Beta-adrenergic regulation of adipose tissue function by PKA and MAP kinases

摘要 肥胖现在处于流行状态，是导致II型糖尿病等几种衰弱疾病的风险因素， 高血压和心血管疾病。 肾上腺素能受体(ARs)是脂肪组织代谢的关键调节因子，可控制(1)脂解作用 棕色脂肪中储存能量的动员和(Ii)适应性产热作用。所有3个已知的AR子类型都用 脂肪细胞和ARs与细胞内cAMP的增加和cAMP依赖的蛋白激酶的激活有关 (PKA)然而，除了这个经典的途径，我们已经证明了额外的信号级联来自ARs 在脂肪细胞中，其中包括ERK和p38 MAP激酶通路。 这项建议的目标是确定PKA和/p38 MAP激酶之间的信号成分，这些信号成分传递 白色和棕色脂肪细胞中AR介导的信号转导，并确定白色脂肪细胞中p38MAPK的功能 脂肪细胞。为了实现这些目标，我们(I)将MKK3和p38MAPK确定为这一目标的必要组成部分 Casade；(Ii)建立了p38的转录靶点；(Iii)产生了p38？MAPK的脂肪特异性缺失 (p38？/)。尽管缺乏p38？，但在p38？/只小鼠中，棕色脂肪中的产热作用被保留或略有升高。 在缺乏p38的情况下，JNK被选择性地激活，而p38？/动物即使在高脂肪的饮食中也是瘦的。 为这个新的项目期提出了三个目标： 目的1：建立从PKA到MKK3/p38pha的信号转导机制的其余组成部分 MAPK。实验通过候选MKKK的层次结构来识别负责激活的MKKK MKK3和p38MAPK在棕色和白色脂肪细胞中的表达。这些实验还检验了假设一个小G 像rac1这样的蛋白质是这条途径的一个组成部分，我们将确定协调 MAPK、MKK和MKKK组成一个运作单元。 目的2：确定缺乏p38(p38/)的脂肪细胞中的AR信号通路如何重新连接到 利用JNK作为“代偿”的激酶。实验将确定p38？/中的信号元件 现在可以激活JNK的脂肪细胞。它评估了MKKK和 目的1，以及它们的参与是否在p38/小鼠中保守。这些实验还确定了 脂肪细胞中已知的p38转录因子靶点(ATF-2和PGC-1)，以及它们现在是否受到 假设脂肪细胞中p38的一个关键靶点是维持JNK的双特异性MAPK磷酸酶 在非活性、去磷酸化状态下，也进行了探索。 目的3：验证p38？/小鼠瘦肉型是能量消耗增加所致的假说。这个 这一目的的实验测试了在缺乏p38的情况下，JNK现在是否是一种更强大的棕色脂肪适应性激活剂 产热和能量消耗，导致较小的白色脂肪细胞和瘦肉型，以及几个特定的 并提出了相应的解决机制。我们还研究了瘦肉型和较小的脂肪库存在于 代价是脂肪酸存储容量不足，矛盾的是，这会导致胰岛素抵抗和脂肪变性。 PHS 398/2590(09/04版，2006年4月4日重新发布)页面续格式页面

ABSTRACT Obesity is now at epidemic proportions and is a risk factor for several debilitating illnesses such as Type II diabetes, hypertension and cardiovascular disease. The ¿-adrenergic receptors (¿ARs) are key regulators of adipose tissue metabolism to control (i) lipolysis for mobilization of stored energy and (ii) adaptive thermogenesis in brown fat. All 3 known ¿AR subtypes are expressed in adipocytes, and ¿ARs are coupled to increases in intracellular cAMP and activation of cAMP-dependent protein kinase (PKA) However, besides this classic pathway, we have shown that additional signaling cascades emanate from ¿ARs in adipocytes, and these include the ERK and p38 MAP kinase pathways. The goals of this proposal are to identity the signaling components between PKA and /p38 MAP kinase that convey the ¿AR-mediated signaling in white and brown adipocytes, and determine the function of p38 MAPK in white adipocytes. Toward achieving these goals we have (i) identified MKK3 and p38¿ MAPK as necessary components of this cascade; (ii) established the transcription targets of p38; (iii) generated an adipose-specific deletion of p38¿ MAPK (p38¿/). Despite the absence of p38¿, thermogenesis in brown fat is preserved or slightly elevated in p38¿/ mice, there is selective activation of JNK in the absence of p38¿, and p38¿/ animals are lean even on a high-fat diet. Three Aims are proposed for this new project period: Aim 1: Establish the remaining components of the signal transduction mechanism from PKA to MKK3/p38alpha MAPK. The experiments proceed through a hierarchy of candidate MKKKs to identify the one responsible for activating MKK3 and p38¿ MAPK in brown and white adipocytes. These experiments also test the hypotheses that a small G protein such as Rac1 is a component of this pathway, and we will identify the scaffolding protein that coordinates the MAPK, the MKK and the MKKK into a functioning unit. Aim 2: Determine how the ¿AR signaling pathway in adipocytes lacking p38¿ (p38¿/) has become "re-wired" to utilize JNK as the "compensatory" kinase. The experiments will determine the signaling elements in p38¿/ adipocytes that now allow JNK to be activated. It assesses the flexibility of the MKKK and other elements identified in Aim 1, and whether their participation is conserved in the p38¿/ mouse. These experiments also determine the status of the known transcription factor targets of p38¿ in adipocytes (ATF-2 and PGC-1¿), and whether they are now regulated by JNK The hypothesis that a key target of p38¿ in adipocytes is a dual-specificity MAPK phosphatases, that maintains JNK in an inactive, dephosphorylated state, is also explored. Aim 3: Test the hypothesis that the lean phenotype in p38¿/ mice is due to increased energy expenditure. The experiments in this Aim test whether in the absence of p38¿, JNK is now a more powerful activator of brown fat adaptive thermogenesis and energy expenditure, resulting in smaller white adipocytes and a lean phenotype, and several specific mechanisms are proposed. We also examine the possibility that the lean phenotype and smaller adipose depots exists at the expense of an insufficient storage capacity for fatty acids, paradoxically resulting in insulin resistance and steatosis. PHS 398/2590 (Rev. 09/04, Reissued 4/2006) Page Continuation Format Page