TY - JOUR
T1 - Effects of mutant and antisense RNA of phospholamban on SR Ca2+-ATPase activity and cardiac myocyte contractility
AU - He, Huaping
AU - Meyer, Markus
AU - Martin, Jody L.
AU - McDonough, Patrick M.
AU - Ho, Peter
AU - Lou, Xiaojing
AU - Lew, Wilbur Y.W.
AU - Hilal-Dandan, Randa
AU - Dillmann, Wolfgang H.
PY - 1999/8/31
Y1 - 1999/8/31
N2 - Background - The delayed cardiac relaxation in failing hearts has been attributed to a reduced activity of sarcoplasmic reticulum Ca2+-ATPase (SERCA2). Phospholamban (PLB) inhibits SERCA2 activity and is therefore a potential target to improve the cardiac performance in heart failure. Methods and Results - Mutants of PLB (Adv/mPLB) or antisense RNA of PLB (Adv/asPLB) was expressed in cardiac myocytes by recombinant adenovirus, and their effects on SERCA2 activity and myocyte contractility were studied. One mPLB, K3E/R14E, pentamerized with endogenous PLB in neonatal myocytes and resulted in a 45% increase in the affinity of SERCA2 for Ca2+ and 27% faster diastolic Ca2+ decline as determined by SR 45Ca uptake assays and by indo 1-facilitated Ca2+ transient measurement, respectively. Edge-detection analysis of adult myocyte contractility showed a 74% increase in fractional shortening, accompanied by 115% increase in velocity of relengthening and 25% decrease in time to half-maximal relengthening. In parallel, infection of neonatal cardiac myocytes by Adv/asPLB decreased the endogenous PLB level by 54%, which was associated with a 35% increase in Ca2+ affinity of SERCA2 and 21% faster diastolic Ca2+ decline. However, in adult cardiac myocytes, Adv/asPLB failed to significantly alter the endogenous PLB level, the SERCA2 activity, or most of the contractile parameters. Conclusions - K3E/R14E is a dominant negative mutant of PLB that disrupts the structural integrity and function of the endogenous PLB and consequently enhances SERCA2 activity and myocyte contractility. In neonatal myocytes, the decrease in steady-state abundance of PLB by asPLB also leads to increased SERCA2 activity.
AB - Background - The delayed cardiac relaxation in failing hearts has been attributed to a reduced activity of sarcoplasmic reticulum Ca2+-ATPase (SERCA2). Phospholamban (PLB) inhibits SERCA2 activity and is therefore a potential target to improve the cardiac performance in heart failure. Methods and Results - Mutants of PLB (Adv/mPLB) or antisense RNA of PLB (Adv/asPLB) was expressed in cardiac myocytes by recombinant adenovirus, and their effects on SERCA2 activity and myocyte contractility were studied. One mPLB, K3E/R14E, pentamerized with endogenous PLB in neonatal myocytes and resulted in a 45% increase in the affinity of SERCA2 for Ca2+ and 27% faster diastolic Ca2+ decline as determined by SR 45Ca uptake assays and by indo 1-facilitated Ca2+ transient measurement, respectively. Edge-detection analysis of adult myocyte contractility showed a 74% increase in fractional shortening, accompanied by 115% increase in velocity of relengthening and 25% decrease in time to half-maximal relengthening. In parallel, infection of neonatal cardiac myocytes by Adv/asPLB decreased the endogenous PLB level by 54%, which was associated with a 35% increase in Ca2+ affinity of SERCA2 and 21% faster diastolic Ca2+ decline. However, in adult cardiac myocytes, Adv/asPLB failed to significantly alter the endogenous PLB level, the SERCA2 activity, or most of the contractile parameters. Conclusions - K3E/R14E is a dominant negative mutant of PLB that disrupts the structural integrity and function of the endogenous PLB and consequently enhances SERCA2 activity and myocyte contractility. In neonatal myocytes, the decrease in steady-state abundance of PLB by asPLB also leads to increased SERCA2 activity.
KW - Enzymes
KW - Phospholamban
KW - RNA
KW - Sarcoplasmic reticulum
KW - Viruses
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U2 - 10.1161/01.CIR.100.9.974
DO - 10.1161/01.CIR.100.9.974
M3 - Article
C2 - 10468529
AN - SCOPUS:0033621151
SN - 0009-7322
VL - 100
SP - 974
EP - 980
JO - Circulation
JF - Circulation
IS - 9
ER -