营养调控与基因表达

营养调控与基因表达随着分子生物学的发展及其在营养学中的应用,从分子水平上弄清养分的代谢过程和规律,准确确定动物群体及个体的营养需要,掌握养分摄入过量及缺乏的后果,预防和治疗营养代谢疾病以及解决其他营养问题将成为可能。营养与基因表达的关系及其作用机制已成为分子生物学的重要研究内容及营养学的新兴研究领域。该领域的研究在过去5～10年中取得了较大进展。营养和基因表达的一般关系表现为两个方面:一是养分的摄入量影响基因表达;二是基因表达的结果影响养分的代谢途径和代谢效率,并决定营养需要量。Animalperformance(meat,milk,egg,wool,etc)(growth,development,etc)Genotype(geneonchromosome)Environment(nutrition,ambient,management)Inter-relationshipsbetweengenotype,environmentandanimalproduction基因表达：是指编码某种蛋白质的基因从转录、mRNA的加工与成熟、RNA的翻译、蛋白质的加工,到活性(功能)蛋白质的形成的过程(Goodridge,1994)。•基因表达调控包括转录调控、RNA加工调控、RNA转运调控、翻译调控、mRNA稳定性调控及翻译后的调控。每一个调控点都与养分直接或间接有关(Clarke,1992)。••研究表明,营养对基因表达的作用主要发生在转录或翻译前水平上,对翻译后的影响较小(Berdanier,1998)。•研究表明,真核细胞的mRNA的5’和3’端的碱基不能翻译成蛋白质,称为非编码区(UTR)。UTR含有控制mRNA的腺苷聚合、稳定性、在细胞中的分布定位以及翻译的调节信号,对基因表达的调节起着核心作用(Kozak,1992;Choi,1995)。•养分对基因表达的调控作用是通过UTR,特别是3’UTR实现的。NutrientintakeGeneexpressionDNARNAproteinNutrientuptakepathwayandmetabolismNutrientrequirementInter-relationshipsofnutritionandgeneexpressionWhichgenes,particularlythoseinvolvedincontrolofmetabolism,growthandpartitionareregulatedbynutrition?Howdonutrientsanddietregulatetheexpressionofspecificgenes?HowistheexpressionofspecificgeneproductsinvolvedinmetabolismandchannellingofnutrientsFundamentalquestionsDietaryconstituentDirectregulationPhysiologicalmodulationSecondarymediatorRegulatedtranscriptionmRNAProteinResponsivegenesRegulatedtranscriptionDirectandindirecteffectsofnutritiononregulationofgeneexpressionDNARNA3’polyadenylationtranscription5’CapAAA200SplicingAAA200MaturemRNANucleusCytoplasmPlasmamembraneExons1234PromoterSomemicronutrientshormonesSignaltransductionpathwaysIntracellularreceptorsAAAmRNATranslocationPolyribosomesProteinTranslationCriticalcontrolpoints1)TranscriptionalregulationActivation,transcriptionalspeed2)Post-transcriptionalregulationmRNAprocessing(splicing)mRNAtransportation,localizationmRNAstabilitymRNAtranslation3)Post-translationregulationPost-transcriptionalcontrolofgeneexpression(3)(1)(2)Regulatorysignalsintheuntranslatedregionsandtheirinteractionswithnutrition1.Interactionbetweennutritionandgeneexpression2.Nutritionalregulationofgeneexpression3.Molecularbiologicalapproachestonutrient-geneinteractions4.Effectsofenvironmentalfactorsonnutrient-geneinteractionsPrinciples:isolation&estimationofmRNA1)mRNAistheprimaryproductofageneandmediatesitsexpressionasprotein.Inmanyinstances,nutrienteffectsontheexpressionofproteinreflectchangesintheconcentrationoftheirmRNA.ItisalsofrequentlyeasiertodeterminemRNAconcentrationsthantoestimatethecorrespondingproteins.2)mRNAcanreadilybeconvertedintotheirequivalentDNAbyfirstsynthesizingacomplementarystrandofDNA(cDNA)andthenasecondstrandcomplementarytothefirst.DNAismucheasiertomanipulatethanRNAinplasmidsorbyPCR.ThecDNAcanbesequencedtoproteinsequenceinformationwhichcanusedinstructuralstudiesandindevelopmentofimmunologicalmethodsofproteinestimation.Principles:isolation&estimationofmRNA3)ThesynthesisofindividualmRNAisrarelybydirectinteractionofanutrientwithagene.Instead,controlisgenerallymediatedthroughoneormoreproteinsreferredtoastranscriptionfactors.Thesebindtoregulatoryregionsofthegeneanddeterminetheefficiencyofitstranscription.Principles:isolation&estimationofmRNAMaintechniques1)Hybridization2)IsolationofmRNA3)Probes4)EstimationofmRNANorthernblottingRibonucleaseprotectionaaasy5)PCR,realtime-PCR6)DNAsequencing7)CloneImportantunderstandinghowanutrientinfluencesexpressionofaparticulargenerequiresastudyofthefactorswhichbindtothatgene’sregulatoryregion1.Interactionbetweennutritionandgeneexpression2.Nutritionalregulationofgeneexpression3.Molecularbiologicalapproachestonutrient-geneinteractions4.Effectsofenvironmentalfactorsonnutrient-geneinteractionsAnimalperformance(meat,milk,egg,wool,etc)(growth,development,etc)Genotype(geneonchromosome)Environment(nutrition)Inter-relationshipsbetweengenotype,environmentandanimalproductionambientmanagementI.NutritionandgeneexpressionFeedindustryAnimalproductionEffluentLessslurryLessodourAnimalbiotechnologyConsumerproducts-MilkMeatEggFishMicrobialbiotechnologySilageinoculantsProbioticsEnzymesAminoacidsGrowthenhancersEnvironmentalbiotechnologyChemicalorbiotechnologypre-treatmentConservationChemicalorbiotechnologypre-treatmentBetterproteinoraminoacidsCurrentcropsPlantbiotechnologyBy-productsConsumerproducts-FlourWhiskyBeerFood/drinkindustryChemicalindustryChemicalsStarchGlutenAdhensivesOilsBy-products一、能量蛋白质对基因表达的影响生长激素(GH)是控制动物出生后生长的主要激素。GH对生长的控制必须通过GH受体(GHR)及类胰岛素生长因子-Ⅰ(IGF-Ⅰ)的作用才能实现,IGF-Ⅰ是GH促进生长的最重要的介导物(Cohick,1993)。哺乳动物体内存在另一种IGF,叫IGF-Ⅱ,主要在胚胎和胎儿组织产生,是胚胎和胎儿的主要生长因子。IGF-Ⅰ和IGF-Ⅱ的大多数作用均需要IGF-Ⅰ受体(Ⅰ型受体)参与(Straus,1994)。除GH外,营养状况是调控IGF-Ⅰ的重要因素。大多数动物试验均表明,营养不良导致的生长受阻往往伴随有血浆GH水平的升高,而不是下降(Buonomo,1991;;Vance,1992)。能量蛋白质对生长调节基因表达的影响可能具有组织特异性。二、氨基酸对基因表达的影响氨基酸除参与IGF-Ⅰ和GHR基因表达的调节外,还与多种其他基因表达的调节有关。Marten(1994)报道,在大鼠肝细胞培养基质中去掉氨基酸后促进了数个基因的表达,提高幅度最大的是CHOP基因。CHOP是一分子量很小的核蛋白,为转录因子C/EBP(CCAAT/促进子结合蛋白)的同源蛋白。CHOP通过与C/EBP结合成二聚体而参与多种基因表达的调节(Cao,1991;Birkenmeier,1989;Umek,1991)。Bruhat(1997,1999)用人体细胞培养证明,低浓度的亮氨酸明显提高CHOP基因的表达,其机制是提高了基因的转录率和CHOPmRN