q000235:genetics / Q000502:physiology*; D006131:Growth Inhibitors / Q000502:physiology*; D006639:Histidine / Q000502:physiology; D051379:Mice; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D017354:Point Mutation; D014162:Transfection
D000199:Actins; D018260:Gelsolin; D006131:Growth Inhibitors; D010982:Platelet-Derived Growth Factor; D006639:Histidine; D004247:DNA; D000242:Cyclic AMP; D000527:Alprostadil