Peripheral insulin resistance and impaired insulin action are the primary characteristics of type 2 diabetes. The first observable defect in this major disorder occurs in muscle, where glucose disposal in response to insulin is impaired. We have developed a transgenic mouse with a dominant-negative insulin-like growth factor-I receptor (KR-IGF-IR) specifically targeted to the skeletal muscle. Expression of KR-IGF-IR resulted in the formation of hybrid receptors between the mutant and the endogenous IGF-I and insulin receptors, thereby abrogating the normal function of these receptors and leading to insulin resistance. Pancreatic ␤-cell dysfunction developed at a relative early age, resulting in diabetes. These mice provide an excellent model to study the molecular mechanisms underlying the development of human type 2 diabetes.

Early neurogenesis progresses by an initial massive proliferation of neuroepithelial cells followed by a sequential differentiation of the various mature neural cell types. The regulation of these processes by growth factors is poorly understood. We intend to understand, in a well-defined biological system, the embryonic chicken retina, the role of the insulin-related growth factors in neurogenesis. We demonstrate the local presence of signaling elements together with a biological response to the factors. Neuroretina at days 6-8 of embryonic development (E6-E8) expressed proinsulin/ insulin and insulin-like growth factor I (IGF-I) mRNAs as well as insulin receptor and IGF type I receptor mRNAs. In parallel with this in vivo gene expression, E5 cultured neuroretinas synthesized and released to the medium a metabolically radiolabeled immunoprecipitable insulin-related peptide. Furthermore, insulin-related immunoreactive material with a HPLC mobility close to that of proinsulin was found in the E6-E8 vitreous humor. Exogenous chicken IGF-I, human insulin, and human proinsulin added to E6 cultured neuroretinas showed relatively close potencies stimulating proliferation, as determined by [methyl-3H]thymidine incorporation, with a plateau reached at 10-8 M. These factors also stimulated neuronal differentiation, indicated by the expression of the neuron-specific antigen G4. Thus, insulin-related growth factors, interestingly including proinsulin, are present in the developing chicken retina and appear to play an autocrine/paracrine stimulatory role in the progression of neurogenesis.Neurogenesis, as many other developmental processes, is likely to be regulated by the orchestrated action of multiple growth factors present in the cellular microenvironment. This results in an equilibrium of events, cell proliferation-cell differentiation-cell survival, that leads to the highly complex cell diversity and cytoarchitecture of the mature central nervous system (CNS). Neural-specific, as well as multifunctional, diffusible factors are involved in the regulation of these processes (1-3).The insulin family of hormones/growth factors (referred to here as insulin-related growth factors) includes in vertebrates insulin, its precursor proinsulin, and insulin-like growth factors I and II (IGF-I and IGF-II). The genes for the IGFs and the two structurally related signaling receptors, the insulin receptor (IR) and the IGF type I receptor (IGF-IR), are expressed in the developing CNS of several species (4, 5). While the biological relevance of IGF-I in neural development is supported by multiple studies on neural primary cultures and recent knockout mice models (5-8), the CNS synthesis and possible physiological role of proinsulin/insulin [here referred to as (pro)insulin] are debatable issues (5, 9).The avian neuroretina is a well-characterized part of the CNS that is suitable for early developmental studies, in vivo and in vitro (Fig.

Insulin and insulin-like growth factor (IGF-I) receptors are heterotetrameric proteins consisting of two ␣-and two ␤-subunits and members of the transmembrane tyrosine kinase receptors. Specific ligand binding to the receptor triggers a cascade of intracellular events, which begins with autophosphorylation of several tyrosine residues of the ␤-subunit of the receptor. The triple cluster in the tyrosine kinase domain of the ␤-subunit is the earliest and major autophosphorylation site. Previous studies have shown that substitutions of these three tyrosines by phenylalanines of both insulin and IGF-I receptors practically abolish any activation of cellular signaling pathways. We have studied the effect of double tyrosine mutations on IGF-I-induced receptor autophosphorylation, activation of Shc and IRS-1 pathways, and cell proliferation and tumorigenicity. Substitution of tyrosines 1131/1135 blocks any detectable autophosphorylation, whereas substitution of tyrosines 1131/1136 or 1135/1136 only reduces autophosphorylation levels in some clones by ϳ50%. Nevertheless, all the cells expressing IGF-I receptors with double tyrosine substitutions demonstrated markedly reduced signaling through Shc and IRS-1 pathways. In addition, they were unable to respond to IGF-I-stimulated cell growth in culture, and tumor formation in nude mice was abrogated. These data suggest that the presence of tyrosine 1131 or 1135 essential for receptor autophosphorylation, whereas the presence of each of these tyrosines is necessary for a fully functional receptor.The multiple physiological actions, including cell growth and differentiation of the insulin-like growth factors (IGFs) 1 are mediated by the IGF-I receptor. While the IGF-I receptor and the structurally related insulin receptor are members of the type II receptor tyrosine kinase family, their in vivo biological functions are quite separate. Both the IGF-I and insulin receptors are heterotetrameric proteins composed of two extracellular ␣-subunits and two membrane-spanning ␤-subunits linked by disulfide bonds (1-3). Sequences found in the ␣-subunits of each receptor are important for determining ligand specificity. The amino-terminal and carboxyl-terminal portions of the ␣ subunit of the insulin receptor are critical for high affinity insulin binding, while the cysteine-rich domain of the IGF-I receptor determines high affinity IGF-I binding (4 -6). Likewise, the ␤-subunits contain a number of structurally distinct domains including the extracellular, transmembrane, juxtamembrane, tyrosine kinase, and carboxyl-terminal regions. Binding of ligand to the ␣-subunit activates the tyrosine kinase activity of the ␤-subunit resulting in autophosphorylation on distinct tyrosine residues. The triple tyrosine cluster within the kinase domain (1131, 1135, and 1136 tyrosines in the IGF-I receptor and the equivalent residues in the insulin receptor; numbering system of Ullrich et al. (2)) is the earliest and major site of autophosphorylation. Phosphorylation of these three tyrosine residues is...