Arta Physiol Scand 1992, 146, 511-518
Modulation of G proteins and second messenger responsiveness by steroid hormones in GH, rat pituitary tumour cells R. H . P A U L S S E N , E. J. P A U L S S E N , K. M. G A U T V I K and J. 0. G O R D E L A D Z E Institute of Medical Biochemistry, University of Oslo, Norway
E. J., GAUTVIK, K. M. & GORDELADZE, J. 0. 1992. PAULSSEN, R. H., PAULSSEN, Modulation of G proteins and second messenger responsiveness by steroid hormones in GH, rat pituitary tumour cells. Acta PhysiolScand 146, 511-518. Received 25 March 1992, accepted 21 July 1992. ISSN 0001-6772. Institute of Medical Biochemistry, University of Oslo, Norway. We have investigated the modulation of different G protein a- and /%subunit levels in prolactin (PRL) and growth hormone producing rat pituitary adenoma cells (GH, cells) in culture after prolonged exposure (6-48 h) to the steroid hormones 17/%oestradioland dexamethasone. G,_,a- and GP-subunits were the only G protein subunits which increased in response to lo-' M oestradiol (to approximately 150 and 200% of controls, respectively), while the other a-subunits investigated (G,a, G,_,a and G,a) remained relatively unchanged. Thyroliberin ( T R H F a n d guanosine 5'-[/ly-imido]trisphosphate (Gpp(NH)p)-elicited adenylyl cyclase (AC) activities were reduced during 6-12 h of oestradiol treatment (by 60 and 20%, respectively), while the inhibitory effect of somatostatin (SRIF) increased by approximately 100%. Dexamethasone (lo-' M) increased levels of the stimulatory G protein G p (to approximately 3400/6) and decreased levels of Gl-3ci (to 25%). After 48 h, the AC response to TRH was reduced by approximately 70%, whereas the effect of the other modulators remained close to controls. We conclude that G protein subunits in GH, cells are subject to specific regulation by steroid hormones and that this may be important in the tuning of the responsiveness of PRL secretion to hormones in the in vzuo situation. Key words : adenylyl cyclase, dexamethasone, G proteins, 17&oestradiol, rat pituitary cells
GH, cells are clonal rat pituitary tumour cells that produce and spontaneously secrete prolactin (PRL) and growth hormone ( G H ) to the culture medium (Tashjian 1979). Hormone synthesis and secretion is subject to regulation by a variety of physiological and pharmacological agents, such as the peptide hormones thyroliberin (TRH), vasoactive intestinal peptide (VIP) and somatostatin (SRIF) (Gautvik et al. 1988) and steroid hormones such as oestrogen (Haug & Gautvik 1976, Amara et al. 1987) and glucoCorrespondence : Ruth H. Paulssen, Institute of Medical Biochemistry, P.O. Box 1112 Blindern, 0317 Oslo 3, Norway.
corticoids (Clausen et a/. 1978, Naess et al. 1980). Steroid hormones have previously been shown to modify membrane signalling systems involving guanine nucleotide-binding proteins ( G proteins). Experiments involving in uiuo treatment of rats with 17P-oestradiol have demonstrated an effect on the content of pituitary G proteins, involving a specific increase of G p D aand a general reduction of most of the other a- and subunits (Bouvier et al. 1991). Similarly, 17boestradiol administration also increases the inhibitory action of SRIF on hormone-sensitive adenylyl cyclase (AC) in rat pituitary (Kimura & Hayafuji 1989). Furthermore, mouse striatal
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neurons in culture have been shovm to respond to 17/l-oestradiol with enhancement of the pertussis toxin-catal>-sed .\KIP-ribos!-lation of Gclzand G,J (hlaus et ( I / . 1990), and an increased response to biogenic amincs on -4C has been obseried (Maus r t a / . 1989). I//l-oestradiol also reduces /I-adrenoreceptor-mediated c.-\\LP production and G, content in rahbit mymietriurn (Riemer LV ul. 1988). tilucocorticoids modulate the steady-state levels ofG protein subunits in liver (Garcia-Sainz rt 1 i I . 1989) and mRN.1 h e l s for G protein j j subunits in rat tit-cells (Ros et ol. 1989). Desamethasone affects the / h d r e n e r g i c receptor and G, and G?,protein lei-els in ROS 17/2.8 cells (Rodan & Rodan 1986),and increases -4C acti\-it!and expression of protein in GH,, cells ((:hang Si Uournc 1987). I n addition to the direct gene regulatory action of' li/i-oestradiol and dexamethasone, modification of membrane signalling components including alterations of stead! -state le\-els of G protein subunits, may be of central importance in the action mechanisms of' these steroid hormones. \Ye have recentI!- in\ estigated the modulation of' G protein a-subunit m R S = \ levels b! T R H , TIP and SKIF in G€& cells (Paulssen et a / . 1991 a), and also demonstrated that long-term treatment a i t h these peptide hormones, Ivhich are invohed in regtilation of PRL, synthesis and release, causes alterations in the transmembrane signalling systems in\-olved in their actions and subsequent changes in second messenger response (Paulssen ct al. 1092). I n this n-ork we have employed GH,, cells to elucidate the influence of l i / h e s t r a d i o l and desamethasont on G protein l e d s and the consequences for peptide hormone action on hornlone-dependent
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Modulation of G proteins and second messenger responsiveness by steroid hormones in GH, rat pituitary tumour cells R. H . P A U L S S E N , E. J. P A U L S S E N , K. M. G A U T V I K and J. 0. G O R D E L A D Z E Institute of Medical Biochemistry, University of Oslo, Norway
E. J., GAUTVIK, K. M. & GORDELADZE, J. 0. 1992. PAULSSEN, R. H., PAULSSEN, Modulation of G proteins and second messenger responsiveness by steroid hormones in GH, rat pituitary tumour cells. Acta PhysiolScand 146, 511-518. Received 25 March 1992, accepted 21 July 1992. ISSN 0001-6772. Institute of Medical Biochemistry, University of Oslo, Norway. We have investigated the modulation of different G protein a- and /%subunit levels in prolactin (PRL) and growth hormone producing rat pituitary adenoma cells (GH, cells) in culture after prolonged exposure (6-48 h) to the steroid hormones 17/%oestradioland dexamethasone. G,_,a- and GP-subunits were the only G protein subunits which increased in response to lo-' M oestradiol (to approximately 150 and 200% of controls, respectively), while the other a-subunits investigated (G,a, G,_,a and G,a) remained relatively unchanged. Thyroliberin ( T R H F a n d guanosine 5'-[/ly-imido]trisphosphate (Gpp(NH)p)-elicited adenylyl cyclase (AC) activities were reduced during 6-12 h of oestradiol treatment (by 60 and 20%, respectively), while the inhibitory effect of somatostatin (SRIF) increased by approximately 100%. Dexamethasone (lo-' M) increased levels of the stimulatory G protein G p (to approximately 3400/6) and decreased levels of Gl-3ci (to 25%). After 48 h, the AC response to TRH was reduced by approximately 70%, whereas the effect of the other modulators remained close to controls. We conclude that G protein subunits in GH, cells are subject to specific regulation by steroid hormones and that this may be important in the tuning of the responsiveness of PRL secretion to hormones in the in vzuo situation. Key words : adenylyl cyclase, dexamethasone, G proteins, 17&oestradiol, rat pituitary cells
GH, cells are clonal rat pituitary tumour cells that produce and spontaneously secrete prolactin (PRL) and growth hormone ( G H ) to the culture medium (Tashjian 1979). Hormone synthesis and secretion is subject to regulation by a variety of physiological and pharmacological agents, such as the peptide hormones thyroliberin (TRH), vasoactive intestinal peptide (VIP) and somatostatin (SRIF) (Gautvik et al. 1988) and steroid hormones such as oestrogen (Haug & Gautvik 1976, Amara et al. 1987) and glucoCorrespondence : Ruth H. Paulssen, Institute of Medical Biochemistry, P.O. Box 1112 Blindern, 0317 Oslo 3, Norway.
corticoids (Clausen et a/. 1978, Naess et al. 1980). Steroid hormones have previously been shown to modify membrane signalling systems involving guanine nucleotide-binding proteins ( G proteins). Experiments involving in uiuo treatment of rats with 17P-oestradiol have demonstrated an effect on the content of pituitary G proteins, involving a specific increase of G p D aand a general reduction of most of the other a- and subunits (Bouvier et al. 1991). Similarly, 17boestradiol administration also increases the inhibitory action of SRIF on hormone-sensitive adenylyl cyclase (AC) in rat pituitary (Kimura & Hayafuji 1989). Furthermore, mouse striatal
1-
511 A C T 146
512
R . H. Pnulssen et al.
neurons in culture have been shovm to respond to 17/l-oestradiol with enhancement of the pertussis toxin-catal>-sed .\KIP-ribos!-lation of Gclzand G,J (hlaus et ( I / . 1990), and an increased response to biogenic amincs on -4C has been obseried (Maus r t a / . 1989). I//l-oestradiol also reduces /I-adrenoreceptor-mediated c.-\\LP production and G, content in rahbit mymietriurn (Riemer LV ul. 1988). tilucocorticoids modulate the steady-state levels ofG protein subunits in liver (Garcia-Sainz rt 1 i I . 1989) and mRN.1 h e l s for G protein j j subunits in rat tit-cells (Ros et ol. 1989). Desamethasone affects the / h d r e n e r g i c receptor and G, and G?,protein lei-els in ROS 17/2.8 cells (Rodan & Rodan 1986),and increases -4C acti\-it!and expression of protein in GH,, cells ((:hang Si Uournc 1987). I n addition to the direct gene regulatory action of' li/i-oestradiol and dexamethasone, modification of membrane signalling components including alterations of stead! -state le\-els of G protein subunits, may be of central importance in the action mechanisms of' these steroid hormones. \Ye have recentI!- in\ estigated the modulation of' G protein a-subunit m R S = \ levels b! T R H , TIP and SKIF in G€& cells (Paulssen et a / . 1991 a), and also demonstrated that long-term treatment a i t h these peptide hormones, Ivhich are invohed in regtilation of PRL, synthesis and release, causes alterations in the transmembrane signalling systems in\-olved in their actions and subsequent changes in second messenger response (Paulssen ct al. 1092). I n this n-ork we have employed GH,, cells to elucidate the influence of l i / h e s t r a d i o l and desamethasont on G protein l e d s and the consequences for peptide hormone action on hornlone-dependent
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