Abstract 5532: Regulation of Cardiac Nitric Oxide Synthase-1 Localization by the Adaptor Protein CAPON
Background: The mechanism that underlies nitric oxide synthase-1 (NOS1) redistribution within cardiac myocytes in cardiomyopathies or following myocardial infarction (MI) remains unknown. Here we tested the hypothesis that CAPON, an adaptor protein with a C-terminal PDZ-binding domain that binds NOS1 in the brain, contributes to NOS1 localization in specific organelles within cardiomyocytes.
Methods and Results: Ventricular cardiomyocytes and whole heart homogenates were isolated from sham and myocardial infarcted (MI) wild-type (C57BL/6) and NOS1-knock-out mice for quantification of CAPON mRNA and protein expression levels. CAPON and NOS1 were present and enriched in isolated cardiac sarcoplasmic reticulum (SR) fractions and co-immunoprecipitated with both μ and α isoforms of NOS1 in whole heart lysates. Confocal microscopy images revealed CAPON exhibiting a t-tubular staining pattern and did not co-localize with caveolin-3. The co-localization of CAPON and NOS1 in the SR and mitochondria was further confirmed by dual immuno-gold labeling with electron microscopy. In mice submitted to MI, there was a decrease in CAPON abundance in whole heart lysates. In addition, electron microscopy showed a co-localization of CAPON and NOS1 with caveolin-3 in the plasma membrane caveolae of cardiac myocytes.
Conclusion: In cardiac myocytes, CAPON plays an important role in NOS1 localization in the SR. This is the first study to show that CAPON abundance is reduced post-MI and this is associated with the translocation of NOS1 from the SR to the plasma membrane caveolae of cardiac myocytes. These findings have important implications for nitroso-redox balance within the heart in health and disease.