Simultaneous transfer of mitochondrial DNA and single chromosomes in somatic cells: a novel approach for the study of defects in nuclear-mitochondrial communication.
The assembly and function of respiratory-competent mitochondria in eukaryotic cells depends on collaboration between the nuclear and mitochondrial genomes, but the molecular mechanisms underlying such cross-talk are poorly understood. Microcell-mediated chromosome transfer has been used to transfer intact chromosomes from one mammalian cell to another, helping to map loci implicated in different diseases and in the senescence process. In the present work, we show that microcells have a significant number of mitochondria which can be transferred to another cell simultaneously with a limited number of chromosomes. By fusing microcells from a colon carcinoma cell line with a mitochondrial DNA (mtDNA)-less osteosarcoma cell line, we were able to isolate transmitochondrial hybrids containing only one of three selectable chromosomes and mtDNA from the donor cell. The proportion of transmitochondrial hybrids containing one chromosomal marker with respect to the total transmitochondrial hybrids and cybrids was approximately 1% and no hybrids were isolated containing more than one nuclear marker. The genetic data correlated well with the composition and structure of the microcell preparations, which showed the presence of cytoplast-like structures and microcells containing mitochondria surrounding the micronuclei. Microcell-mediated mtDNA and chromosome transfer can be used to identify nuclear factors implicated in mtDNA maintenance and gene expression, as well as to investigate nuclear factors which modulate clinical phenotypes in mitochondrial disorders.
Barrientos
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Department of Neurology and Department of Cell Biology and Anatomy, University of Miami, School of Medicine, 1501 NW 9th Avenue, Miami, FL 33136, USA.
Moraes
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D001761:Bleomycin / Q000494:pharmacology; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism; D002875:Chromosomes; D002999:Clone Cells / Q000648:ultrastructure; D003571:Cytochalasin B / Q000494:pharmacology; D004272:DNA, Mitochondrial / Q000235:genetics*; D004351:Drug Resistance / Q000235:genetics; D018014:Gene Transfer Techniques; D005819:Genetic Markers; D005821:Genetic Techniques; D006801:Humans; D006822:Hybrid Cells / Q000648:ultrastructure; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism; D009355:Neomycin / Q000494:pharmacology; D014162:Transfection
D004272:DNA, Mitochondrial; D005819:Genetic Markers; D001761:Bleomycin; C105427:Zeocin; D003571:Cytochalasin B; D009355:Neomycin