Block by extracellular divalent cations of Drosophila big brain channels expressed in Xenopus oocytes.
Drosophila Big Brain (BIB) is a transmembrane protein encoded by the neurogenic gene big brain (bib), which is important for early development of the fly nervous system. BIB expressed in Xenopus oocytes is a monovalent cation channel modulated by tyrosine kinase signaling. Results here demonstrate that the BIB conductance shows voltage- and dose-dependent block by extracellular divalent cations Ca(2+) and Ba(2+) but not by Mg(2+) in wild-type channels. Site-directed mutagenesis of negatively charged glutamate (Glu(274)) and aspartate (Asp(253)) residues had no effect on divalent cation block. However, mutation of a conserved glutamate at position 71 (Glu(71)) in the first transmembrane domain (M1) altered channel properties. Mutation of Glu(71) to Asp introduced a new sensitivity to block by extracellular Mg(2+); substitutions with asparagine or glutamine decreased whole-cell conductance; and substitution with lysine compromised plasma membrane expression. Block by divalent cations is important in other ion channels for voltage-dependent function, enhanced signal resolution, and feedback regulation. Our data show that the wild-type BIB conductance is attenuated by external Ca(2+), suggesting that endogenous divalent cation block might be relevant for enhancing signal resolution or voltage dependence for the native signaling process in neuronal cell fate determination.
Program in Pharmacology and Toxicology, Department of Physiology, College of Medicine, University of Arizona, Tucson, Arizona 85724, USA.
Gina M
GM
Yanochko
Andrea J
AJ
Yool
D000818:Animals; D001464:Barium / Q000494:pharmacology*; D002118:Calcium / Q000494:pharmacology*; D002413:Cations, Divalent / Q000494:pharmacology; D002478:Cells, Cultured; D004330:Drosophila; D029721:Drosophila Proteins / Q000187:drug effects / Q000502:physiology*; D045604:Extracellular Fluid / Q000378:metabolism; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D007473:Ion Channels / Q000187:drug effects / Q000502:physiology*; D008274:Magnesium / Q000494:pharmacology*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008565:Membrane Proteins / Q000187:drug effects / Q000502:physiology*; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000187:drug effects / Q000502:physiology*; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D013329:Structure-Activity Relationship; D014982:Xenopus laevis
D002413:Cations, Divalent; D029721:Drosophila Proteins; D007473:Ion Channels; D008565:Membrane Proteins; D011994:Recombinant Proteins; C070621:bib protein, Drosophila; D001464:Barium; D008274:Magnesium; D002118:Calcium