Systems genetics of premorbid and cocaine use traits in a rat reduced complexity cross

项目总结 兴奋剂使用障碍(如可卡因、甲基苯丙胺)是一个主要的公共卫生问题。尽管有一个 遗传率约为40%-50%，全基因组关联研究(GWAS)已确定极少数基因座，包括一个命中 对于可卡因依赖，映射到FAM53B的基因也是通过表达数量性状鉴定的 基因座(QTL)分析与小鼠COC自我给药相关。主要目标是迅速 在大鼠中识别导致发病前风险(强迫症、冲动性)和 自发性高血压大鼠(SHR)降低复杂性交叉(RCC)的可卡因使用特征。一只啮齿动物 基于发现的遗传和多水平功能基因组分析的系统遗传学方法三角化 并且可以提供对药物作用和潜在的神经可塑性的更快速的遗传和神经生物学洞察 上瘾。几年来，接触式PI一直使用鼠标降低复杂性交叉(RCC) 近等基因近交系亚系之间的差异，以促进基因定位、验证和机制。因为 啮齿动物亚株在基因上99%相同，并且包含的变异少了几个数量级 与经典近交系相比，定位RCCS中的数量性状基因座(QTL)产生了数量级的结果 需要考虑的候选因果基因更少。当与功能基因组学相结合时，RCCS可以迅速导致 致病基因和变异体鉴定。我们的初步研究在发病前建立了强大的、可遗传的差异 SHR/NCr1和SHR/NCr1之间的冲动性和强迫性、蔗糖奖赏敏感性和多COC使用特性 SHR/nhsd亚株，包括COC诱导的运动活动，COC IVSA摄取，寻找和摄取周期， 论证了在RCC中进行基因定位的可行性。在目标1和目标2中，我们将率先使用大鼠RCC 在那里我们将进行全基因组测序(WGS)并定位行为QTL和表达QTL 伏隔核(NAC)和前额叶皮质(PFC)在整个转录和外显子水平的eQTL F2杂交包括COC训练的大鼠和带轭生理盐水(SAL)训练的大鼠。在目标3中，我们将进行蛋白质组学 高置信度条件下COC训练大鼠的PFC和NAC分析 数量性状基因(QTGs)和变体(QTV)当我们在DNA变体之间建立功能联系时， 病前和可卡因潜在的转录调控、蛋白质翻译和细胞信号适应 利用他们的特点。这些研究开创了利用大鼠肾细胞癌结合深度行为表型来快速 确定高置信度候选新的遗传因素和影响病前风险的分子机制 因素和可卡因使用特征。未来候选因果基因变体的基因编辑将以这两个为基础 近等基因SHR背景，以证明必要性(基因突变纠正；“拯救”)和充分性 (诱变)。交付内容包括用于复杂性状分析的未来RCC的SHR亚株的WGS 以及大鼠中脑皮质边缘关键区域的适应性转录和蛋白质组学数据集 研究人员可以进一步挖掘的电路，希望能为治疗学提供信息。

PROJECT SUMMARY Stimulant use disorders (e.g., cocaine, methamphetamine) are a major public health concern. Despite a heritability of ~40-50%, genome-wide association studies (GWAS) have identified very few loci, including one hit for cocaine (COC) dependence that maps to FAM53B, a gene also identified via expression quantitative trait locus (QTL) analysis to be associated with COC self-administration in mice. The primary objective is to rapidly identify novel genetic factors in rats that contribute to premorbid risk (compulsivity, impulsivity) and cocaine use traits in a spontaneously hypertensive rat (SHR) reduced complexity cross (RCC). A rodent systems genetics approach triangulates on discovery-based genetic and multi-level functional genomic analysis and can provide a more rapid genetic and neurobiological insight into drug action and neuroplasticity underlying addiction. For several years, the contact PI has been employing mouse reduced complexity crosses (RCCs) between near-isogenic inbred substrains to facilitate gene mapping, validation, and mechanisms. Because rodent substrains are > 99% genetically identical and contain several orders of magnitude fewer variants compared to classical inbred strains, mapping quantitative trait loci (QTLs) in RCCs yields orders of magnitude fewer causal candidate genes to consider. When combined with functional genomics, RCCs can rapidly lead to causal gene and variant identification. Our preliminary studies establish robust, heritable differences in premorbid impulsivity and compulsivity, sucrose reward sensitivity, and multiple COC use traits between SHR/NCrl and SHR/NHsd substrains, including COC-induced locomotor activity, COC IVSA taking, seeking, and intake cycles, demonstrating feasibility for gene mapping in an RCC. In Aims 1 and 2, we will pioneer the use of a rat RCC where we will conduct whole genome sequencing (WGS) and map behavioral QTLs and expression QTLs (eQTLs) from nucleus accumbens (NAC) and prefrontal cortex (PFC) at the whole transcript and exon levels in an F2 cross comprising COC-trained versus yoked saline (SAL)-trained rats. In Aim 3, we will conduct proteomic analysis of PFC and NAC from COC vs. yoked SAL-trained rats to triangulate on high confidence candidate quantitative trait genes (QTGs) and variants (QTVs) as we build functional connections between DNA variants, transcriptional regulation, protein translation, and cell signaling adaptations underlying premorbid and cocaine use traits. These studies pioneer the use of a rat RCC combined with deep behavioral phenotyping to rapidly identify high-confidence candidate novel genetic factors and molecular mechanisms influencing premorbid risk factors and cocaine use traits. Future gene editing of candidate causal gene variants will be modeled on the two near-isogenic SHR backgrounds to demonstrate necessity (mutation correction; “rescue”) and sufficiency (mutation induction). Deliverables include WGS’s of SHR substrains for future RCCs for complex trait analysis as well as adaptive rat transcriptomic and proteomic datasets in key brain regions of the mesocorticolimbic circuitry that can be further mined by investigators and hopefully inform therapeutics.