title: JPH-2 interacts with Cai-handling proteins and ion channels in dyads: Contribution to premature ventricular contraction-induced cardiomyopathy.
abstract: BACKGROUND
  In a canine model of premature ventricular contraction-induced cardiomyopathy (PVC-CM), Cav1.2 is downregulated and misplaced from transverse tubules (T tubules). Junctophilin-2 (JPH-2) is also downregulated.
  
  
  OBJECTIVE
  The objectives of this study were to understand the role of JPH-2 in PVC-CM and to probe changes in other proteins involved in dyad structure and function.
  
  
  METHODS
  We quantify T-tubule contents (di-8-ANEPPS fluorescence in live myocytes), examine myocyte ultrastructures (electron microscopy), probe JPH-2-interacting proteins (co-immunoprecipitation), quantify dyad and nondyad protein levels (immunoblotting), and examine subcellular distributions of dyad proteins (immunofluorescence/confocal microscopy). We also test direct JPH-2 modulation of channel function (vs indirect modulation through dyad formation) using heterologous expression.
  
  
  RESULTS
  PVC myocytes have reduced T-tubule contents but otherwise normal ultrastructures. Among 19 proteins examined, only JPH-2, bridging integrator-1 (BIN-1), and Cav1.2 are highly downregulated in PVC hearts. However, statistical analysis indicates a general reduction in dyad protein levels when JPH-2 is downregulated. Furthermore, several dyad proteins, including Na/Ca exchanger, are missing or shifted from dyads to the peripheral surface in PVC myocytes. JPH-2 directly or indirectly interacts with Cai-handling proteins, Cav1.2 and KCNQ1, although not BIN-1 or other scaffolding proteins tested. Expression in mammalian cells that do not have dyads confirms direct JPH-2 modulation of the L-type Ca channel current (Cav1.2/voltage-gated Ca channel β subunit 2) and slow delayed rectifier current (KCNQ1/KCNE1).
  
  
  CONCLUSIONS
  JPH-2 is more than a "dyad glue": it can modulate Cai handling and ion channel function in the dyad region. Downregulation of JPH-2, BIN-1, and Cav1.2 plays a deterministic role in PVC-CM. Dissecting the hierarchical relationship among the three is necessary for the design of therapeutic interventions to prevent the progression of PVC-CM.
authors.affiliation: Department of Physiology and Biophysics, Virginia Commonwealth University, Richmond, Virginia.
authors.forename: Min
authors.initials: M
authors.lastname: Jiang
authors.affiliation: Department of Physiology and Biophysics, Virginia Commonwealth University, Richmond, Virginia.
authors.forename: Mei
authors.initials: M
authors.lastname: Zhang
authors.affiliation: Department of Medicine/Cardiology Division, McGuire VA Medical Center, Richmond, Virginia.
authors.forename: Maureen
authors.initials: M
authors.lastname: Howren
authors.affiliation: Department of Physiology and Biophysics, Virginia Commonwealth University, Richmond, Virginia.
authors.forename: Yuhong
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Medicine/Cardiology Division, McGuire VA Medical Center, Richmond, Virginia.
authors.forename: Alex
authors.initials: A
authors.lastname: Tan
authors.affiliation: Cellular and Molecular Arrhythmia Research Program, Department of Medicine, University of Wisconsin, Madison, Wisconsin.
authors.forename: Ravi C
authors.initials: RC
authors.lastname: Balijepalli
authors.affiliation: Department of Physiology and Biophysics, Virginia Commonwealth University, Richmond, Virginia.
authors.forename: Jose F
authors.initials: JF
authors.lastname: Huizar
authors.affiliation: Department of Physiology and Biophysics, Virginia Commonwealth University, Richmond, Virginia. Electronic address: gtseng@vcu.edu.
authors.forename: Gea-Ny
authors.initials: GN
authors.lastname: Tseng
mesh_terms: D000818:Animals; D009202:Cardiomyopathies / Q000209:etiology / Q000378:metabolism* / Q000473:pathology; D002478:Cells, Cultured; D004195:Disease Models, Animal; D004285:Dogs; D015151:Immunoblotting; D008565:Membrane Proteins / Q000096:biosynthesis*; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D009206:Myocardium / Q000378:metabolism* / Q000473:pathology; D032383:Myocytes, Cardiac / Q000378:metabolism / Q000648:ultrastructure; D019831:Sodium-Calcium Exchanger / Q000096:biosynthesis*; D018879:Ventricular Premature Complexes / Q000150:complications / Q000378:metabolism* / Q000473:pathology
chemical_list: D008565:Membrane Proteins; D019831:Sodium-Calcium Exchanger; C412507:junctophilin; C110374:sodium-calcium exchanger 1
keywords: Cardiomyopathy; Dyad; Excitation-contraction coupling; Premature ventricular contractions; Sarcoplasmic reticulum; T tubules