Cross-Species Genetic and Epigenetic Studies of Stress Response and Reward

应激反应和奖励的跨物种遗传学和表观遗传学研究

• 批准号: 8746468
• 负责人: Christina Barr
• 金额: $ 95.94万
• 依托单位: NATIONAL INSTITUTE ON ALCOHOL ABUSE AND ALCOHOLISM
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8746468
• 关键词: Affect; Affinity; Age; Aggressive behavior; Alcohol abuse; Alcohol consumption; Alcohols; Alleles; Amino Acid Substitution; Animals; Anti-Anxiety Agents; Anxiety; Archives; Area; Behavior; Behavior Disorders; Behavioral; Brain; Brain region; Canis familiaris; Catarrhine; Cebidae; Cercopithecidae; Cercopithecus tantalus; Characteristics; Code; DNA; DRD4 gene; Data; Detection; Disease; Dolphins; Dopamine; Elements; Endorphins; Epigenetic Process; Equilibrium; Equus caballus; Erythrocebus patas; Esthesia; Euphoria; European; Evolution; Exhibits; Exons; Family Felidae; Frameshift Mutation; Frequencies; Fright; Gene Expression; Genes; Genetic; Genetic Polymorphism; Genetic Variation; Genomics; Haplotypes; Human; Human Genetics; Impulsivity; Incidence; Individual Differences; Investigation; Length; Ligand Binding Domain; Ligands; Link; Location; Macaca; Macaca mulatta; Measures; Mental Depression; Mental disorders; Mission; Modeling; Names; National Institute on Alcohol Abuse and Alcoholism; Neuropeptides; Opioid; Oxytocin; Oxytocin Receptor; Pan Genus; Pattern; Physical aggression; Pongidae; Primates; Process; Psychopathology; RNA Splicing; Raccoons; Reagent; Receptor Gene; Regulation; Rewards; Risk; Risk Factors; Rodent; Role; Sampling; Site; Social Interaction; Stress; Structure; System; Tandem Repeat Sequences; Taxon; Temperament; Terminator Codon; Therapeutic Agents; Tissues; Translating; Tribes; Ursidae Family; Variant; Whole Blood; addiction; alcohol related problem; alcohol use disorder; base; behavioral genomics; biological adaptation to stress; comparative; deep sequencing; endophenotype; exome; exome sequencing; gain of function; gene environment interaction; genetic variant; genome sequencing; insertion/deletion mutation; interest; loss of function; neurogenetics; neurotransmission; nonhuman primate; receptor; response; sample fixation; screening; sex; social; social attachment; social organization; species difference; stress related disorder; trait; vervet

Identification of Genetic Variation using Massively Paralleled Sequencing This year, we sequenced the exomes of rhesus macaque subjects that were selected based on variation in temperament. One big advantage of using this approach is that DNA extracted whole blood can be used without the need for extracting DNA from the tissue of interest. Whole Exome Sequencing allows for the capture of large insertion/deletions and SNPs, and is best for Mendelian traits where variation is highly penetrant and either non-synonomous or frameshift. Drawbacks to WES include the inability to interrogate sequence sites that are not currently recognized as genes, control/regulatory sequences, structural variants such as translocations and inversions, splice-site variation, and exonic repeats. However, due to exome enrichment and the resulting deep sequencing coverage, the enhanced coding-region interrogation WES provides will allow for the sensitive detection of variants at those sites that are targeted. While there are no commercially available reagents for performing whole exome sequencing in nonhuman primates, a human whole exome platform is available. Given that there would likely be more purifying selection in coding regions and, therefore, less interspecific variation, we proposed using a human exome capture and sequencing kit for performing exome sequencing in rhesus macaques that differed in impulsivity and aggression. This approach was successful, and we identified many snps, including 391 missense variants, 23 stop codon snps, and 4 frameshift mutations. Of these, 58 variants were in genes in which variation is proposed to moderate addiction risk in humans. Whether these variants predict individual differences in alcohol drinking and related behaviors in rhesus macaques is currently under investigation. In certain instances, genetic variants that are functionally similar or orthologous to those that increase risk for human psychiatric disorders are maintained across primate species. We have identified several examples of this phenomenon in performing studies in rhesus macaques in order to model how genetic variation moderates risk for developing psychopathology. Some of these studies have suggested there to be convergent evolution or allelic variants being maintained by selection in both species. We may use naturally occurring genetic variation to look at not only within-species differences in vulnerability to behavioral disorders, but also to examine how genetic variation may alter traits in closely related species. Macaque species, all of which exhibit the same core elements of social organization and demographic structure, differ in characteristic patterns of stress reactivity, temperament and social interaction. We are currently performing whole exome sequencing a higher number of primate species, including the greater and lesser apes, and Old World monkeys (with an enriched sampling of Macaca) and the New World monkeys. One other area of interest for the study of genetic factors that contribute to addiction vulnerability is the genetics underlying domestication in animals.At its most basic, domestication is a suite of heritable traits affecting behavior. There are intriguing phenotypic commonalities among domesticates. Most important among these traits, and the only one common to all domesticates, is the ability to coexist with humans. The systems that likely permitted early domestication range from those involving fear and impulse control to those involving reward and sociality. Genetics contributing to such traits are likely to be relevant to human psychiatric illness and, in particular, the addictions. We have sequenced the genomes of domestic and European Wildcats, two species of felids that differ only in their behavior. Whether genetic variation that might underlie behavioral differences in these species is present is currently under investigation. Neurogenetics of Stress Response and Reward Sensitivity We have been studying how spontaneous genetic variation that influences reward sensitivity, and which could be adaptive in certain contexts, may also increase risk for alcohol related problems. In both humans and rhesus macaques, there are polymorphisms in the mu-opioid receptor gene (OPRM1) that influence affinity of the receptor for its endogenous ligand, -Endorphin. We and others have shown that these polymorphisms predict increased alcohol-induced euphoria and decreased stress response, suggesting them to have gain-of-function roles. We also have demonstrated it to relate to individual differences in response to natural rewards, as shown by measures of both reward bias in humans (and social attachment in both human and macaque. Recently, we have been screening this gene for variation across species in order to look for signs of selection and to determine whether functional variation that predicts species might be present. Of interest, we found a variant that produces a non-neutral amino acid substitution in the OPRM1 receptors ligand-binding domain (Asp12>Lys12). Among the Old World Monkey species that were screened (N = 17) this fixed difference was observed in all 5 species in the Cercopthecini tribe (which includes Patas monkeys and vervets, to name a few), but this allele was not observed outside of this tribe. Field observations of Vervet and Patas suggest they are reward-sensitive and more susceptible to stress compared to other OWMs. Our data suggest that an OPRM1 variant arose and went to fixation after the divergence of Cercopthecini from Papionini. They further support data showing that polymorphism at OPRM1 contributes to behavioral variation in primates. Dopamine neurotransmission underlies many reward-dependent and reinforcing processes. Tandem repeats in the third exon of the dopamine receptor D4 gene (DRD4) exist across a variety of animal species (humans, vervets, dogs, dolphins, bears, raccoons, horses, and chimpanzees). Some species, including macaque species, exhibit variation in the number of repeats present, with various alleles differing in frequencies across species. In humans, the DRD4 7-repeat allele (7R) reduces efficacy of the receptor and is linked with traits such as sensation seeking. This has been replicated in vervet monkeys, in which a DRD4 length variant was also observed. Of interest, in the dog, which has been subject to intense artificial selection, a repeat polymorphism predicts social impulsivity and activity-impulsivity endophenotypes. We recently identified 4R, 5R, 6R, and 7R alleles in a colony of rhesus macaques. Compared to the ancestral allele (4R), the loss-of-function 7R allele predicted increased incidence of physical aggression toward an unknown age- and sex-matched conspecific. One system of potential relevance to the addictions via its effects on anxiety and reward is the oxytocin system. We screened the rhesus OXTR gene and identified 17 SNPs, including 4 non-synonymous SNPs in the first exon for the rhesus Oxytocin receptor gene (OXYR). This is a location at which there is high interspecific conservation yet high frequencies of non-synonymous SNPs in humans as well. Among these, some of them are conserved across primate taxa, suggesting that they have been maintained by selection. We also found there to be ancient polymorphisms maintained in humans and the existence of alternative haplotypes within this region indicating that human variation is being maintained by balancing selection. Oxytocin is a neuropeptide that produces affiliative, amnesic and anxiolytic affects. Given its roles in some of these processes, it has been proposed as a potential therapeutic agent for the treatment of anxiety and stress-related disorders, including the addictions.Functional variation that influences oxytocin system function may also be a good candidate for performing GxE studies.

使用大规模并行测序鉴定遗传变异 今年，我们对根据气质变异选择的恒河猴受试者进行了外显子组测序。使用这种方法的一大优点是可以使用提取 DNA 的全血，而不需要从感兴趣的组织中提取 DNA。全外显子组测序可以捕获大量插入/缺失和 SNP，最适合变异高度渗透且非同义或移码的孟德尔性状。 WES 的缺点包括无法询问目前未被识别为基因的序列位点、控制/调节序列、结构变异（例如易位和倒位）、剪接位点变异和外显子重复。然而，由于外显子组富集和由此产生的深度测序覆盖，WES 提供的增强型编码区询问将允许灵敏地检测目标位点的变异。 虽然没有市售试剂可用于在非人类灵长类动物中进行全外显子组测序，但人类全外显子组平台是可用的。鉴于编码区可能有更多的纯化选择，因此种间变异更少，我们建议使用人类外显子组捕获和测序试剂盒对冲动性和攻击性不同的恒河猴进行外显子组测序。这种方法很成功，我们鉴定了许多 snps，包括 391 个错义变异、23 个终止密码子 snps 和 4 个移码突变。其中，58 个基因变异被认为可以减轻人类成瘾风险。目前正在研究这些变异是否可以预测恒河猴饮酒和相关行为的个体差异。 在某些情况下，灵长类动物中存在与增加人类精神疾病风险的基因变异功能相似或同源的基因变异。我们在对恒河猴进行研究时发现了这种现象的几个例子，以便模拟遗传变异如何调节发生精神病理学的风险。其中一些研究表明，两个物种都存在趋同进化或通过选择维持的等位基因变异。我们不仅可以利用自然发生的遗传变异来观察物种内易受行为障碍影响的差异，还可以研究遗传变异如何改变密切相关的物种的性状。所有猕猴物种都表现出相同的社会组织和人口结构核心要素，但在压力反应、气质和社会互动的特征模式上有所不同。我们目前正在对更多灵长类物种进行全外显子组测序，包括大型和小型猿类、旧世界猴（包括丰富的猕猴样本）和新世界猴。 研究导致成瘾脆弱性的遗传因素的另一个感兴趣的领域是动物驯化背后的遗传学。从最基本的角度来说，驯化是一套影响行为的遗传特征。驯化动物之间存在着有趣的表型共性。这些特征中最重要的、也是所有驯化动物唯一共有的特征是与人类共存的能力。可能允许早期驯化的系统范围从涉及恐惧和冲动控制的系统到涉及奖励和社交的系统。造成这些特征的遗传学可能与人类精神疾病，特别是成瘾有关。我们对家猫和欧洲野猫的基因组进行了测序，这两种猫科动物仅在行为上有所不同。目前正在调查是否存在可能导致这些物种行为差异的遗传变异。 压力反应和奖励敏感性的神经遗传学 我们一直在研究影响奖励敏感性的自发遗传变异，以及在某些情况下可能具有适应性的自发遗传变异，也可能会增加酒精相关问题的风险。在人类和恒河猴中，mu-阿片受体基因 (OPRM1) 存在多态性，影响受体与其内源性配体 - 内啡肽的亲和力。我们和其他人已经证明，这些多态性预测酒精引起的欣快感增加和应激反应减少，表明它们具有功能获得作用。我们还证明了它与对自然奖励反应的个体差异有关，正如人类奖励偏差（以及人类和猕猴的社会依恋）的测量所表明的那样。最近，我们一直在筛选该基因的跨物种变异，以便为了寻找选择的迹象并确定是否可能存在预测物种的功能变异，我们发现了一种在 OPRM1 受体配体结合域（Asp12>Lys12）中产生非中性氨基酸取代的变体。这筛选的旧世界猴种（N = 17）在 Cercopthecini 部落（包括帕塔斯猴和黑长尾猴等）的所有 5 个物种中观察到这种固定差异，但在该部落之外没有观察到这种等位基因。对 Vervet 和 Patas 的现场观察表明，与其他 OWM 相比，它们对奖励敏感且更容易受到压力影响。我们的数据表明，OPRM1 变体在 OWM 分化后出现并进入固定状态。来自 Papionini 的 Cercopthecini。他们进一步支持了表明 OPRM1 多态性导致灵长类动物行为变异的数据。 多巴胺神经传递是许多奖赏依赖性和强化过程的基础。多巴胺受体 D4 基因 (DRD4) 第三外显子中的串联重复存在于多种动物物种（人类、黑长尾猴、狗、海豚、熊、浣熊、马和黑猩猩）中。一些物种，包括猕猴物种，表现出重复数量的变化，各种等位基因在物种之间的频率不同。 在人类中，DRD4 7 重复等位基因 (7R) 会降低受体的功效，并与感觉寻求等特征相关。这已在长尾黑颚猴中得到复制，其中还观察到了 DRD4 长度变异。有趣的是，在经过强烈人工选择的狗中，重复多态性预测了社交冲动和活动冲动内表型。我们最近在一群恒河猴中鉴定出了 4R、5R、6R 和 7R 等位基因。与祖先等位基因 (4R) 相比，功能丧失的 7R 等位基因预测对未知年龄和性别匹配的同种人的身体攻击发生率会增加。 催产素系统是通过其对焦虑和奖励的影响而与成瘾潜在相关的一种系统。我们筛选了恒河猴 OXTR 基因并鉴定了 17 个 SNP，其中包括恒河猴催产素受体基因 (OXYR) 第一个外显子中的 4 个非同义 SNP。这个位置在人类中存在高度种间保守性，但非同义 SNP 的频率也很高。其中，其中一些在灵长类动物类群中是保守的，这表明它们是通过选择而得以维持的。我们还发现人类中保留着古老的多态性，并且该区域内存在替代单倍型，表明人类变异是通过平衡选择来维持的。催产素是一种神经肽，能产生亲和力、遗忘力和抗焦虑作用。鉴于其在其中一些过程中的作用，它已被提议作为治疗焦虑和压力相关疾病（包括成瘾）的潜在治疗剂。影响催产素系统功能的功能变异也可能是进行 GxE 研究的良好候选者。