Thyroid-stimulating hormone activates phospholipase D in FRTL-5 thyroid cells via stimulation of protein kinase C.
We studied whether TSH or phorbol myristate acetate (PMA) stimulates the hydrolysis of phospholipids, predominantly phosphatidylcholine, via phospholipase D (PLD) in FRTL-5 thyroid cells and whether this occurs as a consequence of protein kinase C (PKC) activation. FRTL-5 thyroid cells were labeled with [3H]myristate followed by incubation with 200 mM ethanol before the addition of agonist. PLD activity was assessed by the measurement of [3H]phosphatidylethanol from [3H]phospholipid (predominantly [3H]phosphatidylcholine). Compared to control values, bovine TSH (100 microU/ml) increased PLD activity by 480% and 600%, respectively, after 30 and 60 min of exposure. Studies with purified human and bovine TSH revealed similar results, indicating that this effect was due to TSH itself. PMA (100 nM) increased PLD activity at 10 min (630% of the control value), and this effect persisted up to 60 min (600% of the control value). To determine whether the effects of TSH on PLD occurred as a consequence of PKC activation, FRTL-5 thyroid cells were preincubated with the PKC inhibitor, chelerythrine (1 microM for 10 min), or were pretreated for 24 h with PMA (100 nM) to down-regulate PKC. PLD stimulation by TSH and PMA was largely abolished by such treatments. These studies indicate that in FRTL-5 thyroid cells, TSH and PMA are capable of stimulating PLD, and that PKC activation is responsible for this stimulation. The role of PLD activation could be to amplify and prolong the PKC signal by further production of diacylglycerol.
Gupta
S
S
Department of Medicine, University of Alberta, Edmonton, Canada.
Gomez-Muñoz
A
A
Matowe
W C
WC
Brindley
D N
DN
Ginsberg
J
J
D000818:Animals; D002460:Cell Line; D015536:Down-Regulation; D004789:Enzyme Activation / Q000187:drug effects; D010739:Phospholipase D / Q000032:analysis / Q000378:metabolism* / Q000502:physiology; D010743:Phospholipids / Q000378:metabolism; D011493:Protein Kinase C / Q000187:drug effects / Q000502:physiology*; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013961:Thyroid Gland / Q000166:cytology / Q000201:enzymology*; D013972:Thyrotropin / Q000494:pharmacology*; D013997:Time Factors; D014316:Tritium
D010743:Phospholipids; D014316:Tritium; D013972:Thyrotropin; D011493:Protein Kinase C; D010739:Phospholipase D; D013755:Tetradecanoylphorbol Acetate