Abstract 3294: Blockade of Muscle Lim Protein Nuclear Shuttling Impairs Myocyte Remodelling in Response to Hypertrophic Stimuli
Cardiac hypertrophy in response to increased hemodynamic load is mediated through cellular stretch sensors that convert the mechanical stimulus into a biological response. However, prolonged hemodynamic load may lead to impaired mechanosensing thereby leading to decompensation and heart failure. CSRP3 or Muscle LIM protein (MLP) is a mechanosensor and nucleocytoplasmic shuttling protein in cardiac myocytes. Mutations in the MLP gene lead to human cardiomyopathy but the reasons for this are poorly understood. We assessed the hypothesis that nuclear cycling of MLP is required for myocyte adaptation to hypertrophic stimuli. To test this, cultured neonatal rat myocytes were treated with pharmacological or mechanical stimuli. Cells were then fractionated to assess the subcellular distribution of MLP. Treatment with the calcium channel blocker verapamil decreased nuclear MLP by 71% (p<0.01, n=5 cultures) and the myofilament crossbridge inhibitor 2,3-butanedione monoxime decreased it by 43% (p<0.05, n=5). However, the alpha adrenergic agonist phenylephrine increased nuclear MLP by 132% (p<0.01, n=5). These results suggest that myocyte contractility and calcium cycling regulate MLP subcellular localization. Using cell permeable synthetic peptides containing the nuclear localization signal (NLS) of MLP, nuclear import of the protein was inhibited in cultured rat neonatal myocytes. The peptide containing the NLS of MLP localized to the nucleolus suggesting a mechanism by which MLP may regulate ribosomal protein synthesis. Cellular protein accumulation and brain natriuretic peptide expression increased 40% (p<0.05, n=6) and 65% (p<0.01, n=5) respectively following cyclic stretch. However, these increases were prevented when MLP nuclear cycling was prevented. Furthermore, cyclic strain of myocytes after prior MLP-NLS treatment to remove nuclear MLP resulted in disarrayed sarcomeres and a 67% (p<0.01, n=6) decrease in alpha actinin protein expression. In conclusion, these data suggest that nuclear cycling of MLP is required for remodeling of the myofilaments and gene expression following hypertrophic stimuli in cardiac myocytes. These findings may explain why mutations in this gene lead to cardiomyopathy in the human heart.
This research has received full or partial funding support from the American Heart Association, National Center.