Dissection of GnRH receptor-G protein coupling
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White2009.doc (14.60Mb)
Date
2009Author
White, Colin D.
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Abstract
Hypothalamic gonadotropin-releasing hormone (GnRH) (GnRH I) is the central
regulator of the mammalian reproductive system. Most vertebrates studied also
possess a second form of GnRH, GnRH II. GnRH I acts on its cognate G proteincoupled
receptor (GPCR) on pituitary gonadotropes and activates Gq/11-mediated
signalling pathways to stimulate the biosynthesis and the release of luteinising
hormone (LH) and follicle-stimulating hormone (FSH). Both GnRHs have also been
suggested to inhibit cellular proliferation, an action which has largely been proposed
to be mediated by the coupling of the receptor to Gi/o. However, the range of G
proteins activated by the GnRH receptor and the signalling cascades involved in
inducing antiproliferation remain controversial.
To delineate the G protein coupling selectivity of the mammalian GnRH receptor and
to identify the signalling pathways involved in GnRH I-mediated cell growth
inhibition, I examined the ability of the receptor to interact with Gq/11, Gi/o and Gs in
Gαq/11 knockout MEF cells. My results indicate that the receptor is unable to interact
with Gi/o but can signal through Gq/11. Additionally, my data do not support the
suggestion of GnRH receptor-Gs interaction. Furthermore, I show that the GnRH Iinduced
inhibition of cell growth is dependent on Gq/11, src and extracellular signal
regulated kinase (ERK) but is independent of the activity of protein kinase C (PKC),
Ca2+, jun-N-terminal kinase (JNK) or P38. Based on these findings and previous
research within our group, I propose a mechanism whereby GnRH I may induce
antiproliferation.
Previous studies from our laboratory suggest that the GnRH receptor can adopt
distinct active conformations in response to the binding of GnRH I and GnRH II.
These data thus account for our hypothesis of ligand-induced selective signalling
(LiSS). Given my previous results, I examined the ability of the GnRH receptor to
couple to G12/13. My work indicates that the receptor can directly activate G12/13 and
the downstream signalling cascades associated with this G protein family. Indeed, I
provide evidence, in several cellular backgrounds, to suggest that GnRH receptor-
G12/13-mediated signalling is involved in the regulation of GnRH-induced MAPK
activity, SRE-driven gene transcription and cytoskeletal reorganisation.
Furthermore, I propose a role for these G proteins in the transcriptional regulation of
LHβ and FSHβ. Finally, I confirm previous results from our laboratory indicating
that the GnRH receptor may interact with src Tyr kinase and show that GnRH I but
not GnRH II may, independently of Gq/11, stimulate the Tyr phosphorylation and thus
the activation of this protein. I propose that this differential signalling accounts for
the distinct effects of GnRH I and GnRH II on cellular morphology and SREpromoted
transcriptional activity.
The research presented within this thesis provides evidence to refute published
conclusions based on largely circumstantial experimental data, describes novel
GnRH receptor signalling pathways and offers support for the concept of LiSS. It
may assist in the development of new therapeutic compounds which selectively
target one GnRH-mediated signalling pathway while bypassing others.