Reduction of four-and-a-half LIM-protein 2 expression occurs in human left ventricular failure and leads to altered localization and reduced activity of metabolic enzymes

Abstract

We sought to identify changes in four-and-a-half LIM-protein 2 levels and location in human cardiomyocytes during the transition from compensated aortic stenosis to left ventricular failure. We also sought to characterize four-and-a-half LIM-protein 2 binding with the metabolic enzymes phosphofructokinase 2, adenylate kinase, and creatine kinase M isoform during this transition and their consequential subcellular localization in failing human ventricles. Left ventricular biopsy specimens from selected patients undergoing aortic valve replacement for aortic stenosis were allocated to one of 2 groups: (1) nondilated with preserved left ventricular function (nonfailing group, n = 16) and (2) grossly dilated with poor left ventricular function (failing group, n = 15). These were compared with a control group of unused donor hearts (n = 6). Protein levels and subcellular localization were determined by means of Western blotting and immunofluorescence. Four-and-a-half LIM-protein 2 binding to adenylate kinase, creatine kinase M isoform, or phosphofructokinase 2 was studied by means of coimmunoprecipitation. Phosphofructokinase 2, adenylate kinase, and creatine kinase M isoform activities were assayed in protein extractions. Four-and-a-half LIM-protein 2 levels were preserved in nonfailing hypertrophied hearts but reduced by 53% in failing hearts. The pattern of four-and-a-half LIM-protein 2 staining was disrupted in failing hearts: four-and-a-half LIM-protein 2 was lost from the sarcomere but present in the perinuclear Golgi apparatus complex. Phosphofructokinase 2, adenylate kinase, and creatine kinase M isoform coimmunoprecipitated in vitro and colocalized with four-and-a-half LIM-protein 2 in both hypertrophied and failing hearts. Phosphofructokinase 2 and adenylate kinase activities were reduced to 77% and 58% of normal values in compensated aortic stenosis, with phosphofructokinase 2 activity decreased further to 56% of normal value in failing hearts, but creatine kinase activity remained unchanged. Altered four-and-a-half LIM-protein 2 expression in heart failure is associated with disruption of the normal subcellular localization of phosphofructokinase 2, adenylate kinase, and creatine kinase M isoform and reduced activity of phosphofructokinase 2 and adenylate kinase, which might have important consequences for myocardial energy metabolism in heart failure.