Highly Heterologous Region in the N-Terminal Extracellular Domain of Reptilian Follitropin Receptors Yasuhisa Akazome, Osamu Ogasawara, Min K. Park, and Takao Mori Laboratory of Endocrinology, Department of Biological Sciences, Graduate School of Science, University of Tokyo, Bunkyo-ku, Tokyo 113, Japan Accepted August 7, 1996 The primary structure of the N-terminal extracellular region of the follitropin receptor (FSH-R), which is thought to be responsible for hormone binding specific- ity, was determined in three reptilian species (tortoise, gecko, and lizard). Remarkably low sequence homolo- gies were detected in the C-terminal part of the extracel- lular domain. This region was estimated to be a part of exon 10, which is the last exon of the FSH-R gene. In this region, not only were low homologies detected among the three reptilian species, but also specific deletions and/or insertions were found. In particular, large dele- tions were detected in squamate (gecko and lizard) FSH-Rs. Phylogenetic analysis indicated that these large deletions occurred recently, i.e., after the Triassic period. In another region characterized, sequence homologies were high, with tortoise–rat homology 78.4%, gecko–rat 64.7%, and lizard–rat 69.1%. In this highly conserved region, however, some reptile-specific alterations were detected, such as the loss of a cysteine residue in putative exon 7 and the existence of potential N-linked glycosylation sites in putative exon 9. r 1996 Academic Press, Inc. Follitropin (FSH), one of the pituitary glycoprotein hormones, plays an important role in follicular maturation in the ovary and spermatogenesis in the testis. This hormone is distributed widely throughout vertebrates. Many studies on the binding activity of gonadotropins (FSH and lutropin, LH) to vertebrate gonads have been performed. In general, FSHs are able to bind to gonads in many species of vertebrate, whereas LH binding is more species-specific. In fact, rat FSH affects chick, quail, ostrich, tortoise, lizard, and newt gonads (Ishii, 1988), while pitu- itary FSH preparations derived from ostrich, turkey, turtle, sturgeon, and bullfrog affect estrogen production in the granulosa cells of rat ovary (Dahl et al., 1989). The primary structure of the FSH receptor (FSH-R) has been studied in several species of mammal (rat, Sprengel et al., 1990; human, Minegishi et al., 1991; monkey, Gromoll et al., 1993; sheep, Yarney et al., 1993; horse, Robert et al., 1994; cattle, Houde et al., 1994; pig, Remy et al., 1995) as well as in a bird (Akazome et al., 1996). Based on these reports, other vertebrate FSH-Rs are thought likely to be highly homologous with mammalian and avian FSH-Rs. We performed RT-PCR cloning of reptilian FSH-Rs to determine the primary structures of three species, tortoise, gecko, and lizard. MATERIALS AND METHODS Animals A male tortoise Geoclemys reevesii was obtained from a commercial source. Male geckoes Gekko japonicus and a male lizard Eumeces latiscutatus were captured near the campus. PCR Cloning of Reptilian FSH-Rs Testicular RNA samples were extracted from adult tortoise, gecko, and lizard testes by acid guanidinium General and Comparative Endocrinology 104, 374–381 (1996) Article No. 0183 374 0016-6480/96 $18.00 Copyright r 1996 by Academic Press, Inc. All rights of reproduction in any form reserved.