Overexpressed chloride intracellular channel protein CLIC4 (p64H1) is an essential component of novel plasma membrane anion channels.
Chloride intracellular channel protein CLIC4 is a putative organellar anion channel or channel regulator with an unusual dual cytoplasmic and integral membrane localisation. To investigate its contribution to cellular anion channel activity, the protein was overexpressed in stably transfected HEK-293 cells. Patch-clamp recording revealed CLIC4-associated indanyloxyacetic acid-sensitive (IC(50) approximately 100 microM) plasma membrane currents showing mild outward rectification, and novel low conductance (approximately 1pS) CLIC4-associated anion channels were resolved at the single-channel level. The CLIC4-associated channels were inhibited by anti-CLIC4 antibodies, including a monoclonal antibody directed against a FLAG epitope fused to the C-terminus of CLIC4, but only when these were applied to the cytoplasmic (not the external) face of the membrane. CLIC4 is thus an essential molecular component of novel cellular anion channels and the C-terminus of the integral membrane form of CLIC4 is cytoplasmic.
Proutski
I
I
Department of Biomedical Sciences, University of Edinburgh, UK.
Karoulias
N
N
Ashley
R H
RH
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D018118:Chloride Channels / Q000235:genetics / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism; D006801:Humans; D018408:Patch-Clamp Techniques; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D000911:Antibodies, Monoclonal; D018118:Chloride Channels; D002712:Chlorides; C467578:Clic4 protein, rat; D011993:Recombinant Fusion Proteins