Cryo-Electron Tomography of Mitochondria in Intact Cells Reveals Mitochondria Divide by Budding

Abstract

Eukaryotes rely on mitochondrial division so that new generation of cells can acquire adequate number of mitochondria to maintain their physiological functions. Mitochondrial division has long been thought to occur by binary fission which has recently been considered to be mediated by Drp1 and ER. However, the ultimate verification of the process of mitochondrial division has previously depended heavily on the visualization by fluorescent microscopy and conventional two-dimensional (2D) electron microscopy (EM). The resolution limit of fluorescent microscopy essentially prevents observers from seeing sufficient details of mitochondrial division which involves distance between two separating mitochondrial bodies down to zero. In conventional EM, specimen usually goes through extensive chemical and mechanical treatments including fixation, dehydration, staining and sectioning which may readily disrupt critical structure features that define mitochondrial division. On the other hand, in conventional EM, fine structure features always tend to be buried in 2D images due to overlapping of three-dimensional information. Here, I utilized the Whole Cell Cryo-Electron Tomography to probe mitochondrial division in intact cells. Small mitochondria were clearly observed budding from large mitochondria. Snapshots of large number of intermediates of mitochondrial budding were captured showing relatively larger mitochondrial buds of various sizes connected to large mitochondria by weak tethers or stalk-like structures essentially forming mitochondrial networks. High contrast densities most likely corresponding to mitochondrial DNA could be seen inside each of the mitochondrial buds. Moreover, mitochondrial budding resembles the reproductive budding of alpha-proteobacteria from which mitochondria are believed to be evolved. Therefore, this study has revealed that mitochondria divide by budding.