Structural features of mammalian gonadotropins

Department of Biological Sciences, Wichita State University, KS 67260-0026, USA.
Molecular and Cellular Endocrinology (Impact Factor: 4.41). 01/1997; 125(1-2):3-19. DOI: 10.1016/S0303-7207(96)03945-7
Source: PubMed


There are two species for which both pituitary and placental gonadotropins are readily available, humans and horses. The human gonadotropins are better characterized than equine gonadotropins. Nevertheless, the latter are very interesting because they provide exceptions to some of the general structure-function principles derived from studies on human and other mammalian gonadotropins. For example, separate genes encode the hLH beta and hCG beta subunits while a single gene encodes eLH beta and eCG beta. Thus, eCG and eLH differ only in their oligosaccharide moieties and eLH is the only LH that possesses the O-glycosylated C-terminal extension previously believed to be restricted to chorionic gonadotropins. Truncation experiments involving eLH beta and hCG beta have suggested the C-terminal extension has no effect on receptor binding. However, the largest of three eCG forms which differ only in the extent of O-glycosylation possessed reduced affinity for LH and FSH receptors. This result suggested that effects of O-glycosylation need to be considered when examining the glycosylation differences between eLH and eCG responsible for the 10-fold lower eCG receptor binding affinity compared with that of eLH. Contribution of alpha Asn56 N-linked oligosaccharides to the different biological activities of eLH and eCG has been evaluated following selective removal using peptide-N-glycanase digestion of native equine alpha-subunit preparations. Hormones-specific patterns of glycosylation were observed on alpha Asn56 of eLH, eFSH, and eCG. Removal of alpha Asn56 oligosaccharides increased the rate of subunit association, the extent of association, and receptor binding activity. Some unassociated alpha-subunit oligosaccharides were identified which may interfere with subunit association because they were more abundant in unassociated subunit oligosaccharide maps than in a total oligosaccharide map. This was most striking in the case of eCG alpha in which two minor peaks became the major oligosaccharide peaks detectable in the unassociated eCG alpha fraction following association with eLH beta and eFSH beta. The biological activities exhibited by hybrid hormones, eLH alpha reassociated with oLH beta and pLH beta, found to be greater than those of oLH and pLH provided an interesting exception to the general rule that the beta-subunit determines the potency of the heterodimer. LH receptor binding activities of eLH beta-chimeric ovine/equine alpha-subunits suggested that the equine alpha-subunit N-terminal domain may be responsible for this effect. Equine FSH has higher FSH receptor binding activity than human, ovine, and porcine FSH preparations. This probably results from two factors. First, the presence of the equine alpha-subunit promotes receptor binding as noted above. Second, the overall -2 charge of the eFSH beta determinant loop, which is less negative that the -3 observed in other species, results from the presence of an Asn residue at position 88 instead of Asp. This apparently facilitates binding to the FSH receptor.

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    • "Both subunits contain N-linked heterogeneous oligosaccharide structures that play an important role in many of the functional characteristics of the molecule, including the metabolic fate of the hormone and the interaction with its cognate receptor (Bishop et al., 1994; Helenius and Aebi, 2004; Matzuk and Boime, 1988a, 1988b, 1989; Matzuk et al., 1990; Muyan and Boime, 1998; Sairam, 1989; Sairam and Bhargavi, 1985; Smith et al., 1990; Ulloa-Aguirre et al., 1999). Numerous carbohydrate intermediates formed through posttranslational processing eventually become final forms of the glycans attached to the protein core, and conform the repertoire of distinct oligosaccharides structures that characterize all glycoprotein hormones (Baenziger and Green, 1988; Bousfield et al., 1996; Ulloa-Aguirre et al., 1995). Further, differential terminal glycosylation allows fine-tuning of the protein properties at the target cell without the need for a change in primary 0016-6480/$ -see front matter Ó 2009 Published by Elsevier Inc. doi:10.1016/j.ygcen.2008.12.021 "
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    ABSTRACT: Men with insulinopenic diabetes mellitus frequently present hypogonadism and exhibit circulating luteinizing hormone (LH) molecules with increased biological activity. To further study this latter issue, we analyzed the pattern of isoform distribution and the impact of changes in terminal glycosylation of pituitary LH on the bioactivity of this gonadotropin in experimental diabetes. Adult male rats were treated with streptozotocin or vehicle and euthanized on days 30, 60, or 90 posttreatment. All diabetic groups exhibited a significant decrease in serum insulin and testosterone levels as well as in sperm count; serum gonadotropins and 17beta-estradiol decreased only after 90 days of insulinopenia. Both the immunoreactive concentrations and the biological to immunological ratio of intrapituitary LH significantly increased in all experimental groups, as assessed by an in vitro homologous bioassay in HEK-293 cells expressing a recombinant LH receptor. Chromatofocusing of pituitary extracts revealed the presence of multiple LH charge isoforms; the pH distribution profile of LH in diabetic and control rats was indistinguishable on days 30 and 60 posttreatment. By contrast, the abundance of more basic isoforms (pH 9.99-9.0) decreased and that of isoforms with pH values 8.99-8.0 increased in rats with long-standing diabetes compared to controls. It is concluded that experimental diabetes alters the function of the pituitary-testicular axis, resulting in reduced sex steroids levels and hypogonadotropism. Long-standing insulinopenia leads to a paradoxical accumulation of intrapituitary LH molecules enriched in bioactivity with altered terminal glycosylation, which are apparently subserved by distinct mechanisms involving altered hypothalamic and/or gonadal inputs on the gonadotrope.
    General and Comparative Endocrinology 06/2009; 161(3):304-12. DOI:10.1016/j.ygcen.2008.12.021 · 2.47 Impact Factor
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    • "We do not know the influence of oligosaccharide composition on the overall charge of New World primate FSH, but putative N-linked glycosylation sites at positions 7 and 24 (NetNGlyc 1.0 Server) and twelve cysteines are conserved among species. FSHβ is not thought to undergo O-linked glycosylation (Bousfield et al., 1996). "
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    ABSTRACT: The goal of this study was to characterize the gonadotropins expressed in pituitary glands of the New World squirrel monkey (Saimiri sp.) and owl monkey (Aotus sp.). The various subunits were amplified from total RNA from squirrel monkey and owl monkey pituitary glands by reverse transcription-polymerase chain reaction and the deduced amino acid sequences compared to those of other species. Mature squirrel monkey and owl monkey glycoprotein hormone alpha-polypeptides (96 amino acids in length) were determined to be 80% homologous to the human sequence. The sequences of mature beta subunits of follicle stimulating hormone (FSHbeta) from squirrel monkey and owl monkey (111 amino acids in length) are 92% homologous to human FSHbeta. New World primate glycoprotein hormone alpha-polypeptides and FSHbeta subunits showed conservation of all cysteine residues and consensus N-linked glycosylation sites. Attempts to amplify the beta-subunit of luteinizing hormone from squirrel monkey and owl monkey pituitary glands were unsuccessful. Rather, the beta-subunit of chorionic gonadotropin (CG) was amplified from pituitaries of both New World primates. Squirrel monkey and owl monkey CGbeta are 143 and 144 amino acids in length and 77% homologous with human CGbeta. The greatest divergence is in the C terminus, where all four sites for O-linked glycosylation in human CGbeta, responsible for delayed metabolic clearance, are predicted to be absent in New World primate CGbetas. It is likely that CG secreted from pituitary of New World primates exhibits a relatively short half-life compared to human CG.
    General and Comparative Endocrinology 03/2008; 155(3):534-41. DOI:10.1016/j.ygcen.2007.08.004 · 2.47 Impact Factor
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    • "Stimulation of graft Leydig cells by mouse LH leads to production of bioactive testosterone and establishment of a feedback system between the graft and the mouse pituitary (Honaramooz et al. 2002). Differences between mouse and donor species gonadotropic hormones (Bousfield et al. 1996) may lead to inefficient interaction between the murine gonadotropins and the donor testis tissue, thus leading to insufficient amounts of hormones interacting with the grafted donor tissue. It is possible, therefore, that supplementation of host mice with exogenous gonadotropins could enhance spermatogenesis in testicular xenografts. "
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    ABSTRACT: Grafting of testis tissue from immature animals to immunodeficient mice results in complete spermatogenesis, albeit with varying efficiency in different species. The objectives of this study were to investigate if grafting of horse testis tissue would result in spermatogenesis, and to assess the effect of exogenous gonadotropins on xenograft development. Small fragments of testis tissue from 7 colts (2 week to 4 years of age) were grafted under the back skin of castrated male immunodeficient mice. For 2 donor animals, half of the mice were treated with gonadotropins. Xenografts were analyzed at 4 and 8 months post-transplantation. Spermatogenic differentiation following grafting ranged from no differentiation to progression through meiosis with appearance of haploid cells. Administration of exogenous gonadotropins appeared to support post-meiotic differentiation. For more mature donor testis samples where spermatogenesis had progressed into or through meiosis, after grafting an initial loss of differentiated germ cells was observed followed by a resurgence of spermatogenesis. However, if haploid cells had been present prior to grafting, spermatogenesis did not progress beyond meiotic division. In all host mice with spermatogenic differentiation in grafts, increased weight of the seminal vesicles compared to castrated mice showed that xenografts were releasing testosterone. These results indicate that horse spermatogenesis occurs in a mouse host albeit with low efficiency. In most cases, spermatogenesis arrested at meiosis. The underlying mechanisms of this spermatogenic arrest require further investigation.
    Reproduction (Cambridge, England) 07/2006; 131(6):1091-8. DOI:10.1530/rep.1.01101 · 3.17 Impact Factor
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