本文選題：代謝綜合征 + 全基因組關(guān)聯(lián)分析　； 參考：《浙江大學(xué)》2017年博士論文

【摘要】：代謝綜合征是一組代謝指標(biāo)異常癥候群,主要表現(xiàn)為中心性肥胖、高血壓和糖脂代謝紊亂。研究表明,代謝綜合征會(huì)增加心腦血管疾病、糖尿病的發(fā)病風(fēng)險(xiǎn)。按中國(guó)糖尿病協(xié)會(huì)定義,2010年我國(guó)居民代謝綜合征患病率達(dá)17.6%。雙生子研究表明,遺傳因素在代謝異常的發(fā)生中起了重要的作用。經(jīng)典的全基因組關(guān)聯(lián)研究(genome-wide association study,GWAS)已發(fā)現(xiàn)了一些與代謝異常相關(guān)的遺傳變異,但仍存在以下兩大問題:(1)已發(fā)現(xiàn)的位點(diǎn)僅能解釋少量的遺傳度,即存在大量丟失的遺傳度(missing heritability)有待進(jìn)一步探究;(2)以往研究主要關(guān)注表型相關(guān)的標(biāo)簽SNP(tag SNP)及與之距離最近的基因,但后續(xù)研究發(fā)現(xiàn),大部分的定位信號(hào)并不通過與之距離最近的基因起作用,如現(xiàn)已證明,定位在FTO上的肥胖相關(guān)位點(diǎn)主要通過遠(yuǎn)程調(diào)控IRX3基因表達(dá)影響表型�？傊�,以往對(duì)GWAS定位信號(hào)的解析非常有限,多數(shù)關(guān)鍵基因和效應(yīng)位點(diǎn)(causal variant)未被找到,不利于后續(xù)功能研究。近些年研究發(fā)現(xiàn),大部分表型相關(guān)遺傳變異通過調(diào)控基因表達(dá)水平進(jìn)而影響表型。若以基因表達(dá)調(diào)控作為注釋,可以將全基因組的篩選范圍縮小到基因表達(dá)數(shù)量性狀基因座(expression Quantitative Trait Loci,eQTLs)上,提高發(fā)現(xiàn)表型相關(guān)位點(diǎn)的效率。ENCODE、GTEx、Roadmap等研究項(xiàng)目陸續(xù)公布了大量可用于注釋遺傳變異對(duì)表型調(diào)控路徑中多個(gè)組學(xué)的信息,包括具有組織特異性的基因表達(dá)、DNA甲基化、組蛋白修飾、轉(zhuǎn)錄因子結(jié)合等。這些信息可用于進(jìn)一步精確篩選表型相關(guān)遺傳變異,也可用于對(duì)已知信號(hào)的重新解析。以往的研究主要以歐裔人群為主,在我國(guó)漢族人群中的研究還很有限,由于不同人種間存在遺傳背景的差異,歐裔人群的研究結(jié)果不能直接外推至中國(guó)漢族人群。為了系統(tǒng)地探索代謝綜合征及代謝組分相關(guān)的遺傳易感位點(diǎn),本研究以我國(guó)漢族人群為主要研究對(duì)象,通過基于多組學(xué)注釋的全基因組關(guān)聯(lián)分析,發(fā)現(xiàn)并解析代謝異常相關(guān)的遺傳變異。研究?jī)?nèi)容分為三個(gè)部分(圖1):第一部分采用經(jīng)典的全基因組關(guān)聯(lián)分析策略,篩選與代謝綜合征及代謝組分關(guān)聯(lián)信號(hào)最強(qiáng)的遺傳變異位點(diǎn)并進(jìn)行多階段驗(yàn)證、功能分析和基因環(huán)境交互效應(yīng)分析。第二部分在第一部分研究的基礎(chǔ)上,通過對(duì)轉(zhuǎn)錄調(diào)控的注釋,進(jìn)一步篩選與糖脂代謝相關(guān)的遺傳變異,通過精細(xì)定位和功能實(shí)驗(yàn),解析基本調(diào)控路徑。第三部分針對(duì)已知的糖脂代謝關(guān)聯(lián)信號(hào),利用最新的多組學(xué)注釋信息對(duì)信號(hào)區(qū)域進(jìn)行系統(tǒng)化的重解析,探究關(guān)鍵的調(diào)控基因,為關(guān)聯(lián)分析的成果轉(zhuǎn)化提供支持。圖1研究?jī)?nèi)容示意圖左上側(cè)圖為課題組開展的代謝綜合征GWAS關(guān)聯(lián)信號(hào)(P值)的曼哈頓圖,圖下左側(cè)為第一部分技術(shù)路線,右側(cè)為第二部分技術(shù)路線。右側(cè)圖為NHGRI-EBI GWAS catalog記錄的表型相關(guān)的位點(diǎn),為研究的第三部分,采用多組學(xué)注釋,對(duì)與代謝異常相關(guān)的位點(diǎn)進(jìn)行系統(tǒng)化重解析。第一部分代謝綜合征的遺傳易感性研究一、研究目的:在中國(guó)漢族人群中篩選并驗(yàn)證代謝綜合征相關(guān)的遺傳易感性位點(diǎn)。二、材料和方法:以單核苷酸多態(tài)性位點(diǎn)為遺傳變異標(biāo)記,首先在杭州蕭山地區(qū)1742例樣本中采用全基因組關(guān)聯(lián)分析的方法,篩選出與代謝綜合征及代謝組分關(guān)聯(lián)信號(hào)最強(qiáng)的位點(diǎn),然后對(duì)這些位點(diǎn)進(jìn)行多階段獨(dú)立樣本驗(yàn)證,驗(yàn)證樣本來自我國(guó)東部、北部、東北部等多個(gè)地區(qū),共計(jì)10978例。合并多階段驗(yàn)證結(jié)果后,對(duì)達(dá)到全基因組陽(yáng)性水平的位點(diǎn)進(jìn)行功能預(yù)測(cè)、基因環(huán)境交互效應(yīng)分析。三、研究結(jié)果:通過代謝綜合征的全基因組關(guān)聯(lián)分析及多階段獨(dú)立樣本的驗(yàn)證,研究發(fā)現(xiàn)位于APO45上的rs651821位點(diǎn)和位于ADF2上亞洲人特有的高頻錯(cuò)義突變位點(diǎn)rs671的基因型與代謝綜合征的遺傳易感性相關(guān)。在控制了 APO4基因簇區(qū)域內(nèi)最強(qiáng)信號(hào)rs651821后,位于BUD13上的rs180326位點(diǎn)仍然與血清甘油三酯(triglyceride,TG)水平相關(guān)(Pconbined=2.4E-08),是APO4基因簇內(nèi)一個(gè)新的第二信號(hào)(secondary signal)。在整合了 遺傳變異位點(diǎn) rs651821、rsl80326、血清 APOA5、BUD13的蛋白水平、TG水平后分析發(fā)現(xiàn),除了APO外,BUD13也參與了血清TG水平的調(diào)控。此外,研究發(fā)現(xiàn)rs671位點(diǎn)的多態(tài)性不僅會(huì)通過影響乙醛代謝影響人們的飲酒行為,該位點(diǎn)還與飲酒行為之間存在對(duì)代謝綜合征及相關(guān)代謝表型的交互效應(yīng),其效應(yīng)主要存在于飲酒人群中。四、小結(jié):(1)在中國(guó)漢族人群中,rs651821(APOA5)和rs671(ALH2)位點(diǎn)的基因型與代謝綜合征的遺傳易感性相關(guān);(2)rs180326(BUD13)基因型與血清TG水平相關(guān),其效應(yīng)獨(dú)立于已知位點(diǎn)rs651821(APOA5);(3)rs671(ALDH2)的基因型與飲酒行為存在交互效應(yīng)。第二部分基于注釋信息的糖脂代謝相關(guān)遺傳變異篩選及精細(xì)定位一、研究目的:在第一部分研究的基礎(chǔ)上,結(jié)合多組學(xué)注釋信息,進(jìn)一步篩選和驗(yàn)證糖脂代謝指標(biāo)相關(guān)的遺傳變異位點(diǎn);對(duì)新發(fā)現(xiàn)的糖脂代謝相關(guān)遺傳變異位點(diǎn)進(jìn)行精細(xì)定位,并通過功能實(shí)驗(yàn)確認(rèn)效應(yīng)位點(diǎn)。二、材料和方法:首先,通過對(duì)1742例樣本(同第一部分)的糖脂代謝的全基因組關(guān)聯(lián)分析,得到與表型相關(guān)的但又未被第一部分驗(yàn)證過的遺傳變異位點(diǎn);然后分析這些位點(diǎn)與脂肪、肝臟、胰島和骨骼肌中基因表達(dá)水平的關(guān)聯(lián),并將這些位點(diǎn)的信號(hào)與表型關(guān)聯(lián)信號(hào)共定位,找出與糖脂代謝表型相關(guān)的eQTLs;進(jìn)一步通過多階段獨(dú)立樣本驗(yàn)證基因型與表型的關(guān)聯(lián);對(duì)驗(yàn)證達(dá)到全基因組陽(yáng)性水平的位點(diǎn),通過以基因?yàn)閱挝坏姆治霾呗?gene-based analyses)推測(cè)其可能的調(diào)控基因;并通過ENCODE和Roadmap計(jì)劃提供的相關(guān)組織細(xì)胞中染色質(zhì)狀態(tài)、組蛋白修飾、轉(zhuǎn)錄因子結(jié)合信號(hào)推測(cè)調(diào)控活性區(qū)域及其對(duì)應(yīng)的效應(yīng)位點(diǎn);最后構(gòu)建包含不同等位基因的載體,通過熒光素酶報(bào)告基因?qū)嶒?yàn)確認(rèn)其表達(dá)調(diào)控作用。三、研究結(jié)果:通過全基因組關(guān)聯(lián)分析及eQTL的注釋,發(fā)現(xiàn)了 22個(gè)與糖脂代謝相關(guān)的遺傳變異位點(diǎn)在特定的組織中影響基因表達(dá),多階段獨(dú)立樣本驗(yàn)證確認(rèn)了 rsl880118位點(diǎn)與血清HDL-C水平的關(guān)聯(lián)(Pcombined= 1.4E-10)。該位點(diǎn)可以tag的區(qū)域主要包括DAGLB和RAC1兩個(gè)基因,在加性模型下,rsl880118的基因型可以解釋DAGLB(diacylglycerol lipase,beta)基因在皮下脂肪組織中表達(dá)水平變異的47.7%(P =5.9E-42)。同時(shí),通過TWAS、SMR、Sherlock等以基因?yàn)閱挝坏难芯糠椒?我們發(fā)現(xiàn)D4GLB基因與血清HDL-C水平之間存在關(guān)聯(lián),關(guān)聯(lián)的P值分別為3.0E-08、1.1E-04和1.6E-06。進(jìn)一步通過組蛋白信號(hào)H3K27ac、H3K4me3,H3K9ac及轉(zhuǎn)錄因子結(jié)合區(qū)域、DNA酶Ⅰ超敏位點(diǎn)的定位,找到了位于DAGLB基因5'區(qū)域的調(diào)控活性片段,熒光素酶報(bào)告基因的結(jié)果顯示,該活性片段中rs4724806位點(diǎn)(與rs1880118位點(diǎn)LD r2 = 0.77)可能是真正的效應(yīng)位點(diǎn),其最小等位基因會(huì)增加轉(zhuǎn)錄活性,與eQTL分析的結(jié)果一致。四、小結(jié):rsl880118是一個(gè)在中國(guó)漢族人群中新發(fā)現(xiàn)的與血清HDL-C水平相關(guān)的遺傳變異位點(diǎn),其效應(yīng)位點(diǎn)rs4724806通過調(diào)控轉(zhuǎn)錄活性影響DAGLB基因表達(dá),該部分研究提示了 A4GLB基因在脂代謝中的作用。第三部分利用多組學(xué)注釋系統(tǒng)解析糖脂代謝相關(guān)的遺傳變異一、研究目的:結(jié)合多組學(xué)注釋信息,對(duì)已知的糖脂代謝相關(guān)遺傳變異信號(hào)區(qū)域進(jìn)行系統(tǒng)的解析,探究基因型到表型的調(diào)控路徑,為后續(xù)功能研究提供支持。二、研究方法:首先,整理已報(bào)道的與糖脂代謝相關(guān)的遺傳變異位點(diǎn);然后,利用千人基因組計(jì)劃提供的位點(diǎn)間連鎖不平衡信息,填補(bǔ)出所有與lead SNP高度連鎖不平衡的位點(diǎn)用于后續(xù)的精細(xì)定位和功能預(yù)測(cè);接著,對(duì)這些位點(diǎn)進(jìn)行系統(tǒng)的多組學(xué)注釋,包括基因表達(dá)、染色質(zhì)狀態(tài)、DNA甲基化、組蛋白修飾、轉(zhuǎn)錄因子結(jié)合等。對(duì)位于編碼區(qū)域的遺傳變異位點(diǎn),再進(jìn)行物種間保守性估計(jì)、翻譯后修飾等翻譯水平的注釋;,最后,整理推測(cè)出可能的基因型到表型調(diào)控路徑。三、研究結(jié)果:對(duì)經(jīng)篩選過濾后得到的592個(gè)糖脂代謝相關(guān)遺傳變異位點(diǎn)進(jìn)行填補(bǔ)后,共計(jì)17646個(gè)位點(diǎn)納入后續(xù)精細(xì)定位分析(LD r20.5)。在轉(zhuǎn)錄水平,通過遺傳變異與基因表達(dá)相關(guān)的注釋,發(fā)現(xiàn)了 104個(gè)在內(nèi)臟脂肪、肝臟、骨骼肌或胰島細(xì)胞中與基因表達(dá)水平相關(guān)的位點(diǎn)。同時(shí),發(fā)現(xiàn)了一些與特定環(huán)境刺激相關(guān)的eQTLs,如rs702485位點(diǎn)與DAGLB基因表達(dá)的關(guān)聯(lián)僅出現(xiàn)在LPS刺激后(Pbefore0.05,Pafter=2.52E-16)。133個(gè)糖脂代謝相關(guān)位點(diǎn)與脂肪組織或胰島細(xì)胞中的DNA甲基化相關(guān),其中許多位點(diǎn)與多個(gè)CpG位點(diǎn)的甲基化水平相關(guān)。經(jīng)過對(duì)遺傳變異位點(diǎn)的精細(xì)定位,我們發(fā)現(xiàn)有49個(gè)位點(diǎn)可以關(guān)聯(lián)(tag)到一個(gè)或以上的位于組蛋白修飾信號(hào)峰區(qū)域內(nèi)的位點(diǎn),且具有組織特異性。在翻譯水平,有122個(gè)(r20.5)或43個(gè)(r20.8)位點(diǎn)可以關(guān)聯(lián)到一個(gè)或以上的非同義突變位點(diǎn),其中有16個(gè)位點(diǎn)經(jīng)SIFT和Polyphen注釋均提示存在影響蛋白功能的可能。對(duì)糖代謝相關(guān)位點(diǎn)rsl535500精細(xì)定位和功能預(yù)測(cè)后發(fā)現(xiàn),該位點(diǎn)G到T的變異與附近7個(gè)CpG位點(diǎn)的存在相關(guān)聯(lián),這些位點(diǎn)靠近KCNK17基因5'端的CpG島,存在影響DNA甲基化的可能。通過對(duì)胰島細(xì)胞中多個(gè)組學(xué)的信息整合,發(fā)現(xiàn)該位點(diǎn)確實(shí)可以通過影響附近位點(diǎn)甲基化水平進(jìn)而影響KCNK17的表達(dá),這不同于以往報(bào)道認(rèn)為該信號(hào)主要通過KCNK16起作用。四、小結(jié):(1)通過整合多個(gè)組學(xué)信息,對(duì)糖脂代謝GWAS信號(hào)進(jìn)行重新解析后發(fā)現(xiàn)了許多可能參與到遺傳變異影響表型調(diào)控路徑中的基因及調(diào)控元件。與其他復(fù)雜表型類似,經(jīng)過注釋后,三分之一的基因與以往對(duì)該位點(diǎn)報(bào)道一致。(2)僅有7%-20%的糖脂代謝表型相關(guān)遺傳變異位點(diǎn)可以關(guān)聯(lián)到一個(gè)或以上的非同義或無義突變,其余可能通過轉(zhuǎn)錄水平影響表型。(3)通過對(duì)糖代謝相關(guān)的位點(diǎn)rs1535500的精細(xì)定位,發(fā)現(xiàn)該位點(diǎn)G到T的變異與附近位點(diǎn)CpG位點(diǎn)的存在相關(guān)聯(lián),進(jìn)而影響甲基化水平調(diào)控KCNK17的表達(dá)。結(jié)論:基于以上三部分內(nèi)容,得出以下結(jié)論:(1)在中國(guó)漢族人群中,rs651821(APO45)和rs671(ALDH2)位點(diǎn)的基因型與代謝綜合征的遺傳易感性相關(guān);新發(fā)現(xiàn)的rs180326(BUD13)位點(diǎn)與血清TG水平的關(guān)聯(lián)獨(dú)立于區(qū)域內(nèi)已知位點(diǎn)rs651821;rs671與飲酒行為存在交互效應(yīng)。(2)結(jié)合轉(zhuǎn)錄調(diào)控注釋信息可優(yōu)化GWAS的篩選策略;rs1880118是一個(gè)在中國(guó)漢族人群中新發(fā)現(xiàn)的與血清HDL-C水平相關(guān)的遺傳變異位點(diǎn),與該位點(diǎn)高度連鎖不平衡的rs4724806位點(diǎn)多態(tài)性會(huì)通過調(diào)控DAGLB基因表達(dá)影響表型。(3)80%以上的糖脂代謝相關(guān)位點(diǎn)主要通過調(diào)控轉(zhuǎn)錄水平影響表型;經(jīng)多組學(xué)注釋推測(cè)的調(diào)控基因中有三分之一與原GWAS報(bào)道的基因一致,如文獻(xiàn)報(bào)道糖代謝相關(guān)位點(diǎn)rs1535500可能的調(diào)控基因?yàn)镵CNK16,但注釋信息提示該位點(diǎn)可能通過影響甲基化水平調(diào)控KCNK17的表達(dá)進(jìn)而影響表型。
[Abstract]:Metabolic syndrome is a group of abnormal metabolic syndrome syndrome, mainly characterized by central obesity, hypertension and glucose and lipid metabolism disorder. Studies have shown that metabolic syndrome can increase the risk of cardiovascular and cerebrovascular diseases and diabetes. According to the definition of China Diabetes Association, the prevalence rate of metabolic syndrome in Chinese residents in 2010 was 17.6%. twins. Genetic factors play an important role in the occurrence of metabolic abnormalities. The classical genome-wide association study (GWAS) has found some genetic variations associated with metabolic abnormalities, but there are still two major problems: (1) the found loci can only explain a small number of heritability, that is, there is a large number of lost heredity. Missing heritability remains to be further explored; (2) previous studies mainly focus on phenotypic related label SNP (tag SNP) and the closest genes to it, but subsequent studies have found that most of the location signals do not play a role in the nearest gene, for example, the obesity related loci on FTO have been mainly passed through. Long distance regulation of IRX3 gene expression affects phenotypes. In a word, the previous analysis of GWAS localization signals is very limited. Most of the key genes and effect sites (causal variant) have not been found, which is not conducive to the follow-up function study. In recent years, most of the phenotypic related genetic variations have passed the regulation gene expression level and then affect the phenotype. As a note of expression regulation, the screening range of the whole genome can be narrowed to the gene expression quantitative trait loci (expression Quantitative Trait Loci, eQTLs) to improve the efficiency of the detection of the phenotypic related sites,.ENCODE, GTEx, Roadmap and other research projects have been published in a large number of studies that can be used to annotate genetic variation in the pathway of phenotypic regulation. The information of multiple histones, including tissue specific gene expression, DNA methylation, histone modification, transcription factor binding, and so on. These information can be used to further accurately screen phenotypic related genetic variations and can be used to reinterpret known signals. Because of the differences in genetic background between different human species, the results of the European population can not be directly extrapolated to the Chinese Han population. In order to systematically explore the metabolic syndrome and the genetic susceptibility loci related to the metabolic components, the main object of this study is to study the whole base of the Chinese Han population based on the multi group annotation. The genetic variation related to metabolic abnormalities was found and resolved by group association analysis. The study was divided into three parts (Figure 1): in the first part, the classical whole genome association analysis strategy was used to screen the strongest genetic variation loci of metabolic syndrome and metabolic components and carry out multistage validation, functional analysis and genetic environment interaction. The second part, on the basis of the first part of the study, further screened the genetic variation related to glycolipid metabolism through the annotation of transcriptional regulation, and analyzed the basic regulation path through fine location and functional experiments. The third part aimed at the known glycolipid metabolic signals, using the latest multi group annotation information to the signal. The region carries out systematic reanalysis, explores key regulatory genes, and provides support for the transformation of correlation analysis. The upper left side map of Figure 1 is the Manhattan map of the metabolic syndrome associated signal (P value) carried out by the project group. The left side is the first part of the technical route and the right side is the second part of the technical route. The right side map is the right map. 1 The third part of the phenotypic related loci recorded by NHGRI-EBI GWAS catalog, which is the third part of the study, uses a multi - group annotation to systematically re - analyze the sites associated with metabolic disorders. Susceptibility loci. Two, materials and methods: using single nucleotide polymorphic loci as genetic variation markers, the best loci with metabolic syndrome and metabolic components were screened in 1742 samples of Xiaoshan, Hangzhou, with the method of full genome association analysis. The sample is from the eastern, northern, northeastern and other regions of China, with a total of 10978 cases. After the combined multistage validation results, the functional prediction of the positive levels of the whole genome, the analysis of the interaction effect of the gene environment. Three, the results of the study: through the whole genome association analysis of the metabolic syndrome and the verification of the multi stage independent samples, The study found that the genotype of the rs651821 site on APO45 and the genotype rs671 of the high frequency missense mutation site specific to Asians on ADF2 is related to the genetic susceptibility to metabolic syndrome. After controlling the strongest signal rs651821 in the APO4 gene cluster region, the rs180326 site on BUD13 is still with the serum triglyceride (triglyceride, TG) water. Flat correlation (Pconbined=2.4E-08) is a new second signal (secondary signal) in the APO4 gene cluster. After integrating the genetic variation loci rs651821, rsl80326, serum APOA5, BUD13 protein level and TG level after analysis, it is found that BUD13 also participates in the regulation of serum TG levels except APO. Furthermore, the polymorphism of the locus is found not. Only by influencing the metabolism of acetaldehyde affects people's drinking behavior, and the interaction effect on metabolic syndrome and related metabolic phenotypes exists between the site and drinking behavior. The effect is mainly in the drinking crowd. Four, (1) the genotype and metabolic syndrome of rs651821 (APOA5) and rs671 (ALH2) loci in Chinese Han population Genetic susceptibility is related; (2) rs180326 (BUD13) genotype is related to serum TG level, its effect is independent of the known site rs651821 (APOA5); (3) the genotype of rs671 (ALDH2) has interaction effects with drinking behavior. The second part is based on the annotated information of glycolipid Xie Xiang pass genetic mutation screening and fine location one. The purpose of this study is to study the first part of the study On the basis of the study, the genetic variation loci related to glycolipid metabolic indices were further screened and verified with multi group annotation information, and the newly discovered genetic mutation sites related to glycolipid metabolism were fine located, and the functional loci were confirmed by functional experiments. Two, materials and methods: first, the glycolipid of the 1742 samples (the same part) was obtained. Genetic variation sites associated with phenotypic but not verified by the first part of metabolism; then the association of these sites with the gene expression levels in the fat, liver, islets, and skeletal muscles and the co location of signals from these sites with phenotypic correlation signals to identify eQ related to the glycolipid metabolic phenotype. TLs; further verify the association between genotypes and phenotypes by multistage independent samples; to verify the locus of the positive level of the whole genome and to speculate on the possible regulatory genes by the analysis strategy (gene-based analyses) based on the basis of the unit; and the chromatin state in the related tissue cells provided by the ENCODE and Roadmap plan, and the group of eggs White modification, the transcription factor binding signal conjectured the active region and its corresponding effect loci; finally, the vector containing the different alleles was constructed, and the expression regulation was confirmed by the luciferase reporter gene experiment. Three, the results were found to be related to the metabolism of glycolipid by whole genome association analysis and eQTL annotation. Genetic variation loci affect gene expression in specific tissues. Multi stage independent sample validation confirms the association of rsl880118 loci with serum HDL-C levels (Pcombined= 1.4E-10). The loci can mainly include two genes of DAGLB and RAC1. Under the additive model, the genotype of rsl880118 can explain DAGLB (diacylglycerol Li). Pase, beta) gene expression in subcutaneous adipose tissue is 47.7% (P =5.9E-42). At the same time, we find that there is a association between the D4GLB gene and the serum HDL-C level by TWAS, SMR, Sherlock and so on. The P values of the associated P are 3.0E-08,1.1E-04 and 1.6E-06. further through the histone signals. E3, H3K9ac and transcription factor binding region, the location of DNA enzyme I hypersensitivity loci, found a regulatory active fragment located in the 5'region of the DAGLB gene. The results of the luciferase reporter gene show that the rs4724806 locus (and rs1880118 LD R2 = 0.77) in the active fragment may be a true effect site, and its minimum allele will increase the transcriptional activity. Sex, in accordance with the results of eQTL analysis. Four, summary: rsl880118 is a newly discovered genetic locus of variation associated with serum HDL-C levels in Chinese Han population. Its effect locus rs4724806 affects the expression of DAGLB gene by regulating transcriptional activity. This part of the study suggests the role of A4GLB gene in lipid metabolism. The third part is used in this study. The multi group annotation system parses the genetic variation related to glycollipid metabolism. The purpose of this study is to systematically analyze the known region of genetic variation signals related to glycolipid metabolism by combining multi group annotation information to explore the regulation path of genotype to phenotype to provide support for subsequent functional studies. Two, research methods: first, the report has been reported. Genetic variation loci associated with glycolipid metabolism; then, using interloci linkage disequilibrium information provided by the Millennium genome project to fill out all the highly linked unbalanced sites with lead SNP for subsequent fine location and functional prediction; and then systematically annotate these loci, including gene expression, chromatin State, DNA methylation, histone modification, transcription factor binding, etc., annotations to the genetic variation loci in the coding region, interspecies conservatism estimation, post translation modification and other translation levels; finally, the possible genotype to phenotypic regulation path was conjectured. Three, research results: 592 glycolipids obtained after screening and filtering. After the metabolism related genetic mutation sites were filled, a total of 17646 loci were included in the follow-up fine location analysis (LD r20.5). At the transcriptional level, 104 loci related to the gene expression level in visceral fat, liver, skeletal muscle, or islet cells were found by the annotation related to genetic variation and gene expression. EQTLs related to specific environmental stimuli, such as the association of rs702485 loci with DAGLB gene expression, only occurs after LPS stimulation (Pbefore0.05, Pafter=2.52E-16).133 glycolipid related sites are associated with DNA methylation in adipose tissue or islet cells, many of which are related to the level of methylation at multiple CpG sites. We found that 49 loci can associate (tag) to one or more loci in the peak region of the histone modified signal, and have tissue specificity. At the translation level, 122 (r20.5) or 43 (r20.8) loci can be associated with one or more of the non synonymous mutation sites, of which 16 loci are in SI. FT and Polyphen notes suggest the possibility of affecting protein function. After the fine localization and function prediction of the glucose metabolism related site rsl535500, the mutation of the site G to T is associated with the existence of the 7 CpG loci near the KCNK17 gene, which is near the CpG island of the 5'terminal of the KCNK17 gene, and the possibility of the DNA methylation is possible. It is found that this site can affect the expression of KCNK17 by influencing the methylation level of the nearby loci, which is different from the previous reports that the signal is mainly mediated by KCNK16. Four, a summary: (1) a number of GWAS signals are reinterpreted by integrating a number of Informatics, and many can be found. Genes and regulatory elements that can participate in genetic variation affecting the pathway of phenotypic regulation. Similar to other complex phenotypes, after annotation, 1/3 of the genes are consistent with the previous reports. (2) only 7%-20%'s glycolipid phenotype related genetic variation loci can be associated with one or more unsynonymous or nonsense mutations. The phenotype may be affected by transcription level. (3) by mapping the site of rs1535500 related to glycometabolism, it is found that the locus G to T

【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：R589

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