Propeptide-mediated regulation of procollagen synthesis in IMR-90 human lung fibroblast cell cultures. Evidence for transcriptional control.
We have demonstrated previously that the carboxyl- and amino-terminal propeptides of type I procollagen can inhibit procollagen synthesis by specifically decreasing procollagen mRNA levels. The objective of the present experiments was to determine the mechanism by which propeptides cause these pretranslational effects. IMR-90 fibroblasts were exposed to medium containing carboxyl-terminal propeptide of type I procollagen, and nuclear run-off assays were performed by hybridization to a specific alpha 1 chain type I procollagen cDNA probe. Specific type I procollagen transcription rates were found to be decreased by 50% in the presence of 75 nM carboxyl-terminal propeptide compared with control (untreated) cells. Total cellular transcription rates as well as beta-actin mRNA rates were not affected significantly by any concentration of carboxyl-terminal propeptide. Propeptide radiolabeled with 125I was found to be taken up by cultured cells. Furthermore, exogenous carboxyl-terminal propeptide levels increased in the cytosolic compartment and eventually reached a steady-state level of 18 +/- 2 pmol/g cell protein by 30 min. Of particular interest was the finding that levels of radiolabeled carboxyl-terminal propeptide were also detected in the nuclear fraction and increased with time, reaching a plateau after 60 min of incubation. Incubation of nuclei from IMR-90 cells in medium containing varying concentrations of carboxyl-terminal propeptide resulted in nuclear transcription rates that were decreased by 40% compared with untreated controls. beta-Actin nuclear message levels remained unchanged under identical conditions. We conclude that carboxyl-terminal propeptide of type I procollagen can be internalized and become associated with the nuclear compartment. This suggests a feedback regulatory role on procollagen synthesis by a direct effect on procollagen gene transcription.
Wu
C H
CH
Department of Medicine, University of Connecticut School of Medicine, Farmington 06030.
Walton
C M
CM
Wu
G Y
GY
D000818:Animals; D002478:Cells, Cultured; D002642:Chick Embryo; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000378:metabolism; D008168:Lung / Q000166:cytology / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology*; D011347:Procollagen / Q000096:biosynthesis* / Q000235:genetics; D012333:RNA, Messenger / Q000187:drug effects; D014158:Transcription, Genetic
D010455:Peptides; D011347:Procollagen; D012333:RNA, Messenger