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Am J Physiol Cell Physiol (February 15, 2006). doi:10.1152/ajpcell.00348.2005
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Submitted on July 13, 2005
Accepted on February 2, 2006

Tagging and Tracking Individual Networks within a Complex Mitochondrial Web Using Photoactivatable GFP

Gilad Twig1, Solomon A Graf1, Jakob D Wikstrom1, Hibo Mohamed1, Sarah E Haigh1, Alvaro G Elorza1, Motti Deutsch2, Naomi Zurgil2, Nicole Reynolds1, and Orian S Shirihai1*

1 Pharmacology and Experimental Therapeutics, Tufts University School of Medicine, Boston, MA, USA
2 The Jerome Schottenstein Center, Physics Department, Bar-Ilan University, Ramat-Gan, Israel

* To whom correspondence should be addressed. E-mail: orian.shirihai{at}tufts.edu.

Assembly of mitochondria into networks supports fuel metabolism and calcium transport, and is involved in the cellular response to apoptotic stimuli. A mitochondrial network is defined as a continuous matrix lumen whose boundaries limit molecular diffusion. Observing individual networks has proven challenging in live cells that possess dense populations of mitochondria. Investigation into the electrical and morphologic properties of mitochondrial networks has therefore not yielded consistent conclusions. In this study we employed matrix-targeted, photoactivatable GFP to tag single mitochondrial networks. This approach, coupled with real-time monitoring of mitochondrial membrane potential, permitted the examination of matrix lumen continuity and fusion and fission events over time. We found that adjacent and intertwined mitochondrial structures often represent a collection of distinct networks. We additionally found that all areas of a single network are invariably equipotential, suggesting that a heterogeneous pattern of membrane potential within a cell's mitochondria represents differences between discrete networks. Interestingly, fission events frequently occurred without any gross morphological changes and particularly without fragmentation. These events, which are invisible under standard confocal microscopy, redefine the mitochondrial network boundaries and result in electrically disconnected daughter units.




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