PERSPECTIVE NATURE฀GENETICS฀|฀VOLUME฀40฀|฀NUMBER฀3฀|฀MARCH฀2008฀ 275 What฀causes฀mitochondrial฀DNA฀deletions฀in฀฀ human฀cells? Kim฀J฀Krishnan,฀Amy฀K฀Reeve,฀David฀C฀Samuels,฀Patrick฀F฀Chinnery,฀John฀K฀Blackwood,฀Robert฀W฀Taylor,฀฀ Sjoerd฀Wanrooij,฀Johannes฀N฀Spelbrink,฀Robert฀N฀Lightowlers฀&฀Doug฀M฀Turnbull Mitochondrial฀DNA฀(mtDNA)฀deletions฀are฀a฀primary฀cause฀of฀ mitochondrial฀disease฀and฀are฀likely฀to฀have฀a฀central฀role฀in฀ the฀aging฀of฀postmitotic฀tissues.฀Understanding฀the฀mechanism฀ of฀the฀formation฀and฀subsequent฀clonal฀expansion฀of฀these฀ mtDNA฀deletions฀is฀an฀essential฀first฀step฀in฀trying฀to฀prevent฀ their฀occurrence.฀We฀review฀the฀previous฀literature฀and฀recent฀ results฀from฀our฀own฀laboratories,฀and฀conclude฀that฀mtDNA฀ deletions฀are฀most฀likely฀to฀occur฀during฀repair฀of฀damaged฀ mtDNA฀rather฀than฀during฀replication.฀This฀conclusion฀has฀ important฀implications฀for฀prevention฀of฀mtDNA฀disease฀and,฀ potentially,฀for฀our฀understanding฀of฀the฀aging฀process. Mitochondrial฀DNA฀(mtDNA)฀fascinates฀both฀biologists฀and฀clinicians฀ because฀of฀its฀unique฀genetics฀and฀its฀role฀in฀human฀disease.฀The฀most฀ common฀ genetic฀ defects฀ seen฀ in฀ individuals฀ with฀ mtDNA-associated฀ diseases฀are฀deletions 1,2 ฀or฀point฀mutations 3,4 .฀MtDNA฀deletions,฀mol- ecules฀that฀have฀lost฀large฀sections฀of฀the฀mtDNA฀genome,฀have฀been฀ shown฀to฀have฀an฀important฀role฀in฀human฀pathology฀in฀three฀different฀ clinical฀scenarios.฀First,฀single฀mtDNA฀deletions฀are฀a฀common฀cause฀of฀ sporadic฀mitochondrial฀disease;฀in฀these฀cases,฀an฀identical฀mtDNA฀dele- tion฀is฀detected฀in฀all฀cells฀within฀an฀affected฀tissue 5 .฀Second,฀in฀another฀ large฀group฀of฀individuals฀with฀mitochondrial฀disease,฀there฀are฀multiple฀ mtDNA฀deletions฀in฀affected฀tissues,฀particularly฀in฀the฀muscle฀and฀the฀ central฀nervous฀system 6 .฀The฀primary฀genetic฀defect฀in฀these฀individuals฀ involves฀nuclear฀genes฀encoding฀proteins฀involved฀in฀either฀mitochondrial฀ nucleotide฀metabolism฀(TP฀and฀ SLC25A4,฀the฀gene฀encoding฀the฀heart- muscle฀specific฀isoform฀of฀ANT1)฀or฀mtDNA฀maintenance฀(C10orf2฀ (also฀ known฀as฀ PEO1),฀ POLG฀and฀ POLG2) 7 .฀฀Third,฀there฀are฀numerous฀reports฀ of฀mtDNA฀deletions฀in฀aged฀postmitotic฀tissues฀and฀individuals฀with฀ neurodegenerative฀diseases 8,9 .฀The฀extent฀of฀mtDNA฀deletions฀in฀these฀ cases฀is฀often฀much฀lower฀than฀that฀seen฀in฀individuals฀with฀mitochon- drial฀disease,฀but฀in฀substantia฀nigra฀neurons,฀for฀example,฀approximately฀ 50%฀of฀all฀mtDNA฀molecules฀contain฀an฀mtDNA฀deletion 8,9 . The฀ ratio฀ of฀ mutated฀ versus฀ wild-type฀ mtDNA฀ is฀ a฀ crucial฀ fac- tor฀ in฀ determining฀ whether฀ there฀ will฀ be฀ a฀ biochemical฀ defect 6 .฀ As฀ mtDNA฀mutations฀are฀functionally฀recessive,฀usually฀>60%฀of฀mtDNA฀ molecules฀contain฀a฀deletion฀before฀a฀biochemical฀defect฀is฀observed฀ (detected฀by฀the฀absence฀of฀cytochrome฀c฀oxidase฀(COX)฀activity) 10 .฀In฀ individuals฀with฀somatic฀mtDNA฀mutations,฀there฀are฀different฀mtDNA฀ deletions฀in฀different฀cells,฀suggesting฀that฀clonal฀expansion฀can฀reach฀ levels฀that฀cause฀COX฀deficiency 8,9,11 . To฀understand฀the฀mechanisms฀involved฀in฀the฀formation฀and฀clonal฀ expansion฀ of฀ mtDNA฀ deletions,฀ one฀ must฀ take฀ into฀ consideration฀ the฀very฀different฀clinical฀scenarios฀in฀which฀these฀mtDNA฀deletions฀are฀ observed.฀Thus,฀any฀hypothesis฀must฀explain฀a฀range฀of฀findings,฀from฀ mtDNA-deletion฀formation฀in฀oocytes฀to฀that฀in฀neurons฀from฀elderly฀ subjects,฀for฀example.฀ Common฀features฀of฀mtDNA฀deletions Most฀mtDNA฀deletions฀share฀similar฀characteristics.฀Most฀are฀located฀ in฀ the฀ major฀ arc฀ between฀ two฀ proposed฀ origins฀ of฀ replication฀ (O H ฀ and฀O L ;฀Mitomap),฀and฀are฀predominantly฀(~85%)฀flanked฀by฀short฀ direct฀repeats 11,12 .฀We฀have฀recently฀studied฀the฀types฀of฀mtDNA฀dele- tions฀detected฀in฀neurons฀from฀aging฀human฀brain,฀individuals฀with฀ Parkinson’s฀disease฀and฀an฀individual฀with฀a฀multiple฀mtDNA฀deletion฀ disorder,฀and฀compared฀these฀with฀those฀described฀in฀individuals฀with฀ single,฀large-scale฀mtDNA฀deletions 13 .฀The฀results฀from฀this฀study฀show฀ that฀mtDNA-deletion฀characteristics฀are฀similar฀under฀all฀circumstances,฀ with฀no฀notable฀differences฀in฀size,฀presence฀or฀absence฀of฀flanking฀repeat฀ sequences,฀or฀length฀of฀repeats.฀This฀finding฀suggests฀that฀a฀similar฀mech- anism฀is฀generating฀the฀mtDNA฀deletions฀in฀all฀these฀different฀clinical฀ situations. Are฀mtDNA฀deletions฀generated฀by฀replication฀errors? Currently,฀most฀researchers฀consider฀that฀replication฀is฀the฀likely฀mecha- nism฀behind฀deletion฀formation.฀At฀present,฀there฀are฀two฀main฀models฀of฀ mtDNA฀replication:฀a฀strand-asynchronous฀(strand-displacement)฀mode฀ of฀replication 14,15 ,฀and฀a฀more฀conventional฀coupled฀leading-lagging- strand฀replication฀(Fig.฀1) 16 .฀The฀observation฀that฀most฀reported฀mtDNA฀ deletions฀occur฀in฀the฀major฀arc฀has฀led฀to฀the฀proposal฀that฀mtDNA฀ Kim฀J.฀Krishnan,฀Amy฀K.฀Reeve,฀Patrick฀F.฀Chinnery,฀John฀K.฀Blackwood,฀ Robert฀W.฀Taylor,฀Robert฀N.฀Lightowlers฀and฀Doug฀M.฀Turnbull฀are฀at฀the฀ Mitochondrial฀Research฀Group,฀The฀Medical฀School,฀Newcastle฀University,฀ Newcastle฀upon฀Tyne,฀NE2฀4HH,฀UK.฀Kim฀J.฀Krishnan฀and฀฀ Doug฀M.฀Turnbull฀are฀also฀at฀the฀Institute฀for฀Ageing฀and฀Health,฀ Newcastle฀University,฀Newcastle฀upon฀Tyne,฀NE4฀6BE,฀UK.฀David฀C.฀ Samuels฀is฀at฀the฀Virginia฀Bioinformatics฀Institute,฀Virginia฀Polytechnic฀ Institute฀and฀State฀University,฀Blacksburg,฀Virginia฀24061,฀USA.฀Sjoerd฀ Wanrooij฀and฀Johannes฀N.฀Spelbrink฀are฀at฀the฀Institute฀of฀Medical฀ Technology฀and฀Tampere฀University฀Hospital,฀Tampere฀33014,฀Finland.฀ Sjoerd฀Wanrooj฀is฀also฀at฀the฀Karolinska฀Institute,฀Department฀of฀ Metabolic฀Disease฀Novum,฀SE-141฀86,฀Huddinge,฀Sweden.฀ e-mail:฀d.m.turnbull@ncl.ac.uk Published฀online฀27฀February฀2008;฀doi:10.1038/ng.f.94 © 2008 Nature Publishing Group http://www.nature.com/naturegenetics