Arabidopsis Putative MAP Kinase Kinase Kinases Raf10 and Raf11 are Positive Regulators of Seed Dormancy and ABA Response

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Abstract

We isolated an Arabidopsis ABA-insensitive mutant, ais143, by activation tagging screen. T-DNA was found to be located in the coding region of a putative mitogen-activated protein (MAP) kinase kinase kinase (MAP3K) gene, Raf10, thereby abolishing its expression in the mutant. ais143 exhibited reduced seed dormancy as well as reduced ABA sensitivity. The phenotypes were complemented by the wild-type Raf10 gene, and the overexpression (OX) of Raf10 resulted in delayed seed germination and enhanced ABA sensitivity. Raf10 has high sequence identity to another MAP3K, Raf11. Parallel analysis of Raf11 knockout (KO) and OX lines showed that their phenotypes were similar to those of Raf10 KO and OX lines. An ais143 raf11 double mutant exhibited stronger phenotypes than single mutants, indicating the functional redundancy between Raf10 and Raf11. Transcript analysis revealed that the expression of many ABA-associated genes, including the key regulatory genes ABI3 and ABI5, was altered in the Raf10 and Raf11 OX lines. Recombinant Raf10 and Raf11 proteins exhibited kinase activity, which was inhibited by the MAP3K inhibitor BAY 43-9006 but not by the MAP2K inhibitor U0126. Collectively, our data indicate that Raf10 and Raf11 kinases are important regulators of seed dormancy and ABA response and that they affect the expression of ABI3, ABI5 and other ABA-regulated genes.

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