Tissue Specific Control of Cholesterol Metabolism

胆固醇代谢的改变通常与慢性衰老疾病有关，包括心血管疾病、II型糖尿病、非酒精性脂肪性肝炎、多发性硬化症和阿尔茨海默病。虽然胆固醇升高与许多慢性疾病有关，但在大多数情况下，推动功能病理变化的胆固醇依赖途径尚未得到很好的描述。细胞使用负反馈和正前馈控制相结合的方式来协调维持胆固醇的稳态。当细胞内胆固醇增加时，通过抑制转录因子类固醇调节元件结合蛋白2(SREBP2)的蛋白水解性激活，胆固醇合成所需的编码酶的基因表达受到抑制(负反馈)。相反，高胆固醇通过增加肝脏x受体(LXRs)的转录活性来诱导与胆固醇排泄相关的编码蛋白的表达(正前馈)。LXRs是核激素受体超家族的成员，配体激活的转录因子调节基因表达，以响应胆固醇衍生物的直接结合。我们建议，解开胆固醇在慢性病中的作用的独特方法将是以一种特定细胞类型的方式可逆地破坏胆固醇稳态。为此，我们建议建立一种小鼠系，允许组织特异性表达LXRα，并将色氨酸441突变为苯丙氨酸(W441F)。W441F破坏内源性胆固醇衍生的LXR配体的结合，同时仍允许强大的合成激动剂激活转录，提供了一种独特的工具，阻止LXRα感知胆固醇水平变化的能力，同时仍允许药物控制。我们的初步数据表明，LXRαW441F作为一个显性负性基因，关闭了LXR的转录活性，促进了细胞内胆固醇的积累。重要的是，通过用合成的LXR激动剂处理表达LXRαW441F的细胞可以恢复LXR的活性。因此，我们建议建立一种新的小鼠模型，通过在Cre重组酶控制下表达LxRαW441F来调节胆固醇的时空平衡。

Alterations in cholesterol metabolism are often associated with chronic diseases of aging including cardiovascular disease, type II diabetes, non-alcoholic steatohepatitis, multiple sclerosis, and Alzheimer’s disease. While elevated cholesterol is linked to many chronic diseases, in most cases the cholesterol-dependent pathways that drive pathological changes in function have not been well described. Cells use a combination of negative feedback and positive feed forward control to coordinately maintain cholesterol homeostasis. When intracellular cholesterol increases, expression of genes encoding enzymes required for cholesterol synthesis is repressed (negative feedback) by inhibiting the proteolytic activation of the transcription factor sterol regulatory element binding protein 2 (SREBP2). In contrast, elevated cholesterol induces expression of genes encoding proteins involved in cholesterol excretion (positive feed forward) by increasing the transcriptional activity of the liver x receptors (LXRs). The LXRs are members of the nuclear hormone receptor superfamily of ligand activated transcription factors that regulate gene expression in response to the direct binding of cholesterol derivatives. We suggest that a unique approach to unraveling the roles of cholesterol in chronic diseases will be to reversibly disrupt cholesterol homeostasis in a cell type specific manner. To this end we propose to generate a mouse line that allows tissue specific expression of LXRα with a mutation of tryptophan 441 to phenylalanine (W441F). W441F disrupts binding of endogenous cholesterol-derived LXR ligands while still allowing transcription activation by potent synthetic agonists providing a unique tool that blocks the ability of LXRα to sense changes in cholesterol levels while still allowing pharmacological control. Our preliminary data indicates that LXRα W441F functions as a dominant negative that shuts down LXR transcriptional activity and promotes intracellular cholesterol accumulation. Importantly, LXR activity can be restored by treating cells expressing LXRα W441F with synthetic LXR agonists. Therefore, we propose to develop a novel mouse model that allows temporal and spatial regulation of cholesterol homeostasis by expressing LXRα W441F under Cre recombinase control.