Abstracts - Speakers
First Annual Waggoner Center Advance - March 22, 2013
Luisa Scott, PhD
Research Associate, Aldrich Lab
Identifying Novel BK Channel Modulators
The large-conductance calcium-activated potassium (BK) channel is an evolutionarily conserved target of ethanol. Physiologically relevant concentrations of ethanol increases BK channel opening. Additionally, genetic manipulation of BK channel function affects behavioral sensitivity to ethanol in multiple animal models. These findings suggest that the BK channel is a viable therapeutic target for alcohol intoxication and addiction. To begin to test this, we have developed a screen to identify peptides that alter BK channel function. First, we screened 30 million 9 amino acid peptide sequences using a monovalent phagemid display technique. Twenty-seven unique peptides remained after panning for sequences that bind to the human BKalpha channel but not the rat SK2 channel or human glycine alpha1 receptor. Sequences with clusters of positively charged amino acids were enriched 100-300 fold. Three motifs were enriched 3000-6000 fold. Next, select peptides were rapidly screened for functional effects using the nematode, C. elegans. Several peptides with highly enriched motifs affected an ethanol- and BK-channel dependent behavior in this animal. The behavioral assay suggests that peptide pskan4 enhances the ethanol mediated increase in BK channel opening, but does not alter channel activity in the absence of ethanol. Peptide pskan1 had BK channel- but not ethanol-dependent effects. In contrast, a peptide without a highly enriched motif had non-specific effects, and a 9 amino acid peptide not selected in the panning procedure did not alter locomotion. Finally, we have begun to confirm the function of the peptides electrophysiologically. Preliminary recordings support the putative action of peptides pskan1 and pskan4 as BK channel openers. Overall, these findings show that we have developed and successfully employed a screen for identifying and characterizing novel peptides that alter BK channel-dependent behavior in the presence of physiologically relevant concentrations of ethanol. This screening technique can be applied to identify modulators of other ion channels.