Role of dual-site phospholamban phosphorylation in the stunned heart:: insights from phospholamban site-specific mutants
Role of dual-site phospholamban phosphorylation in the stunned heart:: insights from phospholamban site-specific mutants
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DOI：
10.1152/ajpheart.00209.2003
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发表时间：
2003-09-01
影响因子：
4.8
通讯作者：
Mattiazzi, A
中科院分区：
文献类型：
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作者：
Said, M;Vittone, L;Mattiazzi, A
受磷蛋白（PLB）在Ser（16）（蛋白激酶A位点）和Thr（17）[Ca 2 +/钙调蛋白激酶II（CaMKII）位点]的磷酸化增加肌浆网Ca 2+摄取和心肌收缩性和舒张性。在缺血-再灌注大鼠心脏中，我们先前发现缺血诱导的Ser（16）磷酸化依赖于β-肾上腺素能刺激，缺血和再灌注诱导的Thr（17）磷酸化依赖于Ca 2+内流。为了阐明这两个PLB磷酸化位点和缺血后机械恢复之间的关系，在CaMKII抑制剂KN-93（1 μ M）或β-肾上腺素能阻滞剂dl-普萘洛尔（1 μ M）存在和不存在的情况下，对大鼠心脏进行缺血-再灌注。KN-93可抑制再灌注后Thr（17）磷酸化，抑制缺血后心肌收缩和舒张功能的恢复。dl-普萘洛尔降低缺血诱导的Ser（16）磷酸化，但不能改变收缩恢复。为了进一步了解两个PLB磷酸化位点在缺血后机械恢复中的功能作用，使用表达野生型PLB（PLB-WT）或PLB突变体的转基因小鼠，其中Thr（17）或Ser（16）被Ala（分别为PLB-T17 A和PLB-S16 A）替换成PLB-无效背景。两种PLB突变体均显示出比PLB-WT更低的收缩恢复。然而，在PLB-T17 A中，这种恢复在再灌注的整个沿着都显著受损，而在PLB-S16 A中，这种恢复仅在再灌注开始时被抑制。此外，与PLB-WT相比，PLB-T17 A的舒张恢复延迟，而PLB-S16 A的舒张恢复没有变化。这些发现表明，虽然两个PLB磷酸化位点都参与了缺血后的机械恢复，但Thr（17）似乎起着主要作用。
Phosphorylation of phospholamban (PLB) at Ser(16) ( protein kinase A site) and at Thr(17) [Ca2+/calmodulin kinase II (CaMKII) site] increases sarcoplasmic reticulum Ca2+ uptake and myocardial contractility and relaxation. In perfused rat hearts submitted to ischemia-reperfusion, we previously showed an ischemia-induced Ser(16) phosphorylation that was dependent on beta-adrenergic stimulation and an ischemia and reperfusion-induced Thr(17) phosphorylation that was dependent on Ca2+ influx. To elucidate the relationship between these two PLB phosphorylation sites and postischemic mechanical recovery, rat hearts were submitted to ischemia-reperfusion in the absence and presence of the CaMKII inhibitor KN-93 (1 muM) or the beta-adrenergic blocker dl-propranolol (1 muM). KN-93 diminished the reperfusion-induced Thr(17) phosphorylation and depressed the recovery of contraction and relaxation after ischemia. dl-Propranolol decreased the ischemia-induced Ser(16) phosphorylation but failed to modify the contractile recovery. To obtain further insights into the functional role of the two PLB phosphorylation sites in postischemic mechanical recovery, transgenic mice expressing wild-type PLB (PLB-WT) or PLB mutants in which either Thr(17) or Ser(16) were replaced by Ala (PLB-T17A and PLB-S16A, respectively) into the PLB-null background were used. Both PLB mutants showed a lower contractile recovery than PLB-WT. However, this recovery was significantly impaired all along reperfusion in PLB-T17A, whereas it was depressed only at the beginning of reperfusion in PLB-S16A. Moreover, the recovery of relaxation was delayed in PLB-T17A, whereas it did not change in PLB-S16A, compared with PLB-WT. These findings indicate that, although both PLB phosphorylation sites are involved in the mechanical recovery after ischemia, Thr(17) appears to play a major role.