Abstract
We previously isolated a cDNA encoding a novel large GTP-binding protein (mOPA1) from the mouse brain. In transfected cultured cells, mOPA1 is localized to mitochondria and the overexpression of mOPA1 induced dramatic morphological changes of mitochondria from a tubular to a fragmented shape.
We have identified that the C-terminal region of mOPA1 intensely interacted with each other using a yeast two-hybrid method. In this study, we tried to confirm the intracellular homomeric interaction of mOPA1. FLAG- and Myc-tag sequence were added to the C-terminus of wild type mOPA1 (960 a.a. residues) or C-terminal deletion mutant of mOPA1 (525 a.a. residues). COS-7 cells were cotransfected with both tagged mOPA1, and the immunoprecipitation was carried out using anti-FLAG antibody from cell extracts. We confirmed that Myc-tagged mOPA1 was coprecipitated with FLAG-tagged mOPA1, indicating that wild type mOPA1 was homomultimerized in transfected cells. Unexpectedly, C-terminal deletion mutant of mOPA1 was also coprecipitated although the intense interaction domain was removed.
In transfected COS-7 cells, C-terminal deletion mutant of mOPA1 showed a vesicular distribution in mitochondria, but mitochondrial fragmentation was not observed. These results suggest that homomultimerization of mOPA1 is not sufficient and the C-terminal sequence of mOPA1 is necessary for mitochondrial fragmentation. [Jpn J Physiol 54 Suppl:S82 (2004)]
