Molecular cloning and permanent expression in a neuroblastoma cell line of a fast inactivating potassium channel from bovine adrenal medulla

Abstract

Using a cDNA library from bovine adrenal medulla, we have isolated cDNAs coding for a potassium channel. These cDNAs encode a 660-amino acid protein that has a molecular weight of 73,288 kDa and no amino-terminal signal peptide. We have called it BAK4. Analysis of its sequence reveals close similarity (94% homology) with a recently described potassium channel from rat brain (RCK4) and heart (RHK1). Neuroblastoma cells (Neuro-2a cell line) were stably transfected with BAK4 DNA. Expression of the DNA was under the control of a heat-shock promoter. Several clones, that were isolated by neomycin resistance selection, had integrated the plasmid DNA in a stable form. Upon heat induction, these cells produced BAK4 RNA and a potassium outward current, not present in control non-transfected cells. The current, which was transient and decayed markedly during the duration of 200 ms-pulses, can be described as a Ik(A) potassium current. The expression of these types of channels in brain (RCK4,RHK1), heart (RHK1) and adrenal medulla (BAK4) suggest their possible implication in important functions for the cell.