N-terminal phosphorylation of protein phosphatase 2A/Bβ2 regulates translocation to mitochondria, dynamin-related protein 1 dephosphorylation, and neuronal survival

The neuron-specific B beta 2 regulatory subunit of protein phosphatase 2A (PP2A), a product of the spinocerebellar ataxia type 12 disease gene PPP2R2B, recruits heterotrimeric PP2A to the outer mitochondrial membrane (OMM) through its N-terminal mitochondrial targeting sequence. OMM-localized PP2A/B beta 2 induces mitochondrial fragmentation, thereby increasing susceptibility to neuronal insults. Here, we report that PP2A/B beta 2 activates the mitochondrial fission enzyme dynamin-related protein 1 (Drp1) by dephosphorylating Ser656, a highly conserved inhibitory phosphorylation site targeted by the neuroprotective protein kinase AA kinase anchoring protein 1 complex. We further show that translocation of PP2A/B beta 2 to mitochondria is regulated by phosphorylation of B beta 2 at three N-terminal serines. Phosphomimetic substitution of Ser20, Ser21, and Ser22 renders B beta 2 cytosolic, blocks Drp1 dephosphorylation and mitochondrial fragmentation, and abolishes the ability of B beta 2 overexpression to induce apoptosis in cultured hippocampal neurons. Alanine substitution of Ser20Ser22 to prevent phosphorylation has the opposite effect, promoting association of B beta 2 with mitochondria, Drp1 dephosphorylation, mitochondrial fission, and neuronal death. OMM translocation of B beta 2 can be attenuated by mutation of residues in close proximity to the catalytic site, but only if Ser20Ser22 are available for phosphorylation, suggesting that PP2A/B beta 2 autodephosphorylation is necessary for OMM association, probably by uncovering the net positive charge of the mitochondrial targeting sequence. These results reveal another layer of complexity in the regulation of the mitochondrial fissionfusion equilibrium and its physiological and pathophysiological consequences in the nervous system.