Gonadotropin releasing hormone (GnRH) regulates the reproductive system through a specific G-protein-coupled receptor (GPCR) in pituitary gonadotropes. The existence of two (or more) forms of GnRH in most vertebrates suggested the existence of GnRH receptor subtypes (I and II). Using sequence information for extracellular loop 3 of a putative Type II GnRH receptor from a reptile species, we have looked for a Type II GnRH receptor gene in the human genome EST (expressed sequence tag) database. A homolog was identified which has 45% and 41% amino acid identity with exons 2 and 3 of the known human GnRH pituitary receptor (designated Type I) and much lower homology with all other GPCRs. A total of 27 contiguous ESTs was found and comprised a continuous sequence of 1642 nucleotides. The EST sequences were confirmed in the cloned human gene and in PCR products of cDNA from several tissues. All EST transcripts detected were in the antisense orientation with respect to the novel GnRH receptor sequence and were highly expressed in a wide range of human brain and peripheral tissues. PCR of cDNA from a wide range of tissues revealed that intronic sequence equivalent to intron 2 of the Type I GnRH receptor was retained. The failure to splice out putative intron sequences in transcripts which spanned exon-intron boundaries is expected in antisense transcripts, as candidate donor and acceptor sites were only present in the gene when transcribed in the orientation encoding the GnRH receptor homolog. No transcripts extended 5' to the sequence corresponding to intron 2 of the Type I GnRH as the antisense transcripts terminated in poly A due to the presence of a polyadenylation signal sequence in the putative intron 2 when transcribed in the antisense orientation. These findings suggest that a Type II GnRH receptor gene has arisen during vertebrate evolution and is also present in the human. However, the receptor may have become vestigial in the human, possibly due to the abundant and universal tissue transcription of the opposite DNA strand to produce antisense RNA.
R Millar, D Conklin, C Lofton-Day, E Hutchinson, B Troskie, N Illing, SC Sealfon and J Hapgood
B. E. CLAYTON, J. S. M. HUTCHINSON, R. D. HYDE, D. R. LONDON, I. H. MILLS and F. T. G. PRUNTY
Thirty-five male patients with hypogonadism (excluding those suffering from complete hypopituitarism) have been studied. A system of classification is outlined based on the following features: (1) the clinical picture; in particular, evidence for androgen deficiency, (2) the histology of the Leydig cells, (3) the function of the seminiferous tubules as assessed by histology and sperm counts in semen, and (4) the excretion of gonadotrophins. The patients have been classified into those with primary testicular failure and those with failure of pituitary gonadotrophin secretion; the former group includes the patients with Klinefelter's syndrome. Difficulties arose in deciding when gonadotrophin excretion was definitely subnormal because the values in the lower part of the normal range overlap some of those from patients who were obviously deficient in pituitary gonadotrophin.
Some patients showed evidence of hypothyroidism, probably related to testosterone deficiency.
The excretion of 17-oxosteroids is shown to be a poor guide to the assessment of androgen deficiency. This is partly due to the secretion of adrenal androgen, which may be greater than normal in some patients with Leydig-cell deficiency. Patients with primary Leydig-cell failure responded to stimulation by corticotrophin (ACTH) with a relatively greater increase in 17-oxosteroids than normal. Patients with deficiency of pituitary gonadotrophin responded to ACTH stimulation with a much smaller increase in 17-oxosteroid excretion than normal individuals. The evidence presented suggests that a mechanism exists for the integration of androgen production by both gonad and adrenal.