Mitogen-activated protein kinase kinase 9 (MKK9) is an upstream activator of mitogen-activated protein kinase 3 (MAPK3) and MAPK6 in planta. To investigate MKK9 roles in mitochondrial respiration in Arabidopsis, MKK9DD, the active allele with mutations of Thr-201 and Ser-205 to Asp, and MKK9KR, the allele lacking MKK9 activity with a mutation of Lys-76 to Arg, were used. Results showed that the total respiratory rate (Vt), alternative pathway capacity (Valt) and cytochrome pathway capacity (Vcyt) increased under 0–100 mM NaCl treatments but decreased under 150–300 mM NaCl treatments in Col-0 callus. However, the activation of MKK9 by dexamethasone (DEX) increased Vt, Valt and Vcyt under 200 mM NaCl treatment; moreover, Valt showed more increase than Vcyt. The activation of MKK9 in MKK9DD callus sharply increased AOX protein expression under normal and NaCl conditions, but the increase was not observed in MKK9KR callus. Further results indicated that MAPK3 and MAPK6 were involved in the MKK9-induced increase of AOX protein levels. qRT-PCR results showed that MKK9-MAPK3/MAPK6 was involved in the NaCl-induced AOX1b and AOX1d expression, but only MKK9-MAPK3 was necessary for AOX2 expression; in addition, MAPK3 regulated the AOX1a transcription in an MKK9-independent manner. MKK9 positively regulated SOD and CAT activities by affecting MAPK3 and MAPK6 and negatively regulated APX and POD activities by affecting MAPK3. Moreover, MKK9 functions as a positive factor in H2O2 accumulation under salt stress. The regulation of ethylene on alternative respiration was also associated with MKK9 under salt stress. Taken together, the MKK9-MAPK3/MAPK6 pathway plays a pivotal role in increasing alternative respiration in the salt-treated Arabidopsis callus.

中文翻译：

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活性 MKK9-MAPK3/MAPK6 参与增加盐处理拟南芥愈伤组织的呼吸

丝裂原活化蛋白激酶激酶 9 (MKK9) 是植物中丝裂原活化蛋白激酶 3 (MAPK3) 和 MAPK6 的上游激活剂。为了研究 MKK9 在拟南芥线粒体呼吸中的作用，使用了 MKK9DD（具有 Thr-201 和 Ser-205 突变为 Asp 的活性等位基因）和 MKK9KR（缺乏 MKK9 活性且具有 Lys-76 突变为 Arg 的等位基因）。结果表明，在 Col-0 愈伤组织中，总呼吸频率 (Vt)、旁路容量 (Valt) 和细胞色素通路容量 (Vcyt) 在 0-100 mM NaCl 处理下增加，但在 150-300 mM NaCl 处理下降低。然而，地塞米松 (DEX) 对 MKK9 的激活在 200 mM NaCl 处理下增加了 Vt、Valt 和 Vcyt；此外，Valt 显示出比 Vcyt 更多的增加。在正常和NaCl条件下，MKK9DD愈伤组织中MKK9的活化使AOX蛋白表达急剧增加，但在MKK9KR愈伤组织中未观察到增加。进一步的结果表明，MAPK3 和 MAPK6 参与了 MKK9 诱导的 AOX 蛋白水平的增加。qRT-PCR结果显示MKK9-MAPK3/MAPK6参与了NaCl诱导的AOX1b和AOX1d的表达，但只有MKK9-MAPK3是AOX2表达所必需的；此外，MAPK3 以独立于 MKK9 的方式调节 AOX1a 转录。MKK9 通过影响 MAPK3 和 MAPK6 正向调节 SOD 和 CAT 活性，通过影响 MAPK3 负向调节 APX 和 POD 活性。此外，MKK9 在盐胁迫下作为 H2O2 积累的积极因素。乙烯对替代呼吸的调节也与盐胁迫下的 MKK9 相关。