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.
Yanochko
Gina M
GM
Program in Pharmacology and Toxicology, Department of Physiology, College of Medicine, University of Arizona, Tucson, Arizona 85724, USA.
Yool
Andrea J
AJ
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