Loss-of-function EA2 mutations are associated with impaired neuromuscular transmission

Objective: To examine the functional consequences of episodic ataxia type 2 (EA2)-causing nonsense and missense mutations in vitro and to characterize the basis of fluctuating weakness in patients with E2A. Background: Mutati ons in CACNA1A encoding the Ca(v)2.1 calcium channel subunit cause EA2 thro ugh incompletely understood mechanisms. Although the Ca(v)2.1 subunit is im portant for neurotransmission at the neuromuscular junction, weakness has n ot been considered a feature of EA2. Methods: The disease-causing mutations in three unrelated patients with EA2 and fluctuating weakness were identif ied by mutation screening and sequencing. Mutant constructs harboring mutat ions R1281X, F1406C, R1549X were transfected into COS7 cells and expressed for patch clamp studies. Single-fiber electromyography (SFEMG) was performe d in patients to examine synaptic transmission at the neuromuscular junctio n. Results: Functional studies in COS7 cells of nonsense and missense EA2 m utants demonstrated markedly decreased current densities compared with wild type. SFEMG demonstrated jitter and blocking in these patients with EA2, c ompared with normal subjects and three patients with SCA-6. Conclusion: EA2 -causing missense and nonsense mutations in CACNA1A produced mutant channel s with diminished whole cell calcium channel activity in vitro due to loss of function. Altered biophysical properties or reduced efficiency of plasma membrane targeting of mutant channels may contribute to abnormal neuromusc ular transmission, manifesting as myasthenic syndrome.