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UBC Theses and Dissertations

Characterization of a novel androgen membrane receptor in lampreys that may regulate sexual development : androgen binding in lampreys may have implications for steroid ligand and receptor evolution in vertebrates Didier, David Wesley


Lampreys are basal vertebrates, and their physiology may provide insight into the evolution of physiological systems in vertebrates. To date only progestin, corticoid and estrogen receptors, have been identified in the sea lamprey, Petromyzonmarinus. This is remarkable because 1) more derived vertebrates have evolved six nuclear receptors; 2) if androgen nuclear receptors are absent how might male sexual development be regulated in this ancient group of fishes; 3) androgens have been identified in lampreys so the lack of a nuclear androgen receptor may suggest alternative signaling pathways. I test the hypothesis that lampreys have an active androgen steroid receptor. This hypothesis led to three predictions: first that lamprey synthesize the androgens dehydroepiandrosterone (DHEA) and androstenedione (Ad), second that the master sex hormone gonadotropin releasing hormone regulates androgen synthesis in the testes of male lamprey, and third that androgen signaling in the testes occurs via a novel receptor. Using high performance liquid chromatography, thin layer chromatography and radio-immunoassay I demonstrated the presence of the androgenic steroids DHEA and Ad in the circulation and tissues of sea lamprey and Pacific lamprey. Further, incubation of lamprey testes with lamprey specific GnRH I and III resulted in promoting the conversion of DHEA to Ad. Finally I have demonstrated that androstenedione binds to a membrane fraction isolated from the testes of Pacific lamprey, Entosphenus tridentatus, testes suggesting the presence of a putative androgen membrane receptor (mAR). The binding characteristics indicate a high-affinity (Kd = 7.548 +/- 1.455 nM, R​² = 0.9804,) low capacity (Bmax = 0.0.2366 +/- 0.01345 nM/mg of protein), single binding site androgen receptor. The association rate was determined by non-linear analysis to be 10.2 +/- 3.2 min, with maximum binding achieved at approximately 30 min. The dissociation rate was similar: 9.5 +/- 5.0 min, with maximum binding achieved at approximately 30 min. A partial identification of the receptor was achieved through the use of an affinity column and liquid chromatography. Identification of a novel androgen receptor may point to a novel evolutionary pathway for androgen signaling in vertebrates. It’s significant as it may indicate that this pathway is an ancestral state.

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