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Persistent cAMP signaling by thyrotropin (TSH) receptors is not dependent on internalization
Author(s) -
Neumann Susanne,
GerasRaaka Elizabeth,
MarcusSamuels Bernice,
Gershengorn Marvin C.
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.10-161745
Subject(s) - internalization , adenylyl cyclase , receptor , signal transduction , thyrotropin receptor , g protein coupled receptor , microbiology and biotechnology , endocrinology , medicine , chemistry , camp dependent pathway , g protein , dynamin , biology , endocytosis , biochemistry , thyroid , graves' disease
Evidence was presented that thyrotropin [thyroid‐stimulating hormone (TSH)]‐stimulated persistent cAMP signaling is dependent on receptor (with G‐protein a subunits and adenylyl cyclase) internalization. Because it is not clear whether G proteins and adenylyl cyclase internalize with receptors, we tested whether persistent cAMP signaling by TSH receptor (TSHR) is dependent on internalization. We measured persistent TSHR signaling as an accumulation of cAMP in HEK‐EM293 cells permanently expressing human TSHRs incubated with isobutylmethylxanthine for 30 min after washing the cells to remove unbound TSH, and TSHR internalization by fluorescence microscopy using Alexa‐tagged TSH and binding assays using 125 I‐TSH. TSHRs, but not the closely related lutropin or follitropin receptors, exhibit persistent cAMP signaling. TSHRs were not internalized by 30 min incubation with unlabeled TSH; however, expression of β‐arrestin‐2 promoted TSHR internalization that was inhibited by dynasore, a dynamin inhibitor. Expression of β‐arrestin‐2 had no effect on TSHR cAMP signaling, dynasore inhibited TSHR cAMP signaling in the absence or presence of TSHR internalization, and expression of a dominantnegative mutant dynamin, which inhibited internalization, had no effect on persistent cAMP signaling. Persistent cAMP signaling was specifically inhibited by a small molecule TSHR antagonist. We conclude that TSHRs do not have to be internalized to exhibit persistent cAMP signaling.—Neumann, S., Geras‐Raaka, E., Marcus‐Samuels, B., Gershengorn, M. C. Persistent cAMP signaling by thyrotropin (TSH) receptors is not dependent on internalization. FASEB J. 24, 3992–3999 (2010). www.fasebj.org