Regulation of alcohol consumption by chromatin modification

描述（由申请人提供）：在之前的INIA（酒精中毒综合神经科学倡议）-West项目期间，应用了遗传基因组/表型方法来确定啮齿动物中的候选基因，这些基因通过表达水平的变化导致了摄入不同数量酒精的倾向。其中一个候选基因是Gnb1 （G蛋白β 1亚基，Gpi）。在自交系和重组自交系小鼠以及因酒精消耗差异而选择的小鼠品系中，大脑中Gpi蛋白的水平与酒精消耗水平呈负相关。用表达靶向Gnb1 shRNA的慢病毒载体治疗小鼠，降低了DBA/2小鼠伏隔核中Gpi的水平，增加了酒精消耗。Gpi作为与Gy蛋白的二聚体，影响许多细胞内信号通路，并且也被报道与la类组蛋白去乙酰化酶（HDACs） 4和5相互作用，并改变其转录活性。hdac通常通过修饰组蛋白来抑制转录，hdac可以在细胞核和细胞质之间穿梭，从而调节其转录活性。特别是，Gpy-HDAC的直接相互作用降低了HDAC的活性，因此有可能调节其他基因的转录。考虑到HDAC活性先前被发现对可卡因的奖励作用至关重要，并且HDAC抑制减少酒精消耗，假设Gpy-HDAC相互作用在酒精消耗中起作用。在低酒精摄入量小鼠大脑中Gnb1表达降低后，以及在高酒精摄入量小鼠大脑中抑制或降低hdac表达后，测量酒精摄入量。为了评估Gpi和HDAC之间的联系，将在RNAi降低Gpi后确定HDAC的转录活性（转录物水平和启动子乙酰化）以及HDAC的亚细胞定位。总体而言，该项目将研究分子机制，重点关注染色质修饰，通过该修饰，先前确定的酒精消费/偏好的候选基因影响可能构成该表型的转录网络。

DESCRIPTION (provided by applicant): During the previous INIA (Integrated Neurosciences Initiative on Alcoholism)-West project period a genetical genomic/phenomic approach was applied to identify candidate genes in rodents that, through variation in expression levels, contribute to the predisposition to consume varying amounts of alcohol. One such candidate gene is Gnb1 (the G protein beta 1 subunit, Gpi). Brain levels of the Gpi protein are inversely correlated with levels of alcohol consumption in inbred and recombinant inbred strains of mice and in lines of mice selected for differences in alcohol consumption. Treatment of mice with a lentiviral vector expressing a shRNA targeting Gnb1 lowered Gpi levels in nucleus accumbens and increased alcohol consumption by DBA/2 mice. Gpi, as a dimer with Gy proteins, affects numerous intracellular signaling pathways, and has also been reported to interact with, and modify the transcriptional activity of, the Class lla histone deacetylases (HDACs) 4 and 5. HDACs generally repress transcription via modification of histone proteins and class lla HDACs can shuttle between nucleus and cytoplasm, which can regulate their transcriptional activity. In particular, the direct Gpy-HDAC interaction reduces HDAC activity, and therefore has the potential to regulate the transcription of other genes. Given that HDACS activity has previously been found to be crucial for the rewarding effect of cocaine, and that HDAC inhibition reduces alcohol consumption, a role for the Gpy-HDAC interaction in alcohol consumption is postulated. Alcohol consumption will be measured after lowering Gnb1 expression in brains of low alcohol-consuming mice, and after inhibiting or lowering expression of HDACs in brain of high alcohol-consuming mice. To assess the link between Gpi and HDACs, HDAC transcriptional activity (transcript levels and promoter acetylation), as well as HDAC subcellular localization, will be determined after lowering Gpi with RNAi. Overall, this project will investigate molecular mechanisms, focusing on chromatin modification, by which a previously identified candidate gene for alcohol consumption/preference influences transcriptional networks that may underlie this phenotype.