NOVEL KV GENES, KCNG--CHARACTERIZATION AND DISEASE ASSOCIATION

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Our long term goal is to determine the structural and functional properties of a new voltage-gated K+ ion (Kv) channel gene subfamily, KCNG. Voltage-gated ion channels play an integral role in determining the electrochemical properties and resulting functions of neural, neurosensory, and excitatory cells. Our preliminary results suggest that there are at least two members in the KCNG (Kv7) subfamily. Expression of the KCNG1 gene was observed in the cochlea (inner and outer hair cells), in subpopulations of vestibular and spiral ganglion cells, in specific areas of the brain (cortex, hippocampus, substantia nigra, and brain stem), and in cardiac tissue. We hypothesize that the KCNG1 gene plays a vital role in hearing through an orchestrated association with excitatory cells, as well as neural and neurosensory tissues of the ear and brain. We plan to test this hypothesis by characterizing: 1) the KCNG gene subfamily members; 2) the structure/function relationships of the Kv7 channel proteins; 3) the role of the Kv7.1 gene in the auditory system; and 4) the relationship of this gene with neural, neuromuscular, and neurosensory functions. Thus, we plan to isolate and characterize the Kv7.1 gene and its related subfamily member by initially determining their chromosomal locations through fluorescence in situ hybridization (FISH), and then identifying their genomic sequences, gene organization, and genetic regulatory elements by dideoxynucleotide sequencing of the EMBL3 lambda human lymphocyte genomic clones. In addition, size and tissue distribution of transcripts will be examined by Northern analyses. Sequence data and information about Kv7 transcript heterogeneity, inducing alternatively spliced isoforms, will be obtained by screening cDNA libraries and employing RTPCR and RACE-PCR. The hKcng1 alpha subunits and chimeric channels will be expressed in Xenopus oocytes, and the structure/function relationships will be studied by examining their pharmacological and electrophysiological properties. Experiments will be done to produce and assess mKCNG1 knock out mice. Examination of the Kv7.1 knock out phenotype will assist in determining the physiological role of KCNG1 in the auditory system. We will also examine the possible relationship between the dysfunctional human KCNG1 gene(s) and the Jervell Lange-Niesen families. Recently, our colleague Dr. Bertrand has identified a novel OHC K+ current whose pharmacological properties resemble the IA current but with a slow and very small inactivation. This current may represent functional Kcng1 channels. Collectively, these studies will determine the function and role of the hKCNG genes in neurosensory and neural tissue, as well as define the relationship of voltage-gated KCNG ion channel genes with the auditory system.