Characterization of Insulin and Insulin-Like Growth Factor-I Ovarian Receptors during the Reproductive Cycle of Carp (Cyprinus Carpio)1

Abstract

Insulin and insulin-like growth factor-I (IGF-I) receptors were characterized in glycoprotein fractions prepared by wheat germ agglutinin-agarose affinity chromatography from the ovaries of carp. Insulin-specific overall binding in carp ovaries was 6- to 11-fold lower than IGF-I binding (2.7 +/- 0.48% vs. 22.8 +/- 3.6% per 20 microg glycoprotein). Cold IGF-I displaced radiolabeled IGF-I binding in doses 1000- to 3000-fold lower than cold insulin. On the other hand, cold insulin displaced radiolabeled insulin binding at concentrations 5- to 30-fold lower than cold IGF-I. The alpha-subunit molecular masses of carp insulin and IGF-I receptors were smaller than the alpha-subunit molecular mass of rat insulin receptor (125 and 120 vs. 135 kDa, respectively). Autophosphorylation of carp beta-subunit insulin and IGF-I receptors showed similar molecular masses that did not differ from the molecular mass of rat insulin beta subunit. Receptor tyrosine kinase activity was stimulated in a dose-dependent manner by insulin and IGF-I. Insulin and IGF-I stimulated tyrosine kinase activity and reached a maximum, respectively, of 224 +/- 14% and 279 +/- 7% of basal phosphorylation. Insulin and IGF-I binding characteristics were measured through different stages of follicular development. High specific binding of both peptides in primary oocyte growth (5.6 +/- 0.8% and 50 +/- 10% per 20 microg glycoprotein for insulin and IGF-I, respectively) decreased to a minimum at the end of vitellogenesis, followed by a slight increase later, in the preovulatory stage. The presence of insulin and IGF-I receptors in carp ovaries and the changes in percentage of binding throughout the reproductive cycle suggest that, in carp, the roles of insulin and IGF-I depend on the ovarian maturation stage.

Releasing prophase arrest in zebrafish oocyte: synergism between maturational steroid and Igf1

“…This provides evidence in favour of specificity of different receptor-mediated signalling cascades at the oocyte surface. While Igf1 receptor has been purified and high affinity Igf1 binding sites were demonstrated in amphibian oocytes as well as in carp ovary (Hainaut et al 1991, Maestro et al 1997, the possibility of all four Igf ligands acting through Igf1r in zebrafish oocytes cannot be ruled out at this moment since transcripts for two Igf1 receptor isoforms (igf1ra and igf1rb) have been detected earlier (Nelson & Van Der Kraak 2010b). Moreover, immunoblot analysis reveal that while Igf1 triggers a robust increase in Akt phosphorylation (Ser473), DHP stimulation also promotes a significant increase in Akt activation as early as 15 min of in vitro incubation.…”

“…While Igfs alone can induce GVBD, Igf2 has been shown to up-regulate mPRa and resumption of meiosis providing evidence behind integration of MIS and Igf signalling (Picha et al 2012). Moreover, isolation of Igf1r and detection of high affinity Igf1 binding sites in the carp ovary (Maestro et al 1997), coupled with expression of two different Igf receptor isoforms (Igf1ra and Igf1rb) in zebrafish oocyte (Nelson & Van Der Kraak 2010b) further strengthens the possibility of direct action of Igf on oocyte membrane and regulation of final OM (Reinecke 2010). Although gonadotropin regulation of major follicular events, e.g., biosynthesis of maturational steroid as well as Igf expression might involve common cellular signalling cascades, such as cAMP/PKA and/or MEK/MAPK (Nagahama & Yamashita 2008, Li et al 2011, much less is known about interplay between these pro-maturational factors in teleost ovary.…”

Molecular cloning, identification and characterization of four distinct receptor subtypes for insulin and IGF-I in Japanese flounder, Paralichthys olivaceus

“…The -subunits contain single transmembrane domains and highly conserved cytoplasmic tyrosine kinase domains (Cheatham & Kahn 1995, LeRoith et al 1995. In teleosts, ligand-binding studies using mammalian IGF-I and insulin have consistently demonstrated the presence of high-affinity binding sites on various membranes prepared from teleost tissues (Drakenberg et al 1993, Maestro et al 1997. The binding of IGF-I in teleost muscle ectotherms was abundant and at higher affinity than that of insulin, although IGF-IR mRNA was expressed at the lowest levels in rainbow trout muscle (Greene & Chen 1999a,b).…”

Identification and Transcriptional Modulation of the Largemouth Bass, Micropterus salmoides, Vitellogenin Receptor During Oocyte Development by Insulin and Sex Steroids1

“…Minimal information exists describing normal seasonal plasma levels of INS in teleosts, and few studies have revealed that circulating levels fluctuate during the reproductive cycle, presenting the highest concentrations prior to vitellogenesis [60]. The precise functions of INS during these time frames, particularly during primary ovarian growth, have been described by measuring DNA synthesis and binding receptor studies [61,62]. INS and INSlike growth factor-I (IGF-I) have been shown to bind to fish ovaries primarily in previtellogenic oocytes of carp [62], and recombinant human IGF-I increased the diameter of shortfinned eel previtellogenic oocytes [22].…”

“…The precise functions of INS during these time frames, particularly during primary ovarian growth, have been described by measuring DNA synthesis and binding receptor studies [61,62]. INS and INSlike growth factor-I (IGF-I) have been shown to bind to fish ovaries primarily in previtellogenic oocytes of carp [62], and recombinant human IGF-I increased the diameter of shortfinned eel previtellogenic oocytes [22]. These studies support a role for INS in directing primary growth events in developing oocytes, and our data further suggest a specific role for controlling vtgr mRNA production during this window.…”