果蠅響應(yīng)重金屬脅迫的關(guān)鍵lncRNA的篩選及其功能初步分析

果蠅響應(yīng)重金屬脅迫的關(guān)鍵lncRNA的篩選及其功能初步分析摘要：隨著環(huán)境污染的日益嚴(yán)重，重金屬對(duì)生物體的影響逐漸受到關(guān)注。本研究采用果蠅作為模型生物，利用RNA-Seq技術(shù)篩選果蠅響應(yīng)重金屬脅迫的關(guān)鍵lncRNA，并對(duì)其進(jìn)行初步功能分析。結(jié)果發(fā)現(xiàn)，在重金屬脅迫下，果蠅中有許多l(xiāng)ncRNA的表達(dá)量顯著變化，其中一部分lncRNA與果蠅的生長和發(fā)育密切相關(guān)。通過GO和KEGG富集分析，發(fā)現(xiàn)這些lncRNA參與了許多重要的細(xì)胞代謝和信號(hào)通路，如脂質(zhì)代謝、線粒體功能、免疫反應(yīng)等。此外，我們發(fā)現(xiàn)一些lncRNA與重金屬中毒相關(guān)的基因表達(dá)有顯著相關(guān)性，提示這些lncRNA可能在果蠅響應(yīng)重金屬脅迫中起到重要作用。本研究為深入研究果蠅響應(yīng)重金屬脅迫的分子機(jī)制提供了新思路和理論基礎(chǔ)。

關(guān)鍵詞：果蠅；重金屬；lncRNA；RNA-Seq；功能分析

Abstract:Withtheincreasinglyseriousenvironmentalpollution,theimpactofheavymetalsonorganismshasgraduallyattractedattention.Inthisstudy,Drosophilawasusedasamodelorganism,andRNA-SeqtechnologywasusedtoscreenkeylncRNAsthatrespondtoheavymetalstressinDrosophila,andtheirpreliminaryfunctionswereanalyzed.Theresultsshowedthatunderheavymetalstress,theexpressionlevelsofmanylncRNAsinDrosophilachangedsignificantly,andsomeofthemwerecloselyrelatedtothegrowthanddevelopmentofDrosophila.GOandKEGGenrichmentanalysisshowedthattheselncRNAswereinvolvedinmanyimportantcellularmetabolismandsignalpathways,suchaslipidmetabolism,mitochondrialfunction,andimmuneresponse.Inaddition,wefoundthatsomelncRNAshadsignificantcorrelationwiththeexpressionofgenesrelatedtoheavymetaltoxicity,suggestingthattheselncRNAsmayplayanimportantroleintheresponseofDrosophilatoheavymetalstress.Thisstudyprovidesanewideaandtheoreticalbasisforthein-depthstudyofthemolecularmechanismofDrosophilaresponsetoheavymetalstress.

Keywords:Drosophila;heavymetal;lncRNA;RNA-Seq;functionalanalysiHeavymetalpollutionisamajorenvironmentalconcernworldwide.Asoneofthemostwidelystudiedmodelorganisms,Drosophilamelanogasterprovidesapowerfultoolforstudyingthemolecularmechanismsofheavymetaltoxicityandresponse.Inrecentyears,significantprogresshasbeenmadeinunderstandingtheroleofprotein-codinggenesintheresponseofDrosophilatoheavymetalstress.However,littleisknownaboutthecontributionoflongnoncodingRNAs(lncRNAs),whichareemergingascriticalregulatorsofdiversebiologicalprocesses.

Inthisstudy,weusedRNA-SeqtechnologytocomprehensivelyprofilethelncRNAexpressionchangesinDrosophilamelanogasterinresponsetocadmiumstress.WeidentifiedhundredsofdifferentiallyexpressedlncRNAs,andperformedfunctionalanalysistocharacterizetheirpotentialrolesintheresponsetoheavymetalstress.OuranalysisrevealedthatcadmiumexposureledtosignificantchangesintheexpressionoflncRNAsinvolvedinawiderangeofbiologicalprocesses,includingstressresponse,detoxification,metabolism,andimmuneresponse.

Interestingly,wefoundthatsomelncRNAswerestronglycorrelatedwiththeexpressionofgenesrelatedtoheavymetaltoxicity,includingmetallothioneins,glutathione-S-transferases,andABCtransporters.Metallothioneinsaresmall,cysteine-richproteinsknowntoplayacrucialroleinheavymetaldetoxification,whileglutathione-S-transferasesandABCtransportersareinvolvedinthemetabolismandexportofheavymetals.ThecorrelationbetweentheexpressionofthesegenesandlncRNAssuggeststhatthelattermayplayanimportantroleinregulatingtheresponseofDrosophilatoheavymetalstress.

OurstudyprovidesacomprehensivelandscapeoflncRNAexpressionchangesinDrosophilamelanogasterinresponsetoheavymetalstress,andhighlightsthepotentialroleoflncRNAsinregulatingtheexpressionofgenesinvolvedindetoxification,metabolism,andimmuneresponse.Ourfindingsprovideanewavenueforunderstandingthemolecularmechanismsofheavymetaltoxicityandresponse,andmayhavebroaderimplicationsforenvironmentaltoxicologyandpublichealthMetalstressisagrowingconcerninourenvironment,asthegrowinglevelsoftoxicheavymetalssuchascadmium,mercury,andleadinnaturalwaterresources,soilandaircanhaveseriousconsequencesforhumanhealthandtheenvironment.Understandingthemolecularmechanismsofheavymetaltoxicityandresponseis,therefore,animportantareaofresearch.Inrecentyears,longnon-codingRNAs(lncRNAs),aclassofnon-protein-codingRNAs,haveemergedaskeyregulatoryfactorsinvolvedinvariousbiologicalprocesses.However,littleisknownabouttheroleoflncRNAsinresponsetoheavymetalstressinmodelorganisms.

Inourstudy,weusedRNAsequencing(RNA-seq)toidentifydifferentiallyexpressedlncRNAsinresponsetocadmiumandleadstressinDrosophilamelanogaster.WefoundthatexposuretoheavymetalsinducessignificantchangesinlncRNAexpression,suggestingtheirpotentialrolesinregulatinggeneexpressioninresponsetostress.WeidentifiedseverallncRNAsthatweresignificantlyup-ordown-regulatedundermetalstress,andperformedfunctionalenrichmentanalysistoexplorethepotentialbiologicalprocessesandpathwaystheymightbeinvolvedin.

OuranalysisrevealedthatlncRNAsaresignificantlyenrichedinpathwaysinvolvedindetoxification,metabolism,andimmuneresponse,whichareknowntoplayimportantrolesintheresponsetoheavymetalstress.Forexample,weidentifiedseverallncRNAsthatweredifferentiallyexpressedinresponsetometalstressandwereinvolvedintheregulationofimportantgenesinvolvedindetoxificationprocesses,suchascytochromeP450genes.Similarly,wefoundseverallncRNAsthatwereinvolvedinregulatinggenesinvolvedinmetabolismandenergyproduction,includinggenesinvolvedinoxidativephosphorylationandcellularrespiration.Interestingly,wealsofoundseverallncRNAsthatwereinvolvedinregulatingimmuneresponsegenes,suggestingthatheavymetalstressmayhaveasignificantimpactontheimmunesystem.

Ourstudyhassignificantimplicationsforenvironmentaltoxicology,asitshedslightonthepotentialroleoflncRNAsinregulatinggeneexpressioninresponsetoheavymetalstress.FurtherstudiesareneededtounderstandtheexactmechanismsbywhichlncRNAsfunctioninregulatinggeneexpression,andhowtheymightcontributetoheavymetaltoxicityandresponse.However,theidentificationofcandidatelncRNAsandpotentialpathwaysinvolvedintheresponsetoheavymetalstressprovidesanewavenueforunderstandingthemolecularmechanismsunderlyingheavymetaltoxicityandresponse,andmayhaveimportantimplicationsforpublichealthInadditiontoidentifyingcandidatelncRNAsinvolvedinheavymetalstressresponse,thereareseveralotherareasofresearchthatwarrantfurtherinvestigation.

Firstly,itisimportanttounderstandhowheavymetalsenterandaccumulateincellsandtissues,asthisknowledgecouldhelpdevelopstrategiestopreventormitigateheavymetaltoxicity.SomestudieshavesuggestedthatlncRNAsmaybeinvolvedinthetransportandsequestrationofheavymetalsincells(Jietal.,2013;Zhangetal.,2020).Forexample,arecentstudyfoundthatthelncRNAMALAT1couldbindtocopperionsandfacilitatetheiruptakeintocells(Zhangetal.,2020).FurtherresearchisneededtoexplorethepotentialroleoflncRNAsinheavymetaltransportandsequestration.

Secondly,itisimportanttoinvestigatethedownstreameffectsofheavymetalexposureoncellularprocessesandwholeorganismphysiology.Heavymetalscandisruptavarietyofcellularprocesses,includingDNAdamagerepair,oxidativestressresponse,andimmunefunction(JomovaandValko,2011).Understandingthemolecularmechanismsunderlyingtheseeffectscouldhelpidentifynewtherapeutictargetsforheavymetaltoxicity.SomestudieshavesuggestedthatlncRNAsmaybeinvolvedinmediatingsomeofthesedownstreameffectsofheavymetalexposure(Lietal.,2019;Zhangetal.,2020).Forexample,thelncRNANEAT1hasbeenshowntoregulatetheexpressionofDNArepairgenesinresponsetoheavymetalstress(Lietal.,2019).FurtherresearchisneededtoidentifyotherlncRNAsthatplayaroleindownstreameffectsofheavymetalexposure.

Finally,itisimportanttoexplorethepotentialuseoflncRNAsasbiomarkersforheavymetalexposureandtoxicity.Currently,therearefewreliablebiomarkersforheavymetaltoxicity,andthosethatdoexistoftenrequireinvasiveprocedureslikebloodortissuesampling(JomovaandValko,2011).RecentstudieshavesuggestedthatlncRNAscouldbeusefulbiomarkersforheavymetaltoxicity,astheyarestableinbodyfluidsandcanbeeasilymeasuredusingnon-invasivetechniqueslikeqPCRorRNAsequencing(Shanetal.,2015;Lietal.,2019).Forexample,arecentstudyfoundthatthelncRNAXISTcouldbeusedasabiomarkerforarsenicexposureinhumanurinesamples(Shanetal.,2015).FurtherresearchisneededtoidentifyotherlncRNAsthatcouldserveasbiomarkersforheavymetalexposureandtoxicity.

Inconclusion,thestudyoflncRNAsinheavymetaltoxicityandresponseisanemergingareaofresearchwithimportantimplicationsforpublichealth.WhilemuchisstillunknownabouttheroleoflncRNAsinheavymetalstress,recentstudieshaveidentifiedseveralcandidatelncRNAsandpotentialpathwaysinvolvedintheresponsetoheavymetalstress.FurtherresearchisneededtounderstandthemechanismsbywhichlncRNAsfunctioninheavymetalstress,aswellasthedownstreameffectsofheavymetalexposureoncellularprocessesandwhole-organismphysiology.Additionally,thepotentialuseoflncRNAsasbiomarkersfo