模式生物与人类疾病

模式生物与人类疾病2011.10.24-25）王铸钢上海交通大学医学院附属瑞金医院医学基因组学国家重点实验室上海交通大学医学院医学遗传学学科上海瑞金医院实验医学研究中心上海南方模式生物研究中心―Lifebook‖—composedof4letters(A、G、C、T)5letters/word=====~600,000,000words12words/line=====~50,000,000lines70lines/page======~700,000pages1000pages/volume======~700volumes―GenomeEncyclopedia‖ofHumanHumanVariationandHeredity—weareall99.9%thesame,DNA-wise!differatabout1inevery1,200to1,500pb,orlettersTwinslookthesameaseachother!TheGenomedetermines……LifeCycle—mitosisandmeiosisFromDNAtoChromosomeThegenomeiscondensedandpackagedintochromosomesCopynumber:100-10,000/cell(100-1000/sperm,100000-1000000/egg)mtDNA:16569bpHC:28genesLC:9genes13proteins22tRNA2rRNAHumanMitochondriaDNACentralDogmaofGeneticsAllmedicineisgenetics---MckusickVA.Medicineinfutureismoleculargenetics---RenatoDulbeccoDulbeccoR.,Aturningpointincancerresearch:Sequencingthegenome,Science,1986,231:1055-56.TypeAutosomalX-linkedY-linkedMitochondrialTotalGenewithknownsequence12475611483513169Genewithknownsequenceandphenotype3462002368Phenotypedescription,molecularbasisknown25762274282835Mendelianphenotypeorlocus,molecularbasisunknown1636136501777Other,mainlyphenotypeswithsuspectedmendelianbasis1853134201989Total188861128596520138OMIMStatisticsforAugust14,2010NumberofEntries模式生物与人类TechnicaldifficultyComplexityDrosophilazebrafishmousehumanC.elegansyeast模式生物的定义：生物体由低等向高等、由简单向复杂的进化过程中，很多生物学现象是非常保守的。低等生物的生命现象及分子基础与高等生物相似。因此，便于试验研究人类生命现象或健康问题的其他物种就称为模式生物。最常用的几种模式生物斑马鱼线虫小鼠果蝇•最小的哺乳动物之一(25-40g)•生物进化上与人类接近(60-75百万年)•胎盘形成和早期胚胎发育与人类相近•组织器官结构和细胞功能与人类相似•小鼠基因组测序计划已完成•人类99%的基因存在于小鼠，基因同源性高达78.5%•基因组93%的区域基因排列顺序与人类相同相同•基因组改造的技术手段成熟小鼠作为模式生物的优势Whyme?•基因组–染色体数19对常染色体+XY–双倍体DNA含~6pg–重组单元1600cM–基因数量30-35,000•生殖生物学–怀孕期3周–哺乳期3周–性成熟年龄7周–饲养寿命1.5-2.5年–平均每窝产仔6-9个–雌鼠生育4-8窝小鼠生理特性人类小鼠人和小鼠基因组的比较模式生物技术原理和表型分析疾病相关或功能未知基因功能获得―加法”“减法”功能缺失?转基因小鼠技术流程Dna.mpg基因剔除技术流程Es.mpg人类白血病相关融合基因的转基因小鼠发生类白血病表型W.T.PML-RARaPLZF-RARaNPM-RARa人乙肝病毒基因组转基因小鼠体内持续表达乙肝病毒抗原JournalofMedicalVirology2006;78:551-560.JournalofMedicalVirology2007；79:1478–1484.020406080100020406080100120140160180200Age(days)%oftumorfreeanimalswt(n=20)Line9(n=19)Line42(n=33)PLAG1OverexpressionLeadstoSalivaryGlandPleomorphicAdenomasinMiceInternationalJournalofCancer2006;118(3):643-648.TgTuc-Mycb-cateninPLAG1TheCytoplasmicDomainofMUC1InducesHyperplasiaintheMammaryGlandandCorrelateswithNuclearAccumulationofβ-cateninLiYetal.PLoSONE，2011TheMDM2/MDMXcomplexisrequiredforthecontrolofp53activityinvivoHuangL,etal.PNASrevised.SYNSFgf9wt/wtFgf9wt/S99NFgf9wt/S99Nwt+/-+/-AbnormalitiesintoothandtaildevelopmentinFgf9wt/S99Nmice靶基因表达调控研究的双通道转基因小鼠模型IL-1bpromoter-Luc转基因小鼠EGFP转基因小鼠LPS5'-UTRLucSV40PAInsulatorpCAGEGFP3-UTR化学发光代表靶基因转录水平的表达调控绿色荧光反映内源性miRNA通过靶基因mRNA3‗-UTR对蛋白质表达水平的调控新生鼠心脏脑肾胰腺睾丸Palladin基因剔除致小鼠脑外翻及胚胎造血功能障碍wtM+wtEwtM+mtEmtM+wtEmtM+mtEF4/80Ter119Merge正常小鼠胚胎脑外翻小鼠胚胎MCN2005,29:507;JCB2007,100:1288;Blood2007,110:870.Rig-I基因剔除导致小鼠抗细菌免疫功能障碍Genotypewt+/--/-Sex♂♀total♂♀total♂♀totaln494897395190395089Eyeinfection000000101525%00000025.63028.1Skininfection101011437%20101.91.110.367.8Rig-I基因剔除致小鼠免疫球蛋白IgG3缺失和B淋巴细胞发育障碍29.226.019.410.027.042.0B220+wt-/-IgDIgMB220+T1T2T3T1T2T3B220+AA4.1+IgMCD23B220+AA4.1+B220+AA4.1-MZMZFBFBIgMCD23B220+AA4.1-01020304050607080wt(n=3)-/-(n=4)Percentage(%)T1T2T3MZFO0102030405060wt(n=3)-/-(n=4)Percentage(%)B220+MD0200400600800100012001400wt(n=4)-/-(n=5)IgMIgG1IgG2aIgG2bIgG3IgA(/10)(/10)SerumIgs(mg/ml)*024681012wt(n=4)-/-(n=4)IgMIgG1IgG2aIgG2bIgG3IgALPS-inducedIgsecretion(folds)*结肠炎小鼠模型CellResearch2007,17(10):858-868.ABIncreasedfatmassandinsulinresistanceinmicelackingpancreaticlipase-relatedprotein1.JNutrBiochem.2010Nov5.+/++/--/-OPG基因剔除小鼠发生骨质疏松00.20.40.60.811.21.4野生型杂合子纯合子密度（g/cm3)wt-/-ProgressinBiochemistryandBiophysics2007,34(3):260-266.AdiponectindeficiencyimpairsliverregenerationthroughattenuatingSTAT3phosphorylationinmiceLabInvest.2009Sep;89(9):1043-52.Epub2009Jun29.DelayedliverinjuryandimpairedhepatocyteproliferationaftercarbontetrachlorideexposureinBPOZ2-deficientmice.ToxicolLett.2009Aug10;188(3):201-7.Epub2009Apr23.Kif18Amutationcauses―Sertoli-cell-onlysyndrome‖wt+/--/-00.511.522.5wt+/--/-SerumFSH(ng/ml)0123456781W2W4W8W12WTestis/bodyweight(%)wt+/--/-0123456781W2W4W8W12WTestis/bodyweight(%)wt+/--/-00.511.52wt+/--/-Testosterone(ng/ml)p=0.005p=0.01ABECD-/-Te-/-EpwtTewtEpKif18aisspecificallyexpressedinadultmousetestes,restrictedtospermatogeniccellsduringmitosisandmeiosisMergeKif18a/a-tubulinKif18awtKif18a-/-wtKif18a-/-ABCDE17d18d19d20d21d22dKif18a-LKif18a28SSa.GSpThTestisKif18a1CoomassieBluestainingwt-/-wt-/-wt-/-wt+/--/-Gpr26deficientmicedisplayincreasedanxietyanddepression-likebehaviorsaccompaniedbyreducedpCREBlevelincentralamygdalaGpr26deficientmicedisplayincreasedanxietyanddepression-likebehaviorsaccompaniedbyreducedpCREBlevelincentralamygdala00.010.020.030.040.050.06BLACeAMeApCREBMeanDensityWTKO00.020.040.060.080.1BLACeAMeACREBMeanDensityWTKO0123456WBCoHiCepCREB/CREBWTKO00.20.40.60.811.21.4WBCoHiCeCREB/LaminBWTKOWbCoHiCeWbCoHiCepCREBCREBLaminBWTKOWTKOWTKOCREBpCREBMeACeABlAABCDEF*MouseGpr