Graduate School of Agricultural Science, Kobe University, 1-1 Rokkodai-cho, Nada-ku, Kobe, Hyogo, 657-8501, Japan.
J Plant Res. 2013 Nov;126(6):833-40. doi: 10.1007/s10265-013-0576-0. Epub 2013 Jul 16.
The Arabidopsis mitogen activated protein kinase kinase kinase (MEKK1) plays an important role in stress signaling. However, little is known about the upstream pathways of MEKK1. This report describes the regulation of MEKK1 activity during cold signaling. Immunoprecipitated MEKK1 from cold-treated Arabidopsis seedlings showed elevated kinase activity towards mitogen activated protein kinase kinase2 (MKK2), one of the candidate MEKK1 substrates. To clarify how MEKK1 becomes active in response to cold stress signaling, MEKK1 phosphorylation was monitored by an enzyme extracted from the seedlings grown under cold stress with or without EGTA. MEKK1 was phosphorylated after cold stress, but EGTA inhibited the phosphorylation. MKK2 was also phosphorylated by the same extract, but only when EGTA was absent. These results suggested that Ca(2+) signaling occurred upstream of the MEKK1-MKK2 pathway. Full-length MEKK1 showed almost no activity but MEKK1 without the N-terminal region (MEKK1 KD) that retained the kinase domain had a strong ability to phosphorylate MKK2, demonstrating the inhibitory role of the N-terminal region of MEKK1. In addition, MEKK1 was phosphorylated by calcium/calmodulin-regulated receptor-like kinase (CRLK1), which suggested that CRLK1 is one of candidates located upstream of MEKK1.
拟南芥丝裂原活化蛋白激酶激酶激酶（MEKK1）在应激信号转导中发挥着重要作用。然而，关于 MEKK1 的上游途径知之甚少。本报告描述了 MEKK1 活性在冷信号转导过程中的调节。从经过冷处理的拟南芥幼苗中免疫沉淀的 MEKK1 显示出对丝裂原活化蛋白激酶激酶 2（MKK2）的激酶活性升高，MKK2 是候选 MEKK1 底物之一。为了阐明 MEKK1 如何在响应冷应激信号时变得活跃，通过从在冷胁迫下生长的幼苗中提取的酶来监测 MEKK1 的磷酸化。MEKK1 在冷胁迫后被磷酸化，但 EGTA 抑制了磷酸化。同样的提取物也使 MKK2 磷酸化，但只有在没有 EGTA 的情况下才会发生。这些结果表明，Ca（2+）信号发生在 MEKK1-MKK2 途径的上游。全长 MEKK1 几乎没有活性，但保留激酶结构域的无 N 端区域（MEKK1 KD）的 MEKK1 具有强烈的磷酸化 MKK2 的能力，表明 MEKK1 的 N 端区域具有抑制作用。此外，MEKK1 被钙/钙调蛋白调节的类受体激酶（CRLK1）磷酸化，这表明 CRLK1 是位于 MEKK1 上游的候选物之一。
