Molecular analysis of a Shaker-related K+ channel in leech

Four subfamilies of voltage-gated K"*" channels {Shaker, Shal, Shab and Shaw) have been defined that are present in many species to regulate neuronal excitability. The well characterized simple leech nervous system is ideal to study the roles K+ channels have in regulating neuronal circuits. As a starting point for studies to examine the role of K+ channels in well defined leech circuitry, we report the isolation of a Shaker-related K+ channel, called LK-1, in the two leech species Haemopis marmorota and Hirudo medicinalis. Haemopis and Hirudo LK-1 share 96% identity at the nucleic acid level and in the core region of the channel protein suggesting that the LK-1 K+ channel may play a functionally conserved role in leech neurons. LK-1 sequence encodes a protein of 454 amino acids with a predicted molecular mass of 51 kD. Phylogenetic analysis of LK-1 places it as a separate subfamily that is related to the Shaker subfamily. Tissue specific RT-PCR results detect LK-1 message in adult leech CNS and in El 5-20 stage embryos. LK-1 antisera, generated to the N terminus, detect a 54 kD protein band in leech CNS extract. The LK-1 antisera staining pattern shows that LK-1 protein is present in most neurons in the adult and embryonic leech CNS and is also found in axons and neurites in the PNS of different stage embryos. These results suggest that LK-1 encodes a novel Shaker-relaXed K+ channel which is expressed in the adult leech and developing embryo. iNeuroscience Program, Department of Zoology and Genetics, Signal Transduction Training Group, Iowa State University of Science and Technology, Ames Iowa 50011 2 Department of Anesthesiology Research, University of Texas Southwestern Medical School, Dallas Texas 75235