Retrotransposons in Schizophrenia

精神分裂症中的反转录转座子

• 批准号: 9127614
• 负责人: Wade H Berrettini
• 金额: $ 67.73万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-21 至 2021-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9127614
• 关键词: Adolescence; Affect; Alleles; Anterior; Antibodies; Autopsy; Biological Assay; Brain; Brain Diseases; Brain region; CRISPR/Cas technology; Cardiovascular Diseases; Cell Line; Cell Nucleus; Cell Separation; Cell membrane; Centrifugation; Chromatin; Chronic; Collaborations; Copy Number Polymorphism; DNA; DNA Sequence; Data; Daughter; Development; Disease; Doctor of Philosophy; Electroencephalography; Electrophysiology (science); Elements; Fluorescence; Gene Expression; Gene Frequency; Genes; Genetic; Genetic Transcription; Genome; Genomic DNA; Genomic Segment; Grant; Heritability; High-Throughput Nucleotide Sequencing; Human Genome; Inherited; Interneurons; Jumping Genes; L1 Elements; Label; Letters; Life; Life Expectancy; Membrane Proteins; Morbidity - disease rate; Mosaicism; Neurodevelopmental Disorder; Neurons; Nuclear; Odds Ratio; Ontology; Organism; Parvalbumins; Paste substance; Pathogenesis; Pathway Analysis; Pathway interactions; Patients; Persons; Population; Prefrontal Cortex; Procedures; Proteins; Psychotic Disorders; Public Health; RNA; RNA Polymerase II; RNA-Directed DNA Polymerase; Regulation; Rest; Retrotransposon; Risk; Sample Size; Sampling; Schizophrenia; Scientist; Site; Staining method; Stains; Suicide; Synapses; Technology; Testing; Tissues; Transcriptional Activation; Translations; Variant; base; digital; disability; emerging adult; endonuclease; gene function; genome wide association study; hippocampal pyramidal neuron; kindred; mortality; neuroimaging; promoter; public health relevance; reference genome; transposon/insertion element; young adult

 DESCRIPTION (provided by applicant): Schizophrenia (SZ) is a common, chronic group of psychotic brain disorders, affecting 1% of the US population, creating a significant public health problem because of the associated disability, morbidity and mortality. The pathogenesis of SZ is poorly understood, but it is thought to be a neurodevelopmental disorder. Neuroimaging evidence has accumulated to indicate that the dorsolateral prefrontal cortex (DLPFC) functions abnormally in SZ, both when activated and at rest. Further, EEG evidence has identified abnormalities of γ oscillations in SZ, suggesting that parvalbumin positive GABAergic interneuron regulation of cortical pyramidal neurons is dysfunctional. In the past 5 years, data have accumulated to prove that activation of LINE1 (L1) retrotransposons (RTPs) may occur wherever chromatin assumes a relaxed state to permit transcriptional activation. The abnormal DLPFC activation and γ oscillations in SZ may drive chronic aberrant transcriptional activation, thereby providing opportunities for L1s to retrotranspose in the developing CNS. These somatic de novo L1s may interfere with normal neuronal activity because they have inserted into a gene needed by that neuron for normal function. If one or more functional de novo L1s occur early in CNS development, all the daughter neurons that derive from that neuronal precursor will also carry the L1, perhaps leading to a dysfunctional population of neurons destined to increase risk for SZ. This project builds on preliminary results indicating that intragenic de novo L1s are found significantly more often in SZ DLPFC neuronal DNA in relevant gene ontologies (eg, synapse part and plasma membrane part; p = ~10-4), compared to control tissue. This proposal will employ DLPFC and anterior cingulate tissue (obtained at autopsy) from 100 SZ patients and 100 matched controls. Using differential centrifugation and fluorescence assisted cell sorting (FACS), followed by PCR to enrich the DNA for L1 sequences, DNA amplicons will be sequenced and aligned to the reference genome to detect intragenic de novo (not in the reference genome) L1s in brain-expressed genes. Allele frequencies of the most promising intragenic de novo L1s will be estimated by droplet digital PCR. Selected intragenic de novo L1s will be re-created in neuronal cell lines via CRISPR/Cas9 technology. The functional effect of the de novo L1 on transcription and translation of the gene will be determined. In this manner, it is expected that intragenic de novo L1s, which increase risk for SZ, will be discovered.

 描述（由申请人提供）：精神分裂症 (SZ) 是一种常见的慢性精神性脑部疾病，影响 1% 的美国人口，由于相关的残疾、发病率和死亡率而造成严重的公共健康问题。 SZ 的发病机制尚不清楚，但它被认为是一种神经发育障碍。神经影像学证据表明，背外侧前额叶皮层 (DLPFC) 在 SZ 中功能异常，无论是激活时还是休息时。此外，EEG 证据发现 SZ 中 γ 振荡异常，表明小白蛋白阳性的皮质锥体神经元的 GABA 能中间神经元调节功能失调。 在过去的 5 年中，积累的数据证明，只要染色质呈现松弛状态以允许转录激活，LINE1 (L1) 逆转录转座子 (RTP) 的激活就可能发生。 SZ 中异常的 DLPFC 激活和 γ 振荡可能会驱动慢性异常转录激活，从而为发育中的 CNS 中的 L1 逆转录转座提供机会。这些体细胞从头 L1 可能会干扰正常的神经元活动，因为它们已插入该神经元正常功能所需的基因中。如果在 CNS 发育早期出现一个或多个功能性的从头 L1，则源自该神经元前体的所有子神经元也将携带 L1，可能会导致神经元群体功能失调，从而增加 SZ 的风险。 该项目建立在初步结果的基础上，初步结果表明，与对照组织相比，在相关基因本体（例如突触部分和质膜部分；p = ~10-4）的 SZ DLPFC 神经元 DNA 中发现基因内从头 L1 的频率明显更高。该提案将采用来自 100 名 SZ 患者和 100 名匹配对照者的 DLPFC 和前扣带回组织（尸检时获得）。使用差速离心和荧光辅助细胞分选 (FACS)，然后通过 PCR 富集 DNA 中的 L1 序列，对 DNA 扩增子进行测序并与参考基因组进行比对，以检测脑表达基因中的基因内从头（不在参考基因组中）L1。最有希望的基因内从头 L1 的等位基因频率将通过液滴数字 PCR 进行估计。选定的基因内从头 L1 将通过 CRISPR/Cas9 技术在神经元细胞系中重新创建。将确定 de novo L1 对基因转录和翻译的功能影响。通过这种方式，预计会发现增加 SZ 风险的基因内从头 L1。