Nuclear Lipids in Steroidogenesis

描述（由申请人提供）：类固醇激素是多种生理过程的关键调节剂，包括钠稳态、生殖和第二性征的发育。这些分子允许组织以协调的方式对内部和外部环境的变化作出反应，作为核和质膜受体的配体。由于类固醇激素控制几乎所有细胞类型中许多基因的表达，类固醇生成细胞利用多种机制确保严格控制这些分子的合成。我们研究的一个主要目标是了解垂体衍生激素促肾上腺皮质激素（ACTH）调节肾上腺皮质皮质产生皮质醇的机制。我们的研究已经确定了生物活性鞘脂和磷脂的关键作用。这些分子通过控制类固醇基因的转录来控制激素的产生。我们推测ACTH通过调节肾上腺皮质细胞核中生物活性鞘脂和磷脂的可用性来控制类固醇激素的生物合成。具体目标1将定义核鞘脂和磷脂代谢的能力。初步数据已经确定，生物活性磷脂和鞘脂通过作为核受体类固醇生成因子-1（SF-1）的配体来控制类固醇生成基因的转录。由于主要位于细胞核中，我们假设局部合成的生物活性脂质是控制SF-1功能的关键。脂质分析将用于全面和定量评估细胞核中鞘脂和磷脂物质的浓度以及ACTH对这些脂质量的影响。特异性目的2确定酸性神经酰胺酶（由ASAH 1b编码）调节核SPH浓度和SF-1功能的机制。ASAH 1b是产生鞘氨醇（SPH）的三种神经酰胺酶之一。我们有证据支持ASAH 1b和SF-1之间的直接相互作用以及通过磷酸化调节ASAH 1功能的作用。我们将研究这种酶作为SF-1共调节蛋白的作用。具体目标3将定义cAMP信号传导的激活促进类固醇基因表达的配体（PA）依赖性激活的机制。类固醇生成基因表达的增加与SF-1的结合激动剂配体（磷脂酸）以及随后的转录激活有关。串联质谱和酶测定将被用来确定翻译后修饰在控制合成磷脂酸的酶的活性中的作用。总之，这一建议解决了主要问题的机制，基因调控本地产生的生物活性脂质。从这些研究中获得的知识将提供有关细胞核代谢能力的有价值的信息，以及生物活性脂质的动态通量在基因表达控制中发挥的关键作用。 公共卫生相关性：了解基因是如何在细胞中调节，使类固醇激素将提供深入了解病理生理浓度的皮质醇和肾上腺雄激素的产生机制。这项工作将提供深入了解多种内分泌疾病，包括肾上腺皮质增生，多囊卵巢综合征，和库欣病。

DESCRIPTION (provided by applicant): Steroid hormones are key regulators of a diverse array of physiological processes, including sodium homeostasis, reproduction, and the development of secondary sex characteristics. These molecules allow tissues to respond in a coordinated manner to changes in the internal and external environments by functioning as ligands for both nuclear and plasma membrane receptors. Because steroid hormones control the expression of numerous genes in virtually all cell types, steroidogenic cells utilize multiple mechanisms that ensure tight control of the synthesis of these molecules. A major goal of our research is to understand the mechanisms by which the pituitary-derived hormone adrenocorticotropin (ACTH) regulates cortisol production by the adrenal cortex. Our research has identified a key role for bioactive sphingolipids and phospholipids. These molecules control hormone production by controlling the transcription of steroidogenic genes. We hypothesize that ACTH controls steroid hormone biosynthesis by modulating the availability of bioactive sphingolipids and phospholipids in the nuclei of adrenocortical cells. Specific Aim 1 will define the capacity for nuclear sphingolipid and phospholipid metabolism. Preliminary data has established that bioactive phospholipids and sphingolipids control steroidogenic gene transcription by serving as ligands for the nuclear receptor steroidogenic factor-1 (SF-1). Since is primarily localized in the nucleus, we hypothesize that locally synthesized bioactive lipids are key for controlling SF-1 function. Lipid profiling will be used to gain a comprehensive and quantitative assessment of the concentrations of sphingolipid and phospholipid species in the nucleus and the effect of ACTH on the amounts of these lipids. Specific Aim 2 determine the mechanism by which acid ceramidase (encoded by ASAH1b) regulates nuclear SPH concentrations and SF-1 function. ASAH1b is one of three ceramidases that produces sphingosine (SPH). We have evidence to support a role for direct interaction between ASAH1b and SF-1 and for the regulation of ASAH1 function by phosphorylation. We will investigate the role of this enzyme as a SF-1 coregulatory protein. Specific Aim 3 will define the mechanism by which activation of cAMP signaling promotes ligand (PA)-dependent activation of steroidogenic gene expression. Increased steroidogenic gene expression occurs with the binding agonist ligand (phosphatidic acid) to SF-1 and the subsequent activation of transcription. Tandem mass spectrometry and enzymatic assays will be used to define the role of posttranslational modification in controlling the activity of enzymes that synthesize phosphatidic acid. In sum this proposal addresses major questions about the mechanisms underlying gene regulation by locally produced bioactive lipids. The knowledge gained from these studies will provide valuable information about the metabolic capacity of the nucleus and the key role that dynamic flux of bioactive lipids plays in the control of gene expression. PUBLIC HEALTH RELEVANCE: Relevance Understanding how genes are regulated in cells that make steroid hormones will provide insight into the mechanisms by which pathophysiological concentrations of cortisol and adrenal androgens are produced. This work will provide insight into multiple endocrine disorders, including adrenal hyperplasia, polycystic ovary syndrome, and Cushing's disease.