Searchable abstracts of presentations at key conferences in endocrinology
Endocrine Abstracts (2002) 4 OC22

SFE2002 Oral Communications Steroid hormone action (8 abstracts)

Agonist activated adrenocorticotropin receptor internalises via a clathrin-mediated G-protein coupled receptor kinase dependent mechanism

AH Baig 1 , L Hunyady 2 & AJL Clark 1


1Dept. of Endocrinology, Queen Mary University of London, London, UK; 2Dept. of Physiology, Semmelweis University, Budapest, Hungary.

The physiological effects of the pituitary hormone, adrenocorticotropic hormone (ACTH) on the adrenal are mediated by the melanocortin 2 receptor (MC2R), a G protein coupled receptor (GPCR) that signals via adenylate cyclase to elevate intracellular cyclic AMP (cAMP) levels. Therefore the expression and function of the receptor are likely to be the major determinants of the response to ACTH. Following repeated stimulation, the cAMP signal is diminished or desensitised. Desensitisation of receptors is known to occur due to phosphorylation by specific protein kinases. We have previously shown that the MC2R desensitises over a prolonged time and this effect is largely controlled by cAMP dependent protein kinase A (PKA). Prolonged desensitisation may involve internalisation of the receptor, which is also a known mechanism for the regulation of signalling. Internalisation occurs by two mechanisms, receptor mediated endocytosis (via clathrin coated pits) and caveolae mediated internalisation. The mode of internalisation for the endogenous MC2R in Y1 cells was determined by acid wash studies using radiolabelled ACTH, in which the degree of internalisation was determined over time. Treatment of Y1 cells with hypertonic sucrose, which acts as a clathrin inhibitor, was able to block internalisation suggesting that the MC2R is able to internalise via a clathrin mediated mechanism. Dominant negative (DN) mutant forms of beta-arrestin-1, dynamin-1 and G-protein coupled receptor kinase 2 (GRK2) were stably transfected in Y1 cells. Internalisation was then determined for these cell lines. The DN -arrestin and DN -dynamin forms did not inhibit internalisation whereas the DN- GRK2 was able to do so, suggesting that this kinase is involved in internalisation. The PKA inhibitor, H89 had no effect on internalisation, suggesting PKA is not involved. Therefore it is possible to suggest that PKA and GRKs have distinct roles in signalling regulation of the MC2R.

Volume 4

193rd Meeting of the Society for Endocrinology and Society for Endocrinology joint Endocrinology and Diabetes Day

Society for Endocrinology 

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