Cell signaling pathways regulating myosin regulatory light chain (LC20) phosphorylation contribute to determining contractile responses in smooth muscles. Following excitation and contraction, phasic smooth muscles, such as digestive tract and urinary bladder, undergo a relaxation due to a decline of cellular [Ca²⁺] and a decreased Ca²⁺ sensitivity of LC20 phosphorylation, named Ca²⁺ desensitization. Here, we determined mechanisms underlying the temporal Ca²⁺ desensitization of LC20 phosphorylation in phasic smooth muscles using permeabilized strips of mouse ileum and urinary bladder. Upon the stimulation with pCa6.0 at 20°C, the contraction and the LC20 phosphorylation peaked within 30 sec and then declined to about 50% of the peak force at 2 min after stimulation. During the relaxation phase after the contraction, the LC20 kinase (MLCK) was inactivated, but no fluctuation in the LC20 phosphatase activity occurred, suggesting that the MLCK inactivation is a cause of the Ca²⁺-induced Ca²⁺-desensitization of LC20 phosphorylation. The MLCK inactivation was associated with phosphorylation at the calmodulin binding domain of the kinase. Treatment with antagonists for CaMKKß (STO-609 and TIM-063) attenuated both the phasic response of the contraction and MLCK phosphorylation, whereas neither CaMKII, AMPK nor PAK induced the MLCK inactivation in phasic smooth muscles. Conversely, PP2A inhibition amplified the phasic response. Signaling pathways through CaMKKß and PP2A may contribute to regulating the Ca²⁺ sensitivity of MLCK and the contractile response of phasic smooth muscles.

中文翻译：

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CaMKKß/PP2A 通路诱导的阶段性平滑肌肌球蛋白轻链激酶的时间 Ca2+ 脱敏。

调节肌球蛋白调节轻链 (LC20) 磷酸化的细胞信号通路有助于确定平滑肌的收缩反应。在兴奋和收缩之后，阶段性平滑肌，例如消化道和膀胱，由于细胞 [Ca ²⁺ ] 的下降和 LC20 磷酸化的Ca ²⁺敏感性降低而松弛，称为 Ca ²⁺脱敏。在这里，我们确定了时间 Ca ^{2+ 的}潜在机制使用透化小鼠回肠和膀胱条带对阶段性平滑肌中的 LC20 磷酸化脱敏。在 20°C 下用 pCa6.0 刺激后，收缩和 LC20 磷酸化在 30 秒内达到峰值，然后在刺激后 2 分钟下降到峰值力的约 50%。在收缩后的松弛阶段，LC20 激酶（MLCK）失活，但 LC20 磷酸酶活性没有发生波动，表明 MLCK 失活是 Ca ²⁺诱导的 Ca ^{2+ 的原因}- LC20 磷酸化的脱敏。MLCK 失活与激酶钙调蛋白结合域的磷酸化有关。用 CaMKKß 拮抗剂（STO-609 和 TIM-063）治疗减弱了收缩的阶段性反应和 MLCK 磷酸化，而 CaMKII、AMPK 和 PAK 都不会诱导阶段性平滑肌中的 MLCK 失活。相反，PP2A 抑制放大了阶段性反应。通过 CaMKKß 和 PP2A 的信号通路可能有助于调节MLCK的 Ca ²⁺敏感性和阶段性平滑肌的收缩反应。