OVARY-SELECTIVE KNOCKOUT OF IGF-I

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A body of information relevant to the rodent documents a complete (ovarian) intrafollicular insulin-like growth factor (IGF)-I system, replete with a ligand (IGF-I), a receptor (type I), IGF binding proteins (IGFBPs 4&5) and IGFBP-4 and 5-directed endopeptidases. According to current views, shaped in large measure by numerous in vitro observations, the primary and presumably obligatory autocrine role of bioavailable intrafollicular IGF-I is the amplification of FSH action in granulose cells. It is this FSH-amplifying property of IGF-I which underlies the hypothesis that intrafollicular IGF-I constitutes an obligatory determinant of antral (FSH-dependent) follicular development. Rigorous validation of this so-called "amplification" hypothesis requires demonstration of the in vivo indispensability of intrafollicular IGF-I. Although female null mutants for the igf-1 gene were reported to display reproductive dysfunction, these preliminary observations remain inconclusive given the high (<90%) neonatal mortality rate and the small (<6) number of the mice studied. Moreover, the employment of systemic (as opposed to ovary-specific) gene targeting technology precluded the determination of the relative role of circulating as opposed to intraovarian (locally-generated) IGF-I in the maintenance of ovarian function. To provide unequivocal evidence for or against the "amplification" hypothesis, they propose a series of complementary in vivo approaches designed to effect ovary (granulose cell)-selective "knockout" of IGF-I. In all cases, use will be made of the promoter of mouse alpha-inhibin, chosen for its proven in vivo ability to strongly drive transgene expression in an ovary-selective fashion. Specifically, they propose to generate and characterize mice engineered to overexpress rat IGFBP-1 and thus to sequester/deplete bioavailable intrafollicular IGF-I. Mice bearing Cre/loxP-driven granulose cell-selective igf-1 gene deletion; and mice bearing Tamoxifen-inducible, Cre/loxP-driven granulose cell-selective igf-1 gene deletion. Preliminary data include generation and validation of alpha-inhibin/IGFBP-1 and alpha-inhibin/Cre transgenics, successful "floxing" of the ifg-1 gene, and the generation of viable and fertile ifg-1-"floxed" mutants. Insight derived from this investigation may result in the confirmation or rejection of IGF-I as an obligatory intraovarian amplifier; and development of novel widely-applicable transgenic reagents designed to effect ovary-selective deletion of a gene of interest.