The C-terminal region of fibroblast growth factor-1 is crucial for its biological activity and high level protein expression in mammalian cells.
We have studied the role of the carboxyl(C)-terminus of fibroblast growth factor(FGF)-1 using prokaryotic and eukaryotic expression systems. The full-length FGF-1 protein and its mutants lacking 6- and 9-amino acids at the C-terminus IFGF-1 (Cdel6) and FGF-1 (Cdel9), respectively] could be expressed in E. coli cells at the similar levels. The deletion mutants bound very weakly to FGF receptor and to heparin, and did not stimulate DNA synthesis in BALB/c3T3 cells. In contrast to E. coli cells, in NIH3T3 transfectants and L6 transfectants, the protein expression level of FGF-1 (Cdel6) was significantly lower than that of FGF-1, and longer C-terminal deletions further decreased the protein expression levels. However, the level of transcripts in the transfectants and the level of translates in in vitro system were equivalent for all the FGF-1 constructs. Treatment with proteasome inhibitors of the NIH3T3 transfectants expressing FGF-1(Cdel6) increased the protein level six-fold. The results indicate that the C-terminus of FGF-I is crucial for its biological activity and high-level expression in mammalian cells and suggest that deletion of the C-terminus of FGF-1 induces its post-translational degradation by proteasome system.
Miyakawa
K
K
Biosignaling Department, National Institute of Bioscience and Human Technology, Tsukuba, Ibaraki, Japan.
Ozawa
K
K
Uruno
T
T
Imamura
T
T
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D004305:Dose-Response Relationship, Drug; D016220:Fibroblast Growth Factor 1; D016222:Fibroblast Growth Factor 2 / Q000737:chemistry* / Q000502:physiology*; D015870:Gene Expression; D006493:Heparin / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D014176:Protein Biosynthesis; D013997:Time Factors; D014158:Transcription, Genetic
D016222:Fibroblast Growth Factor 2; D016220:Fibroblast Growth Factor 1; D006493:Heparin