In respect to growth, the adrenal is a dynamic organ that requires constant stimulation from peptides derived from the ACTH precursor pro-opiomelanocortin (POMC) to maintain its tonic state. Previous studies have suggested that the adrenal mitogen resides in the N-terminus of pro-gamma-MSH, which upon cleavage results in the generation of shorter mitogenic peptides not containing the gamma-MSH sequence. This hypothesis has recently been substantiated by our characterisation of a serine protease expressed by the adrenal that cleaves the peptide leading to the identification of the adrenal mitogen as N-POMC (1-52) (1).
We have now extended these studies to identify the receptor through which this peptide elicits its mitogenic activity. Since it has previously be shown that N-POMC (1-28) has mitogenic properties we attempted to use this peptide linked to solid phase to affinity purify the receptor from solubilised membranes prepared from the Y1 adrenal cortical tumour cell line. Using this approach we identified a single protein species of approx 80 KDa which following trypsinisation and N-terminal sequencing will allow the cloning of the gene encoding the receptor. To further investigate the actions of N-POMC peptides we stimulated Y1 cells with N-POMC (1-28) that had been transfected with one of the vectors from the Mercury Pathway Profiling System (Clontech). These vectors contain a specific cis acting enhancer element coupled to a reporter gene that allow the effect of a compound on specific signalling pathways to be investigated. The results from this study suggest that stimulation with N-POMC (1-28) results in the activation of Activator protein 1 (AP1), N_kappaB and Serum response element (SRE). These results provide a platform for further investigation into the signalling pathways of the N-POMC peptides and provide a useful quantitative measure for the characterisation of the receptor.
(1) Bicknell, A.B. et al (2001) Cell 105 903-912.
03 - 04 Dec 2001
Society for Endocrinology