Activation of a Potassium Channel Mutation That Causes Spinocerebellar Ataxia Promotes Aggregation of the RhoGEF Domain-Containing Protein Plekhg4.

Kv3.3 potassium channels are highly expressed in cerebellar Purkinje neurons and contribute to the ability of these neurons to fire at high rates. In addition to their role in regulating excitability, Kv3.3 channels form a complex with several cytoplasmic proteins, including Hax-1, Arp2/3, Rac1, and TBK1. This stimulates the nucleation of actin filaments under the plasma membrane. Using biochemical and confocal laser scanning microscopy techniques, we have found that the Kv3.3 channel binds and colocalizes with Plekhg4, a guanine nucleotide exchange factor (GEF) that regulates Rac1 activity, in Purkinje neurons and in Kv3.3-expressing auditory brainstem neurons. In addition to binding Kv3.3, Plekhg4 immunoreactivity is distributed uniformly in the cytoplasm of these cells, as well as in CHO cells expressing wild-type Kv3.3. The Kv3.3-G592R mutation differs from wild-type channels in that it fails to trigger actin nucleation, constitutively activates Tank-Binding Kinase-1 (TBK1), and, in humans, leads to spinocerebellar ataxia. We find that Plekhg4 forms cytoplasmic aggregates in the cells expressing Kv3.3-G592R, and that the formation of these aggregates is further enhanced by depolarization of the plasma membrane. Pharmacological inhibition of TBK1 reduces the number of Plekhg4 aggregates in Kv3.3-G592R-expressing cells. These results suggest that Purkinje cell activity, mediated by Kv3.3 channels, may regulate Pelkhg4 aggregation and provide a potential new therapeutic approach for the treatment of spinocerebellar ataxias.