Alternate Exons in Caenorhabditis Elegans Slo-1 Diversify BK Channel Calcium Dependence PDF Download

Author: Brandon Eugene Johnson
Publisher: Stanford University
ISBN:
Category :
Languages : en
Pages : 172

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Book Description
Ion channels coordinate the movement of ions across biological membranes in response to a diversity of cellular signals to regulate most if not all aspects of physiology. The large-conductance Ca2+- and voltage-activated potassium (BK) channel is widely expressed across tissues and functions to repolarize action potentials, regulate smooth muscle tone and endocrine function, and tune auditory hair cells. In all tissues, BK channels provide a functional link between cytosolic Ca2+ and membrane excitation. Extensive alternate splicing of the slo-1 gene, encoding the BK channel, significantly diversifies channel function. We identified all twelve predicted BK channel splice variants expressed by the nematode Caenorhabditis elegans, and measured the Ca2+ and voltage dependence for each. Alternate exons encode portions of two regions of the cytoplasmic tail: the RCK domain, important for Ca2+ coordination, and an unstructured region, termed the C-linker. Using the patch clamp technique, we determined that alternate splicing across multiple sites diversifies activation kinetics and Ca2+ sensitivity by manipulating the functional interaction between the RCK domain and C-linker. We recovered two slo-1 alleles (pg34 and pg52) that contain point-mutations near an alternative splice site in the C-linker from a mutant screen. Using real-time PCR, we determined that both mutations change slo-1 splice variant expression profiles. The pg34 mutation encodes an A698T substitution, and the pg52 mutation is located in an intron. When heterologously expressed, the A698T mutation changes BK channel Ca2+ sensitivity in specific splice variant backgrounds by disrupting the functional connection between the C-linker and the RCK domain. Using in vivo electrophysiology, we determined that the pg34 mutation reduces but does not abolish BK channel function in C. elegans neurons. These results suggest that alternate splicing of slo-1 modulates the functional connection between the RCK domain and the C-linker of the BK channel to establish a proper balance between intracellular Ca2+ and membrane excitation in a variety of cell type and physiological conditions.