Regulation of protein kinase C isoforms in FRTL-5 thyroid cells by TSH and phorbol ester.
Activation of protein kinase C (PKC) has been observed following TSH exposure in FRTL-5 thyroid cells. However, PKC exists as a family of isoforms displaying individual characteristics. Recently, Wang et al. identified alpha-, delta-, epsilon-, and zeta-PKC in FRTL-5 thyroid cells chronically exposed to TSH. To determine if these PKC isoforms are regulated by TSH, we examined PKC isoforms by Western blotting in FRTL-5 thyroid cells after depletion of TSH followed by short-term TSH exposure (100 microU/mL; 30 min). Phorbol 12-myristate 13-acetate (PMA; 100 nM, 10 min) served as a positive control. In untreated cells, alpha-, epsilon-, and zeta-PKC isoforms were identified in both cytosolic and membrane fractions. Using the specific antigenic peptide to the respective PKC isoforms identified, each band could be appropriately displaced. PMA caused the translocation of alpha-PKC from the cytosol to the membrane-bound fraction. Although membrane-bound epsilon-PKC and zeta-PKC were increased following PMA exposure, the cytosolic fraction was unaffected. TSH had no effect on the cytosolic fractions of any of the PKC isoforms identified but was able to increase the membrane-bound fractions of alpha-, epsilon-, and zeta-PKC. Although delta-PKC was observed in FRTL-5 thyroid cells cultured in TSH-supplemented medium, delta-PKC disappeared within 24 h of TSH depletion and reappeared 24 h after TSH reexposure with further increases up to 72 h. These studies indicate that the PKC isoforms present in FRTL-5 thyroid cells are regulated by both phorbol ester and TSH and that the duration of TSH exposure differentially regulates delta-PKC. These observations provide further evidence that PKC is a mediator of TSH action in FRTL-5 thyroid cells in vitro.
Matowe
W C
WC
Department of Medicine, University of Alberta, Edmonton, Canada.
Gupta
S
S
Ginsberg
J
J
D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D013961:Thyroid Gland / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D013972:Thyrotropin / Q000494:pharmacology*
D007527:Isoenzymes; D013972:Thyrotropin; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate