Polymorphisms at the 5' end of the human gonadotropin-releasing hormone receptor gene are not associated with the timing of menarche in Japanese girls

Abstract

We read with interest the article by Kakar (1) and the commentary by Miller (2) in the European Journal of Endocrinology, which showed the diversity and species speci®city of the regulatory elements in the gonadotropin-releasing hormone receptor (GnRHR) gene and reviewed the important role of GnRHR gene regulation in the control of the hypothalamic±pituitary±gonadal axis respectively. Thus we hypothesized that polymorphism(s) in the regulatory region in the GnRHR gene could be a genetic factor that in uences the timing of the onset of puberty. By direct sequencing of the human GnRHR gene following PCR ampli®cation of genomic DNA, we identi®ed seven polymorphisms at the 5 end of the gene (Table 1). Allele patterns obtained by direct sequencing and sequencing followed by subcloning of the region containing all seven polymorphisms suggested that the polymorphisms were tightly linked on the same allele. The allele frequency with a 10 bp insertion (allele I) in the 86 healthy Japanese subjects was 0.46, which was determined by PCR ampli®cation of the region and electrophoresis in a 4% agarose gel. The primer pairs used were as follows: PP2-F, 5-CCTAATGAATCAGTCCATCATTAAC-3; PP2-R, 5-TAGGGGAAAAAGATTAGAGCATATC-3. The length of allele I was 198 bp, whereas that of the wild type (allele D) was 188 bp. Then we genotyped the polymorphism in 196 Japanese girls (aged 18±20) in a school of nursing. Ages at menarche were obtained by means of a questionnaire and the heights were measured at the time of blood sampling. Age at menarche and height for the whole group were (means 6 S.D.) 12.2 6 1.1 years and 157.7 6 5.3 cm respectively. These values were consistent with a previous report on the Japanese population (3). The frequency of allele I was 0.47. However, there were no signi®cant differences in the timing of menarche or the height among the three genotypes (I/I, I/D and D/D; tested by one-way analysis of variance) or among the two homozygous genotypes (I/I and D/D; tested by paired t-test). Thus, this high-frequency polymorphic allele at the 5 end of the GnRHR gene is not likely to be associated with the timing of menarche or with ®nal height in Japanese girls. To date, only one polymorphism in the coding region of the GnRHR gene has been reported for a subset of pituitary adenomas (4). As far as the timing of menarche is concerned, there are no reports studying polymorphisms of the genes related to the hypothalamic±pituitary±gonadal axis, though positive associations with CYP17 gene (5) and vitamin D receptor gene (6) polymorphisms have been reported. In this study, we expected that the polymorphism(s) in the 5 end of the GnRHR gene would be one of the strong candidate regions affecting the timing of menarche and, therefore, ®nal height, but the results excluded this possibility.