Background-Removal of the Bax gene from mice completely protects the somas of retinal ganglion cells (RGCs) from apoptosis following optic nerve injury. This makes BAX a promising therapeutic target to prevent neurodegeneration. In this study, Bax +/− mice were used to test the hypothesis that lowering the quantity of BAX in RGCs would delay apoptosis following optic nerve injury.Methods-RGCs were damaged by performing optic nerve crush (ONC) and then immunostaining for phospho-cJUN and quantitative PCR were used to monitor the status of the BAX activation mechanism in the months following injury. The apoptotic susceptibility of injured cells was directly tested by virally introducing GFP-BAX into Bax −/− RGCs after injury. The competency of quiescent RGCs to reactivate their BAX activation mechanism was tested by intravitreal injection of the JNK pathway agonist, anisomycin.Results-24 weeks after ONC, Bax +/− mice had significantly less cell loss in their RGC layer than Bax +/+ mice 3 weeks after ONC. Bax +/− and Bax +/+ RGCs exhibited similar patterns of Terms of use and reuse: academic research for non-commercial purposes, see here for full terms. http://www.springer.com/gb/openaccess/authors-rights/aam-terms-v1 *

BAX, a member of the BCL2 gene family, controls the committed step of the intrinsic apoptotic program. Mitochondrial fragmentation is a commonly observed feature of apoptosis, which occurs through the process of mitochondrial fission. BAX has consistently been associated with mitochondrial fission, yet how BAX participates in the process of mitochondrial fragmentation during apoptosis remains to be tested. Time-lapse imaging of BAX recruitment and mitochondrial fragmentation demonstrates that rapid mitochondrial fragmentation during apoptosis occurs after the complete recruitment of BAX to the mitochondrial outer membrane (MOM). The requirement of a fully functioning BAX protein for the fission process was demonstrated further in BAX/BAK-deficient HCT116 cells expressing a P168A mutant of BAX. The mutant performed fusion to restore the mitochondrial network. but was not demonstrably recruited to the MOM after apoptosis induction. Under these conditions, mitochondrial fragmentation was blocked. Additionally, we show that loss of the fission protein, dynamin-like protein 1 (DRP1), does not temporally affect the initiation time or rate of BAX recruitment, but does reduce the final level of BAX recruited to the MOM during the late phase of BAX recruitment. These correlative observations suggest a model where late-stage BAX oligomers play a functional part of the mitochondrial fragmentation machinery in apoptotic cells.

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