NEURAL CONTROL OF A MOTOR PROGRAM

University Of Washington, Seattle WA

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Abstract

The goal of the proposed research is to investigate neuron and muscle excitability in the nematode C. elegans. We have identified over 30 genes that regulate excitation of defecation, egg-laying, and body-wall muscles, and we have molecularly cloned five of these genes. The genetics of four of these is strikingly similar: each has dominant gain-of-function mutations (gf) that cause strong defects in muscle excitation, and loss-of-function (lf) mutations cause little or no obvious phenotype. All four genes encode K+ channels. We think that the gf mutations in each K+ channel cause channel activation in vivo, accounting for their strong excitation defects. The fact that lf mutations cause relatively minor defects suggests that the many K+ channels have overlapping functions in vivo. We will continue analysis of these genes and other similar genes identified in genetic screens. The K+ channels will be expressed in cultured cells to study their electrophysiological properties. It will be particularly interesting to study how the gf mutations affect channel properties. Two of these K+ channels are related to the human HERG channel, defects in which cause a cardiac malfunction called long-QT. Long-QT can also be caused by tricyclic antidepressants and certain cardiac anti-arrhythmic drugs. We have evidence that these drugs also block one (but not the other) of the C. elegans HERG-related channels in vivo and in vitro. We will study the C. elegans channels and their human equivalents to understand the basis for this specificity and its implications for long-QT disorder. We have shown that the fifth muscle excitation gene, called unc- 43, encodes the nematode homologue of calcium-calmodulin dependent protein kinase II (CaMKII). CaMKII is implicated as a key regulator of synaptic activity, particularly of synaptic plasticity that underlies learning and memory. We have many lf mutations in unc-43, including null mutations. These mutants are viable and have complex behavioral abnormalities. There is also one gf mutation in unc-43, and we think that this mutant CaMKII is partially Ca++ independent (activated). This gf mutant is also viable and confers complex defects that are the opposite of those in null mutants. We have begun to use the unc-43 activated mutation to identify extragenic suppressors of its various phenotypes, some of which we expect to encode direct CaMKII substrates. We propose to use a combination of genetic analysis, molecular cloning, and biochemical analysis to characterize these potential targets and to determine whether they are directly phosphorylated by the unc-43 CaMKII. We will also continue genetic screens to identify additional potential targets.