Activation of the thyrotropin receptor (TSH-R) exhibits growth stimulatory as well
as differentiating effects upon normal and neoplastic thyroid epithelium. The TSH-R
recruits Gs, Gi/o, Gq/11, and G12/13 proteins and thus may instigate a plethora of
signalling pathways. With respect to proliferative responses, involvement of ERK1/2
was tested in previous investigations, yet with somewhat incoherent results. In the
present study, a human follicular thyroid carcinoma cell line (FTC-133) without functional
TSH-R and FTC-133 cells stably expressing either wild-type (FTC-133wt) or mutated
(FTC-133Y601H) TSH-R were investigated. In FTC-133Y601H-cells, Gq/11 coupling of TSH-R
is selectively disrupted while other pathways, e.g. cAMP formation, are still functional.
In FTC-133wt and FTC-133Y601H cells – but not in FTC-133 cells without TSH-R – stimulation
with TSH activated ERK1/2. This was mediated by EGF receptor (EGF-R) transactivation
since EGF-R inhibitor AG1478, anti-HB-EGF antibodies, and matrix metalloproteinase
inhibitors blocked ERK1/2 activation. To further characterize the distinct G-protein-dependent
pathway of ERK1/2 activation, inhibitors of phospholipase C and protein kinase C as
well as the Ca2+-chelator BAPTA and pertussis toxin were tested. None of them inhibited ERK1/2 activation
by TSH suggesting a Gq/11- and Gi/o-independent mechanism. In addition, inhibition
of cAMP-dependent protein kinase did not abolish ERK1/2 activation and adenylyl cyclase
stimulation by forskolin did not activate ERK1/2, thus indicating, that Gs was not
involved either. However, dominant negative Gα13 mutants hampered ERK1/2 activation
whereas dominant negative Gα12 mutants were ineffective. This could be substantiated
using Gα12 and Gα13-specific siRNA whereas inhibitors of possible down-stream effectors,
i.e. RhoA and Rho kinase, were without effect. These data demonstrate the first biological
read-out for TSH-R mediated Gα13 activation and underline the non-redundancy of Gα12
and Gα13 dependent signalling.
