Canine hepatocyte growth factor: molecular cloning and characterization of the recombinant protein
2003
Miyake, Masashi | Saze, Koh-ichi | Yaguchi, Takahiro | Wang, Jianhui | Suzuta, Yasuyuki | Haga, Yoshihisa | Takahashi, Susumu Y. | Yamamoto, Yoshimi | Iwabuchi, Shigehiro
Hepatocyte growth factor (HGF) is a pleiotropic cytokine originally identified and cloned as a potent mitogen for hepatocytes. The HGF receptor is the transmembrane tyrosine kinase encoded by c-MET proto-oncogene. Various lines of evidence suggest that the HGF/c-MET receptor system plays essential roles in monocyte–macrophage function, mammalian development, angiogenesis and organ regeneration. We have cloned canine HGF (CaHGF) cDNA from leukocytes by the methods of reverse transcription (RT)-polymerase chain reaction (PCR) and rapid amplification of cDNA ends (RACE). Canine HGF contains an open reading frame (ORF) of 2193 nucleotides, coding for 730 amino acids. The deduced amino acid sequence of canine HGF shows 97.5, 92.3, 92.1, and 92.0% homologies with those of feline, human, mouse, and rat, respectively. The possible glycosylation sites, cysteine residues linking the α and β chains and the proteolytic processing site are conserved in all species. In addition, we have found a variant cDNA that deleted a sequence of 15 base pairs in the first kringle domain (K1) and resulted in the deletion of five amino acids. To confirm the biological activities of canine HGF cDNAs, both cDNAs were transiently expressed in COS-7 cells. The conditioned medium from the canine HGF-transfected COS-7 cells stimulated the growth of BNL CL.2 cells (a mouse hepatocyte cell) and scattering activity of Madin–Darby canine kidney (MDCK) cells. The materials reported here will be a crucial resource for further studies of canine HGF.
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