title: Rescue of a trafficking defective human pacemaker channel via a novel mechanism: roles of Src, Fyn, and Yes tyrosine kinases.
abstract: Therapeutic strategies such as using channel blockers and reducing culture temperature have been used to rescue some long QT-associated voltage-gated potassium Kv trafficking defective mutant channels. A hyperpolarization-activated cyclic nucleotide-gated HCN4 pacemaker channel mutant (D553N) has been recently found in a patient associated with cardiac arrhythmias including long QT. D553N showed the defective trafficking to the cell surface, leading to little ionic current expression (loss-of-function). We show in this report that enhanced tyrosine phosphorylation mediated by Src, Fyn, and Yes kinases was able to restore the surface expression of D553N for normal current expression. Src or Yes, but not Fyn, significantly increased the current density and surface expression of D553N. Fyn accelerated the activation kinetics of the rescued D553N. Co-expression of D553N with Yes exhibited the slowest activation kinetics of D553N. Src, Fyn, and Yes significantly enhanced the tyrosine phosphorylation of D553N. A combination of Src, Fyn, and Yes rescued the current expression and the gating of D553N comparable with those of wild-type HCN4. In conclusion, we demonstrate a novel mechanism using three endogenous Src kinases to rescue a trafficking defective HCN4 mutant channel (D553N) by enhancing the tyrosine phosphorylation of the mutant channel protein.
authors.affiliation: Department of Physiology and Pharmacology, West Virginia University School of Medicine, Center for Cardiovascular and Respiratory Sciences, Morgantown, West Virginia 26506, USA.
authors.forename: Yen-Chang
authors.initials: YC
authors.lastname: Lin
authors.forename: Jianying
authors.initials: J
authors.lastname: Huang
authors.forename: Hong
authors.initials: H
authors.lastname: Kan
authors.forename: Jefferson C
authors.initials: JC
authors.lastname: Frisbee
authors.forename: Han-Gang
authors.initials: HG
authors.lastname: Yu
mesh_terms: D002460:Cell Line; D054815:Cyclic Nucleotide-Gated Cation Channels / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D009124:Muscle Proteins / Q000235:genetics* / Q000378:metabolism; D050505:Mutant Proteins / Q000378:metabolism; D020125:Mutation, Missense; D010766:Phosphorylation; D015221:Potassium Channels; D021381:Protein Transport; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D051076:Proto-Oncogene Proteins c-fyn / Q000502:physiology; D051575:Proto-Oncogene Proteins c-yes / Q000502:physiology; D019061:src-Family Kinases / Q000502:physiology
chemical_list: D054815:Cyclic Nucleotide-Gated Cation Channels; C119587:HCN4 protein, human; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D009124:Muscle Proteins; D050505:Mutant Proteins; D015221:Potassium Channels; D011505:Protein-Tyrosine Kinases; C494957:FYN protein, human; D051076:Proto-Oncogene Proteins c-fyn; D051575:Proto-Oncogene Proteins c-yes; C489834:YES1 protein, human; D019061:src-Family Kinases