Cocaine Addiction and Retrotransposons

描述（由申请人提供）：可卡因成瘾（CA）是一种常见的脑部疾病，影响到美国人口中约100万成年人，由于其慢性性质和相关的发病率和死亡率，造成了重大的公共卫生问题。尽管双胞胎研究与遗传成分一致，但通过研究血液或唾液中的DNA来识别增加风险的等位基因一直很困难。逆转录转座子（RTPs）是人类基因组中常见的可移动DNA元件。在过去的5年中，积累的数据证明，神经元胚胎发生伴随着LINE1 (L1) RTP的激活，其程度出乎意料，以至于每个发育中的神经元可能积累多个从头开始的L1 RTP基因组插入，可能多达80个。这导致了中枢神经系统神经元群体中大量的镶嵌现象。虽然大多数体细胞新生L1 RTP插入对神经元功能几乎没有影响（可能是因为它们发生在基因沙漠或大内含子或细胞功能不需要的基因中），但有些可能会干扰正常的神经元活动，因为它们插入了特定神经元正常功能所需的基因。如果一个或多个功能性L1 RTP插入事件发生在中枢神经系统发育早期，那么源自该神经元前体的所有子神经元也将携带L1插入，可能导致功能失调的神经元群体，从而传递CA的风险。该应用将利用死于可卡因过量的CA患者的死后脑组织，以及与对照组年龄/性别/种族匹配的死后脑组织。激光捕获显微解剖将用于获取内侧额叶皮层、杏仁核和纹状体（CA动物模型中涉及神经适应的大脑区域）的不同种群。来自这些神经元的DNA将通过高通量测序进行分析，以检测新生L1 RTP插入。在脑表达基因中检测到的任何从头（不在参考基因组中）L1 RTP插入将使用标准转录分析来研究其功能意义。通过这种方式，预计将发现增加CA风险的新的新生体细胞L1 RTP插入。

DESCRIPTION (provided by applicant): Cocaine addiction (CA) is a common brain disorder, affecting ~ 1,000,000 adults in the US population, creating a significant public health problem because of its chronic nature and the associated morbidity and mortality. Although twin studies are consistent with an inherited component, it has been difficult to identify risk-increasing allels by studying DNA from blood or saliva. Retrotransposons (RTPs) are mobile DNA elements that are common in the human genome. In the past 5 years, data have accumulated to prove that neuronal embryogenesis is accompanied by activation of LINE1 (L1) RTPs to an unexpected degree, such that each developing neuron may accumulate multiple de novo L1 RTP genomic insertions, perhaps as many as ~80. This results in a substantial mosaicism within populations of CNS neurons. While most of these somatic de novo L1 RTP insertions will have little effect on neuronal function (perhaps because they occur in gene deserts or in large introns or in genes not required for that cell's function), some may interfere with normal neuronal activity because they have inserted into a gene needed by that particular neuron for normal function. If one or more functional L1 RTP insertion events occur early in CNS development, all the daughter neurons that derive from that neuronal precursor will also carry the L1 insertion, perhaps leading to a dysfunctional population of neurons destined to convey risk for CA. This application will leverage post-mortem brain tissue from CA patients, who have died from a cocaine overdose, and age/gender/ethnicity matched post-mortem brain tissue from controls. Laser capture microdissection will be used to harvest distinct populations of medial frontal cortex, amygdala and striatum (brain regions implicated in neuroadaptation in CA animal models). DNA from these neurons will be analyzed by high-throughput sequencing, to detect de novo L1 RTP insertions. Any de novo (not in the reference genome) L1 RTP insertion detected in a brain-expressed gene will be investigated for functional significance using standard assays of transcription. In this manner, it is expected that novel de novo somatic L1 RTP insertions, that increase risk for CA, will be discovered.