PI 3-Kinase Isoforms in Insulin Action

描述（由申请人提供）：这是NIH资助DK 55545的竞争性更新，重点是PI 3-激酶在胰岛素作用中的作用。PI 3-激酶是胰岛素代谢调控的关键节点，也是胰岛素信号传导的关键分支。使用体外和体内方法，包括RNA干扰（RNAi）和小鼠和细胞系的产生和表征，其中PI 3-激酶的特定亚型已经在全身或组织特异性水平上缺失，即敲除，我们已经证明了这种酶以阳性和阴性方式控制胰岛素信号传导的多种重要方式。这包括PI 3-激酶的各种调节亚基的活性和性质的差异，PI 3-激酶信号传导和胰岛素作用中调节亚基和催化亚基之间化学计量的重要作用，以及PI 3-激酶允许Akt和非典型PKC之间下游信号发散的能力。此外，我们已经证实了疾病状态下PI 3-激酶活性的改变以及该通路与其他信号通路的关系，包括两个以前未被认识到的联系：一个是PI 3-激酶调节亚基与应激激酶JNK和p38的激活之间的联系，这可能将p85亚基与胰岛素抵抗状态下IRS蛋白的丝氨酸磷酸化联系起来;第二个在PI 3-激酶调节亚基和细胞中主要的PIPS磷酸酶PTEN的活性之间。这使我们对PI 3-激酶的重要作用提出了新的假设，PI 3-激酶不仅是胰岛素信号通路的分歧点，而且是生理和病理状态下的正调节和负调节点，以及与其他信号系统，特别是应激激酶的串扰点。在接下来的五年里，我们建议：1）剖析PI 3-激酶调节亚基的多重、差异性作用，（p85 a/B、p50 a和p55 a/AS 53）在胰岛素信号传导中的作用，着重于来自不依赖于PI 3激酶活性的调节亚基的不同N-末端结构域的潜在作用和相互作用; 2）通过组织特异性缺失，确定PI 3-激酶催化亚基p110 a和p110 b在体内不同胰岛素信号转导中的作用;探索新的化学生物学方法来修饰PI 3-通过使用小分子筛选和使用纳米颗粒递送系统的新的体内基因沉默技术，在正常和胰岛素抵抗状态下激酶介导的信号传导。

DESCRIPTION (provided by applicant): This is a competitive renewal of NIH grant DK55545 which is focused on the role of PI 3-kinase in insulin action. PI 3-kinase is important as a critical node in insulin control of metabolism and a key point of divergence of insulin signaling. Using, both in vitro and in vivo approaches, including RNA interference (RNAi) and creation and characterization of mice and cell lines in which specific isoforms of PI 3-kinase have been deleted, i.e. knocked out, at the whole body or tissue specific levels, we have demonstrated multiple, important ways in which this enzyme controls insulin signaling in both positive and negative ways. This includes differences in the activity and properties of the various regulatory subunits of PI 3-kinase, the important role of stoichiometry between regulatory and catalytic subunits in PI 3-kinase signaling and insulin action, and the ability of PI 3-kinase to allow divergence of the downstream signal between Akt and atypical PKCs. In addition, we have demonstrated alterations in PI 3-kinase activity in disease states and the relationship of this pathway to other signaling pathways, including two previously unrecognized connections: one between PI 3-kinase regulatory subunits and activation of the stress kinases JNK and p38, which may link the p85 subunit to serine phosphorylation of IRS proteins in insulin resistant states; and a second between the PI 3-kinase regulatory subunit and the activity of the major PIPS phosphatase PTEN in cells. This has led us to new hypotheses about the important role of PI 3-kinase not only as a site of divergence of the insulin signaling pathways, but also a site of both positive and negative regulation in physiological and pathological states, and a site for cross-talk with other signaling systems, especially the stress kinases. In the next five years, we propose to 1) Dissect the multiple, differential roles of PI 3-kinase regulatory subunits (p85a/b, p50a and p55a/AS53) in insulin signaling, focusing on the potential actions and interactions emanating from the different N-terminal domains of the regulatory subunits that are independent of PI 3- kinase activity; 2) Determine the role of the PI 3-kinase catalytic subunits p110a and p11 Ob in divergent insulin signaling in vivo through tissue specific deletion ; and 3) Explore new chemical biology approaches to modifying PI 3-kinase mediated signaling in normal and insulin resistant states through the use of small molecule screening and new in vivo gene silencing techniques using nanoparticle delivery systems.