Monophosphorylation of cardiac troponin-I at Ser-23/24 is sufficient to regulate cardiac myofibrillar Ca2+ sensitivity and calpain-induced proteolysis


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Martin-Garrido A., Biesiadecki B. J. , Salhi H. E. , Shaifta Y., dos Remedios C. G. , Ayaz-Guner Ş. , et al.

JOURNAL OF BIOLOGICAL CHEMISTRY, cilt.293, ss.8588-8599, 2018 (SCI İndekslerine Giren Dergi)

  • Cilt numarası: 293 Konu: 22
  • Basım Tarihi: 2018
  • Doi Numarası: 10.1074/jbc.ra117.001292
  • Dergi Adı: JOURNAL OF BIOLOGICAL CHEMISTRY
  • Sayfa Sayısı: ss.8588-8599

Özet

The acceleration of myocardial relaxation produced by -adrenoreceptor stimulation is mediated in part by protein kinase A (PKA)-mediated phosphorylation of cardiac troponin-I (cTnI), which decreases myofibrillar Ca2+ sensitivity. Previous evidence suggests that phosphorylation of both Ser-23 and Ser-24 in cTnI is required for this Ca2+ desensitization. PKA-mediated phosphorylation also partially protects cTnI from proteolysis by calpain. Here we report that protein kinase D (PKD) phosphorylates only one serine of cTnI Ser-23/24. To explore the functional consequences of this monophosphorylation, we examined the Ca2+ sensitivity of force production and susceptibility of cTnI to calpain-mediated proteolysis when Ser-23/24 of cTnI in mouse cardiac myofibrils was nonphosphorylated, mono-phosphorylated, or bisphosphorylated (using sequential incubations in -phosphatase, PKD, and PKA, respectively). Phos-tag gels, Western blotting, and high-resolution MS revealed that PKD produced >90% monophosphorylation of cTnI, primarily at Ser-24, whereas PKA led to cTnI bisphosphorylation exclusively. PKD markedly decreased the Ca2+ sensitivity of force production in detergent-permeabilized ventricular trabeculae, whereas subsequent incubation with PKA produced only a small further fall of Ca2+ sensitivity. Unlike PKD, PKA also substantially phosphorylated myosin-binding protein-C and significantly accelerated cross-bridge kinetics (k(tr)). After phosphorylation by PKD or PKA, cTnI in isolated myofibrils was partially protected from calpain-mediated degradation. We conclude that cTnI monophosphorylation at Ser-23/24 decreases myofibrillar Ca2+ sensitivity and partially protects cTnI from calpain-induced proteolysis. In healthy cardiomyocytes, the basal monophosphorylation of cTnI may help tonically regulate myofibrillar Ca2+ sensitivity.