Regulation of insulin sensitivity by the Src family non-receptor tyrosine kinase,

描述（由申请人提供）：最近，我们的实验室已经开始研究胰岛素受体信号传导与非受体酪氨酸激酶Src、林恩和Fyn之间的串扰。我们已经发现，Fyn基因敲除小鼠显示出显著减少的肥胖，增加的脂肪酸氧化，增强的cateholamine刺激的脂解和改善的胰岛素敏感性。令人惊讶的是，该小鼠在高脂肪饮食下也获得了显著的体重和脂肪组织质量，尽管有显著的血脂异常，但即使与其瘦的野生型对照相比，仍保持高度的葡萄糖耐受性。骨骼肌和脂肪组织氧化的增强伴随着亚基苏氨酸172（活化位点磷酸化）上AMPK磷酸化的增加和其负调控位点上乙酰辅酶A羧化酶磷酸化的增加而发生。脂解速率的增加是由于周脂蛋白磷酸化和激素敏感脂肪酶激活位点磷酸化增加所致。尽管常规Fyn敲除小鼠在能量利用和胰岛素敏感性方面显示出显著的改善，但这些数据并没有解决Fyn功能的丧失是否通过信号传导的急性改变或通过发育适应表现出来。此外，Fyn有三种剪接变体，似乎具有不同的功能和组织特异性分布模式。因此，常规Fyn缺失小鼠的分析不能解决这些问题。因此，我们在该R21申请中提出通过产生同种型组织特异性Fyn转基因表达小鼠和诱导型组织特异性Fyn敲除小鼠来确定这些效应是否以细胞自主与非自主方式发生。然后，我们建议使用敲入技术，以产生骨骼肌和脂肪组织中的同种型特异性Fyn表达。这些动物将通过详细分析调节与全身整合系统生理学相关的代谢信号传导的分子信号传导途径进行检查。糖尿病、肥胖和胰岛素抵抗状态都可以被描述为身体调节能量消耗和输出差异的能力缺陷。我们已经发现，酶Fyn仅在禁食状态下调节脂肪酸氧化，而在进食状态下不调节。这意味着可以调节Fyn活性，从而增加脂肪酸氧化和能量消耗（例如：减少肥胖）。因此，该项目的重点是开发和表征小鼠模型，以直接测试Fyn功能改变的综合生理后果。

DESCRIPTION (provided by applicant): Recently, our laboratory has begun to investigate the cross talk between insulin receptor signaling with that of the non-receptor tyrosine kinases Src, Lyn and Fyn. We have found that Fyn null mice display markedly reduced adiposity, increased fatty acid oxidation, enhanced cateholamine-stimulated lipolysis and improved insulin sensitivity. Surprisingly, this mice also gain substantial weight and adipose tissue mass on a high fat diet, yet remain highly glucose tolerant even compared to their lean wild type controls despite marked dyslipidemia. The enhancement of skeletal muscle and adipose tissue oxidation occurred concomitant with increase AMPK phosphorylation on a subunit threonine 172 (activation site phosphorylation) and with increased acetylCoA carboxylase phosphorylation on its negative regulatory site. The increased rate of lipolysis resulted from increased phosphorylation of perilipin and activation site phosphorylation of hormone sensitive lipase. Although the conventional Fyn null mice display a remarkable improvement in energy utilization and insulin sensitivity, these data do not address whether the loss of Fyn function manifests through acute alterations in signaling or through developmental adaptation. Moreover, there are three splice variants of Fyn that appear to have distinct functional and tissue-specific distribution patterns. Thus, the analysis of the conventional Fyn null mice cannot address these issues. We therefore propose in this R21 application to determine whether these effects occur in a cell autonomous versus non-autonomous manner by generating isoform tissue-specific Fyn transgenic expressing mice and inducible tissue-specific Fyn knockout mice. We then propose to use knock-in technology to generate isoform specific Fyn expression in skeletal muscle and adipose tissue. These animals will be examined by detailed analysis of the molecular signaling pathways regulating metabolic signaling in relationship to whole body integrative system physiology. Diabetes, obesity and insulin resistant states can all be characterized as defects in the body's ability to adjust for differences in energy consumption and output. We have found that the enzyme Fyn regulates fatty acid oxidation only in the fasted state but not in the fed state. This means that it may be possible to modulate Fyn activity and thereby increase fatty acid oxidation and energy expenditure (e.g.: decrease obesity). Thus, this project is focused on developing and characterizing mouse models to directly test the integrative physiologic consequences of altered Fyn function.