TRANSFEKSI DNA KE DALAM MITOKONDRIA SEL BETA PANKREAS TIKUS DAN SEL PRIMER MYOBLAST MANUSIA: LANGKAH MENUJU REKAYASA DNA MITOKONDRIA DAN TERAPI GEN
DOI:
https://doi.org/10.21776/ub.jkb.2004.020.03.1Abstract
The delivery of an exogenous or modified DNA into the mitochondria of mammalian cells has been proposed as an alternative way to prevent or modify mitochondrial genetic defect. In  this study, a lipofection method was developed to transfer exogenous DNA into mitochondria of BRIN BD11 cells, a rat beta pancreas cell line, and human myoblast primary cells. An artificial mini-mitochondrial genome carrying a reporter gene encodes for a mitochondrial green fluorescent protein (mtGFP) was usedto track the fate of the exogenous DNA following its transfection into cellular mitochondria. Epifluorescence microscopic observation was carried out to detect the green fluorescence resulted from the GFPexpression. Immunohistochemical assays were also performed to validate and confirm the GFP expression. Collectively, the results indicate that the exogenous DNA pmtGFP could be  delivered into the mitochondrial compartment and was functional withinthe mitochondria of BRIN BD11 and human myoblast cells. Such achievement is a significant progress as both the artificial DNA construct and the mitochondrial transfection method could serve as a model to study the dynamic of mutant mtDNA as well as to provide a basis for the development of gene therapy for human mitochondrial diseases. Key words: DQAsome, pmtGFP, beta pancreas cell line, human myoblast primary cellsDownloads
References
DAFTAR KEPUSTAKAAN
Pinkert CA and Trounce IA. Production of transmitochondrial mice. Methods, 2002; 26: 348-357.
Lyrawati D, Trounson A and Cram D. (submitted) Expression of GFP In Mitochondrial Compartment Using DQAsome-mediated Delivery Of An Artificial Mini-mitochondrial Genome. Majalah Kedokteran Fakultas Kedokteran Universitas Brawijaya.
Yamaoka M, Mikami T, Ono T, Nakada K and HayashiJI. Mice With Only Rat mtDNA Are Required As Models Of Mitochondrial
Diseases. Biochem Biophys Res Commun, 2001; 282:707-711.
Yamaoka M, Isobe K, Shitara H, Yonekawa H, Miyabayashi S and Hayashi JI. Complete repopulation Of Mouse Mitochondrial DNA-less Cells With Rat Mitochondrial DNA restores Mitochondrial Translation But Not Mitochondrial Respiratory Function. Genetics,2000;155:301-307.
Dey R, Barrientos A and Moraes CT. Functional Constraints Of Nuclear-Mitochondrial DNAInteractions In Xenomitochondrial Rodent Cell Lines. J Biol Chem, 2000;275:31520-31527.
McKenzie M and Trounce I. Expression Of Rattus Norvegicus mtDNA In Mus Musculus Cells Results In Multiple Respiratory Chain Defects. J. Biol. Chem., 2000; 275: 31514-31519.
Rinaudo P, Niven-Fairchild T, Buradagunta S, Massobrio M, Revelli A and Keefe DL. Microinjection Of Mitochondria Into Zygotes Creates A Model For Studying The Inheritance Of Mitochondrial DNA During Preimplantation Development.Fertility and Sterility,1999; 71:912-918.
Kenyon L and Moraes CT. Expanding The Functional Human Mitochondrial Dna Database By The Establishment Of Primate
Xenomitochondrial Cybrids. Proc Natl Acad Sci USA, 1997; 94:9131-9135.
Barrientos A, Muller S, Dey R, Wienberg J and Moraes CT. Cytochrome C Oxidase Assembly In Primates Is Sensitive To Small Evolutionary Variations In Amino Acid Sequence. Mol Biol Evol,2000;17:1508-1519.
Tang Y, Schon EA, Wilichowski E, Vazquez-MemijeME, Davidson E and King MP. Rearrangements Of Human Mitochondrial DNA (mtDNA): New Insights Into The Regulation Of mtDNA copy Number And Gene Expression. Mol Biol Cell, 2000; 11:1471-1485.
Azpiroz R and Butow RA. Mitochondrial Inheritance In Yeast. Methods Enzymol, 1995; 260:453-465.
Nunnari J, Marshall WF, Straight A, Murray A, Sedat JW and Walter P. Mitochondrial Transmission During Mating In
Saccharomyces Cerevisiae Is Determined By Mitochondrial Fusion And Fission And The Intramitochondrial Segregation Of Mitochondrial DNA.Mol Biol Cell, 1997; 8:1233-1242.
Okamoto K, Perlman PS and Butow RA. The Sorting Of Mitochondrial DNA And Mitochondrial Proteins In Zygotes: Preferential Transmission Of Mitochondrial Dna To The Medial Bud. J Cell Biol, 1998; 142:613-623.
Wallace DC. Mitochondrial Diseases In Man And Mouse.Science, 1999; 283:1482-1488.
Downloads
Published
Issue
Section
License
Authors who publish with this journal agree to the following terms:- Authors retain copyright and grant the journal right of first publication with the work simultaneously licensed under a Creative Commons Attribution License that allows others to share the work with an acknowledgement of the work's authorship and initial publication in this journal.
- Authors are able to enter into separate, additional contractual arrangements for the non-exclusive distribution of the journal's published version of the work (e.g., post it to an institutional repository or publish it in a book), with an acknowledgement of its initial publication in this journal.
- Authors are permitted and encouraged to post their work online (e.g., in institutional repositories or on their website) prior to and during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work (See The Effect of Open Access).
