The Role of the Calpain-Calpastatin System in Thyrotropin-releasing Hormone-induced Selective Down-regulation of a Protein Kinase C Isozyme, nPKCε, in Rat Pituitary GH4C1 Cells (∗)

• Published in: The Journal of biological chemistry 1995-10, Vol.270 (42), p.25115-25120
• Main Authors: Eto, Akiko, Akita, Yoshiko, Saido, Takaomi C., Suzuki, Koichi, Kawashima, Seiichi
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Calcium-Binding Proteins - physiology; Calpain - physiology; Cell Line; Down-Regulation; Isoenzymes - analysis; Molecular Sequence Data; Pituitary Gland - enzymology; Protease Inhibitors - pharmacology; Protein Kinase C - analysis; Rats; Thyrotropin-Releasing Hormone - pharmacology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.270.42.25115

We have examined the mechanism for the selective down-regulation of protein kinase C ε (nPKCε) in rat pituitary GH4C1 cells responding to thyrotropin-releasing hormone (TRH) stimulation. Among various low molecular weight protease inhibitors examined, only a cysteine protease inhibitor (calpain inhibitor I, N-acetyl-Leu-Leu-norleucinal) blocked the down-regulation of nPKCε. Furthermore, the introduction of a synthetic calpastatin peptide, an exclusively specific inhibitor of calpain, into the cells also reduced the down-regulation, suggesting the involvement of calpain among all the intracellular cysteine proteases in this process. In accordance, we observed TRH-induced translocation of m-calpain from the cytosol to the membrane and the concomitant up-regulation of calpastatin isoforms; presumably, the former represents activation of the protease initiating the kinase degradation, while the latter constitutes a negative feedback system protecting the cells from activated calpain. These results suggest that in GH4C1 cells, TRH mobilizes both protease (m-calpain) and inhibitor (calpastatin) as a strictly regulating system for the nPKCε pathway mediating TRH signals.