Regulation of Akt and glucose transport by prolactin

描述（由申请人提供）：哺乳对新生动物的生存非常重要。催乳素通过调节乳腺上皮细胞向分泌性上皮细胞的分化在此过程中起着关键作用。许多关于催乳素信号转导的研究集中在其在调节乳蛋白基因转录中的作用。然而，由于牛奶含有25 - 30%的脂肪和约3%的乳糖，因此控制葡萄糖可用性和脂质生物合成的途径可能对最佳牛奶生产至关重要。我们以前的研究表明，丝氨酸/苏氨酸蛋白激酶Akt在抑制退化乳腺细胞凋亡中起着关键作用。我们推测催乳素调节葡萄糖转运和脂质生物合成，Akt蛋白激酶是催乳素介导的葡萄糖转运激活所必需的。在这项研究中，我们计划绘制催乳素受体激活Akt和葡萄糖转运所需的区域。我们将集中在磷酸化的酪氨酸残基，因为信号分子的结合，这些网站往往调节其激活，特别是SH 2结构域的分子。我们将使用一种嵌合催乳素受体，其激活可以通过化学二聚化剂控制。这种嵌合受体将在乳腺上皮细胞中表达，以绘制调节葡萄糖转运和Akt激活的区域。将产生在乳腺中表达嵌合催乳素受体的转基因小鼠，以测试特定受体突变体的功能。这些研究将在垂体产生催乳素被阻断从而阻止内源性催乳素受体激活的条件下进行。乳腺发育在Src-/-小鼠中改变，但在Fyn-/-小鼠中没有改变。来源于Src-/-和Fyn-/-小鼠的原代乳腺上皮细胞将用于确定这些激酶在调节Akt活化和葡萄糖转运中的作用。我们还将测试特定信号分子（c-Cbl、Cbl-B、IRS-1、Gab 2、Gab 3、SHP 1和SHP 2）在调节PI 3-激酶/Akt通路和葡萄糖转运中的作用。最后，我们将确定催乳素增加乳腺上皮细胞葡萄糖转运的机制。到目前为止，乳腺上皮细胞中葡萄糖转运的调节机制仍然是一个谜。这些研究应提供有关催乳素在乳腺中的其他作用的重要信息，并确定催乳素调节这些不同过程的机制。

DESCRIPTION (provided by applicant): Lactation is of great importance to survival of newborn animals. Prolactin plays a critical role in this process by regulating the differentiation of mammary epithelial cells into secretory epithelial cells. Much of the research on prolactin signal transduction has focused upon its role in regulating the transcription of milk protein genes. However, since milk is 25 - 30% fat and about 3% lactose, pathways that control glucose availability and lipid biosynthesis are likely to be critical for optimal milk production. Our previous studies indicate that the serine/threonine protein kinase Akt, plays a critical role in suppressing apoptosis in the involuting mammary gland. We hypothesize that prolactin regulates glucose transport and lipid biosynthesis and the Akt protein kinase is required for prolactin-mediated activation of glucose transport. In this grant we propose to map the region(s) of the prolactin receptor that are required for activation of Akt and glucose transport. We will focus on phosphorylated tyrosine residues because the binding of signaling molecules to these sites often regulates their activation, particularly SH2 domain containing molecules. We will use a chimeric prolactin receptor whose activation can be controlled by a chemical dimerizer. This chimeric receptor will be expressed in mammary epithelial cells to map the regions that regulate glucose transport and Akt activation. Transgenic mice will be generated that express the chimeric prolactin receptor in the mammary gland to test the function of specific receptor mutants. These studies will be done under conditions where the production of prolactin by the pituitary is blocked thereby preventing activation of the endogenous prolactin receptor. Mammary gland development is altered in Src-/- mice but not in Fyn-/- mice. Primary mammary epithelial cells derived from Src-/- and Fyn-/- mice will be used to determine the role of these kinases in regulating Akt activation, and glucose transport. We will also test the role of specific signaling molecules (c-Cbl, Cbl-B, IRS-1, Gab2, Gab3, SHP 1, and SHP2) in regulating the PI3-kinase/Akt pathway and glucose transport. Finally, we will determine the mechanisms by which prolactin increases glucose transport in mammary epithelial cells. Until now, the mechanisms by which glucose transport in mammary epithelial cells is regulated have remained a mystery. These studies should provide important information about other roles for prolactin in the mammary gland and identify the mechanisms by which prolactin regulates these diverse processes.