CLONING AND EXPRESSION OF THE LARGE-CONDUCTANCE CA2-ACTIVATED K+ CHANNEL FROM COLONIC SMOOTH-MUSCLE()

We have cloned cDNAs encoding the alpha- and beta-subunits of a large- conductance Ca2+-activated K+ channel (BK channel) from canine colonic smooth muscle (cslo-alpha and cslo-beta). Nucleotide sequence homolog y of cslo-alpha with mslo and dslo suggests that it is the canine homo logue of these genes. The carboxy-terminal end of the protein is the m ost diverse between species, and we have also found alternative exons in cslo-alpha in this region. We have identified a unique splice site in the carboxy-terminal region of cslo-alpha, which we term site 5. No rthern analysis demonstrates expression of both alpha- and beta-subuni ts in all canine vascular and visceral smooth muscles tested. Expressi on of alpha- alone and alpha + beta-subunit cRNA in Xenopus oocytes re sults in a Ca2+- and voltage-dependent conductance. The activity of al pha/beta-channels, measured as either changes in the voltage of half-m aximal activation (V-0.5) in open probability (NPo) or in the normaliz ed conductance (G/G(max)), was more sensitive to [Ca2+](free) than cha nnels composed of the alpha-subunit alone. Neither alpha- nor alpha/be ta-channels expressed in membrane patches of Xenopus oocytes were foun d to be regulated by protein kinase G.