Zhou Chaoming, Ramaswamy Swarna S, Johnson Derrick E, Vitturi Dario A, Schopfer Franciso J, Freeman Bruce A, Hudmon Andy, Levitan Edwin S
Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, 200 Lothrop Street, Pittsburgh, PA 15261 USA.
Department of Biochemistry and Molecular Biology and Stark Neuroscience Research Institute, Indiana University School of Medicine, 320 West 15th Street, Indianapolis, IN 46202 USA.
Sci Rep. 2016 Apr 15;6:23416. doi: 10.1038/srep23416.
Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) oxidation controls excitability and viability. While hydrogen peroxide (H2O2) affects Ca(2+)-activated CaMKII in vitro, Angiotensin II (Ang II)-induced CaMKIIδ signaling in cardiomyocytes is Ca(2+) independent and requires NADPH oxidase-derived superoxide, but not its dismutation product H2O2. To better define the biological regulation of CaMKII activation and signaling by Ang II, we evaluated the potential for peroxynitrite (ONOO(-)) to mediate CaMKII activation and downstream Kv4.3 channel mRNA destabilization by Ang II. In vitro experiments show that ONOO(-) oxidizes and modestly activates pure CaMKII in the absence of Ca(2+)/CaM. Remarkably, this apokinase stimulation persists after mutating known oxidation targets (M281, M282, C290), suggesting a novel mechanism for increasing baseline Ca(2+)-independent CaMKII activity. The role of ONOO(-) in cardiac and neuronal responses to Ang II was then tested by scavenging ONOO(-) and preventing its formation by inhibiting nitric oxide synthase. Both treatments blocked Ang II effects on Kv4.3, tyrosine nitration and CaMKIIδ oxidation and activation. Together, these data show that ONOO(-) participates in Ang II-CaMKII signaling. The requirement for ONOO(-) in transducing Ang II signaling identifies ONOO(-), which has been viewed as a reactive damaging byproduct of superoxide and nitric oxide, as a mediator of GPCR-CaMKII signaling.
钙（Ca2+）/钙调蛋白依赖性蛋白激酶II（CaMKII）的氧化作用控制着细胞的兴奋性和活力。虽然过氧化氢（H2O2）在体外会影响Ca2+激活的CaMKII，但心肌细胞中血管紧张素II（Ang II）诱导的CaMKIIδ信号传导不依赖于Ca2+，需要NADPH氧化酶衍生的超氧化物，而不是其歧化产物H2O2。为了更好地界定Ang II对CaMKII激活和信号传导的生物学调节作用，我们评估了过氧亚硝酸盐（ONOO-）介导Ang II激活CaMKII以及使下游Kv4.3通道mRNA不稳定的可能性。体外实验表明，在没有Ca2+/钙调蛋白（CaM）的情况下，ONOO-会氧化并适度激活纯CaMKII。值得注意的是，在使已知的氧化靶点（M281、M282、C290）发生突变后，这种对无活性激酶的刺激仍然存在，这表明存在一种增加基线Ca2+非依赖性CaMKII活性的新机制。然后，通过清除ONOO-并抑制一氧化氮合酶来阻止其形成，测试了ONOO-在心脏和神经元对Ang II反应中的作用。这两种处理均阻断了Ang II对Kv4.3、酪氨酸硝化以及CaMKIIδ氧化和激活的影响。总之，这些数据表明ONOO-参与了Ang II-CaMKII信号传导。在转导Ang II信号中对ONOO-的需求表明，一直被视为超氧化物和一氧化氮的一种具有反应性的有害副产物的ONOO-，是G蛋白偶联受体（GPCR）-CaMKII信号传导的一种介质。
