Silencing calcineurin A subunit reduces SERCA2 expression in cardiac myocytes
| Autor: | Giuseppe Inesi, Anand Mohan Prasad |
|---|---|
| Rok vydání: | 2011 |
| Předmět: |
Thapsigargin
Physiology Protein subunit Blotting Western Fluorescent Antibody Technique Biology Sarcoplasmic Reticulum Calcium-Transporting ATPases chemistry.chemical_compound Physiology (medical) Animals Gene silencing Myocyte Myocytes Cardiac RNA Small Interfering Cells Cultured Volume concentration CALCINEURIN A Analysis of Variance Reverse Transcriptase Polymerase Chain Reaction Calcineurin Molecular biology Rats Up-Regulation Cell biology Sarcoplasmic Reticulum chemistry cardiovascular system Calcium Signaling and Stress Response Cardiology and Cardiovascular Medicine Intracellular |
| Zdroj: | American Journal of Physiology-Heart and Circulatory Physiology. 300:H173-H180 |
| ISSN: | 1522-1539 0363-6135 |
| DOI: | 10.1152/ajpheart.00841.2010 |
| Popis: | Resting intracellular Ca2+can be raised, in neonatal rat cardiac myocytes, by exposure to very low concentration of thapsigargin (TG). Such a Ca2+rise yields calcineurin (CN) activation demonstrated by increased expression of transfected luciferase cDNA under control of nuclear factor of activated T-cells (NFAT) promoter and increased translocation of NFAT to nuclei. We found that exposure of cardiac myocytes to TG is followed by increase of sarcroplasmic reticulum Ca2+transport ATPase (SERCA2) expression, which is further increased when CN inactivation by CAMKII (calmodulin-dependent kinase) is prevented with KN93 (CAMKII inhibitor). On the other hand, SERCA2 expression is reduced by CN inhibition with cyclosporine. We have now induced calcineurin A (CNA) α- or β-subunit gene silencing with small interfering RNA (siRNA) and observed strong interference with expression of SERCA2, both in control myocytes and following exposure to TG. Such interference is also obtained following NFAT displacement from CN with 9,10-dihydro-9,10[1′,2′]-benzenoanthracene-1,4-dione (INCA-6). We have also observed analogous effects on expression of phospholamban (PLB) and Na+/Ca2+exchanger (NCX). Pertinent to these findings, we have identified, by in-silico analysis, NFAT binding sites in SERCA2, PLB, and NCX1 promoters. Our experiments indicate that activation of the calcineurin-NFAT pathway by rise of resting cytosolic Ca2+elevates transcription/expression of SERCA2, PLB, and NCX1, providing a homeostatic mechanism for long-term control of cytosolic Ca2+. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|