Cloning of TTX-resistant Na⁺channels in NB-1 cell lines

Abstract

Both Tetrodotoxin-sensitive (TTX-S) and resistant (TTX-R) voltage-gated Na⁺ channel are expressed in human neuroblastoma NB-1 cells , but the gene encoding the TTX-R Na⁺ channel is not identified. In this study, we have cloned α subunit of TTX-R Na⁺ channel from NB-1 cells by reverse transcriptase-polymerase chain reaction (RT-PCR) and designated it as hNbR1. The longest open reading frame of NbH1 encodes 2016 amino acid residues. The presence of a cysteine residue (c373) in the pore region domain I suggests that hNbR1 is resistant to TTX. Sequence comparisons indicated that hNbR1 is highly homologous to human cardiac SCN5A (99.2% nucleotide homology) except the region between domain I S3 and S4. At least one alternative splicing variant, which is characterized by a deletion (54 amino acids) the intracellular loop between domains II and III, is found. These results suggest that SCN5A is more widely distributed than previously thought and contributes to TTX-R Na⁺ current in neural tissues. [Jpn J Physiol 54 Suppl:S135 (2004)]