Multiple components of the insulin-like growth factor (IGF) system have been shown by gene targeting to be required for normal growth and differentiation in the mouse. The major goal of this proposal is to determine the in vivo role during development of the six-member IGF binding protein (IGFBP) family, which is thought to be a major regulator of the IGF system at multiple levels. It is generally thought that circulating IGFBPs primarily function to inhibit IGF action, while tissue-associated IGFBPs can enhance or attenuate IGF action and potentially subserve additional roles. They have produced by gene targeting mice that lack IGFBP-2, a prominent fetal IGFBP, and have shown that these mice exhibit a reduced adult spleen size. In addition, the serum of adult mice homozygous for the IGFBP-2 mutation has increased levels of several additional IGFBPs that themselves are expressed by early post-implantation development. To extend their functional analysis of the IGFBP system, they will produce by gene targeting mice lacking functional copies of four additional IGFBPs (BPs-- 3, -4, -5, and -6) that each have distinct spatial and temporal expression patterns in the embryo that suggest prospective developmental roles in multiple organ systems. Developmental deficits in mutant mice will be identified; if individual homozygous mutants are viable, postnatal growth, organ histopathology and endocrine status will he evaluated with specific emphasis on analysis of the connective tissue and urogenital systems, sites that would appear to be particularly sensitive to IGFBP mutation. As mutant lines are produced, combinatorial mating to produce strains defective in several IGFBPs will be initiated with the expectation that additional required roles for this gene family during development will emerge. They also propose to continue to characterize the IGFBP-2 mutation in combination with other IGF system mutants. They will produce mice doubly-homozygous for the IGFBP-2 mutation and other alleles (including a null allele for H19) that increase the circulating levels of IGF-2 either pre- or post-natally. They will determine whether these genetic challenges in the absence of IGFBP-2 alter growth, further increase expression of the other IGFBPs, or reverse the deficit in adult spleen size of homozygous IGFBP-2 mice. These studies will clarify the specific role of IGFBP-2 as well as additional IGFBPs in regulating IGF-2 activity. Taken together, the proposed experiments will determine the role of the IGFBP system in prenatal growth and in development of several organ systems, test the hypothesis that IGFBPs functionally compensate for each other during development, and produce new lines of mutant mice lacking specific IGFBPs that will be useful models for developmental and endocrine manipulation.
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