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，驱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的组合来协调维持胆固醇稳态。当细胞内胆固醇升高时，通过抑制转录因子固醇调节元件结合蛋白2 （SREBP2）的蛋白水解激活，编码胆固醇合成所需酶的基因表达受到抑制（负反馈）。相反，升高的胆固醇通过增加肝脏x受体（LXRs）的转录活性，诱导编码胆固醇排泄相关蛋白的基因表达（正前馈）。LXRs是配体激活转录因子核激素受体超家族的成员，在直接结合胆固醇衍生物时调节基因表达。我们认为，揭示胆固醇在慢性疾病中的作用的一种独特方法将是以细胞类型特定的方式可逆地破坏胆固醇稳态。为此，我们建议产生一种小鼠系，使色氨酸441突变为苯丙氨酸（W441F）的LXRα能够组织特异性表达。W441F破坏内源性胆固醇衍生的LXR配体的结合，同时仍然允许强效合成激动剂激活转录，提供了一种独特的工具，可以阻断LXRα感知胆固醇水平变化的能力，同时仍然允许药理控制。我们的初步数据表明，LXRα W441F作为显性阴性基因，关闭LXR转录活性，促进细胞内胆固醇积累。重要的是，用合成的LXR激动剂处理表达LXRα W441F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，在Cre重组酶控制下，通过表达LXRα W441F来调节胆固醇稳态的时间和空间。

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.