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Bok regulates mitochondrial fusion and morphology

Abstract

Bok (Bcl-2-related ovarian killer) is a member of the Bcl-2 protein family that governs the intrinsic apoptosis pathway, but the cellular role that Bok plays is controversial. Remarkably, endogenous Bok is constitutively bound to inositol 1,4,5-trisphosphate receptors (IP3Rs) and is stabilized by this interaction. Here we report that despite the strong association with IP3Rs, deletion of Bok expression by CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein-9 nuclease)-mediated gene editing does not alter calcium mobilization via IP3Rs or calcium influx into the mitochondria. Rather, Bok deletion significantly reduces mitochondrial fusion rate, resulting in mitochondrial fragmentation. This phenotype is reversed by exogenous wild-type Bok and by an IP3R binding-deficient Bok mutant, and may result from a decrease in mitochondrial motility. Bok deletion also enhances mitochondrial spare respiratory capacity and membrane potential. Finally, Bok does not play a major role in apoptotic signaling, since Bok deletion does not alter responsiveness to various apoptotic stimuli. Overall, despite binding to IP3Rs, Bok does not alter IP3R-mediated Ca2+ signaling, but is required to maintain normal mitochondrial fusion, morphology, and bioenergetics.

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Acknowledgements

We thank Dr. Nicholas Deakin for support and suggestions regarding microscopy; Dr. Danielle Sliter and Dr. Richard Youle for facilitating mitochondrial fusion rate measurements; Dr. Forrest Wright and Caden Bonzerato for helpful suggestions; Dr. Ebbing De Jong for assistance with mass spectral analysis; Dr. Thomas Kaufmann for providing anti-Bok; Dr. Jan B Parys for providing anti-IP3R1–3; and grant support to RJHW from NIH (DK107944 and GM121621) and the Carol M. Baldwin Breast Cancer Research Fund.

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Correspondence to Richard J. H. Wojcikiewicz.

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