The dominant negative LQT2 mutation A561V reduces wild-type HERG expression

HERG(1) K+ channel mutations are responsible for one form of dominantly inh erited Long QT syndrome (LQT), Some LQT mutations exert a dominant negative effect on wild-type current expression. To investigate mechanisms of domin ant-negative behavior, we co-expressed wild-type HERG with the A561V mutant in mammalian cells. Transfection with various cDNA ratios produced HERG K current densities that approached a predicted binomial distribution where mutant and wild-type subunits co-assemble in a tetramer with nearly complet e dominance. Using C terminus myc-tagged wild-type HERG we specifically fol lowed the mutant's effect on full-length wild-type HERG protein expression. Co-expression with A561V reduced the abundance of full-length wild-type HE RG protein comparable to the current reduction. Reduction of wild-type prot ein was due to decreased synthesis and increased turnover. Conditions facil itating protein folding (growth at 30 degrees C, or in 10% glycerol) result ed in partial rescue from the dominant effect, as did the 26 S proteosome i nhibitor ALLN. Thus, for A561V, dominant negative effects result from assem bly of wild-type subunits with mutant very early in production leading to r apid recognition of mutant channels and targeting for proteolysis. These re sults establish protein misfolding, cellular proofreading, and bystander in volvement as contributing mechanisms for dominant effects in LQT2.