Epilepsy Related Slack Channel Mutants Lead to Channel Over-Activity by Two Different Mechanisms

So far, 12 sodium-activated potassium channel (KCNT1, Slack) genetic mutants had been found in severe early-onset epilepsy patients by whole genome sequence. However, the biophysical properties change of these mutated Slack channels and the underlying mechanisms had not been fully investigated. In this abstract, we first measured the KD value of sodium sensitivity of these mutant channels by inside-out patch. We found that two mutants (R409Q and R455H) on the RCK1 domain and one mutant (Y775H) on the RCK2 domain decreased the KD value of sodium sensitivity. Furthermore, we found that the double mutant (R455H/R409Q) on RCK1 domain can decrease KD value of sodium sensitivity further, reflecting these mutants allosterically regulated sodium affinity of Slack channel. In addition, electrophysiology and molecular simulation reflected the mutant Y775H may directly facilitate sodium binding of Slack channel. Second, single channel recording data revealed that all mutants lead to over-activity of Slack channel even if they did not change the sodium sensitivity. We set up a two-step activation model to explain the gating mechanism change and concluded that these mutant channels lead to channel over-activity can be categorized as two different mechanisms.