Sumoylation of transcription factor MYB30 by the small ubiquitin-like modifier E3 ligase SIZ1 mediates abscisic acid response in Arabidopsis thaliana

Abstract

The phytohormone abscisic acid (ABA) plays an essential role in plant development and during the response of the plant to abiotic stress. In this study, we report that the R2R3-type transcription factor MYB30 is involved in the regulation of ABA signaling. Arabidopsis mutants lacking MYB30 are hypersensitive to ABA during germination and seedling growth. A K283R substitution in MYB30 blocks its SUMO E3 ligase SIZ1-mediated sumoylation in Arabidopsis protoplasts, indicating that MYB30 is sumoylated by SIZ1 and that K283 is the principal site for small ubiquitin-like modifier conjugation. Expression of MYB30 K283R in myb30 partially rescues the mutant ABA-hypersensitive phenotype, but expression of wild-type MYB30 complements the mutant phenotype. Overexpression of MYB30 in wild-type results in an ABA-insensitive phenotype, whereas overexpression of MYB30 in the siz1 mutant does not alter siz1 hypersensitivity to ABA. The siz1-2 myb30-2 double-mutant exhibits greater ABA sensitivity than either single mutant, but a mutation in the SIZ1-sumoylated ABI5 transcription factor suppresses the ABA hypersensitivity of myb30-2 to wild-type levels. Our results suggest that coordination of ABI5 and MYB30 sumoylation by SIZ1 may balance gene expression, which is required for regulation of ABA signaling during seed germination.A bscisic acid (ABA) plays important roles throughout the life cycle of the plant, including during seed development and dormancy, seed germination, early seedling development, flowering, and in response to abiotic and biotic stress. Genetic studies have revealed that a number of loci are essential for ABA biosynthesis (1-5), catabolism (6, 7), and signal transduction (8, 9) in plants.A number of studies have focused on understanding ABA signaling from perception to response. Recently, three groups of ABA receptors (ABAR/GUN5, GRGC/GTG, and PYR/PYL/ RCAR) have been identified in Arabidopsis and shown to bind ABA with high affinity (10-14). The various functions and cellular locations of these ABA receptors point to the complexity of ABA signaling. For example, PYR/PYL/RCAR-modulated ABA signaling has been studied extensively and found to involve a double-negative regulatory system that includes the receptor, protein phosphatases (PP2C), and protein kinases (SnRK2) (15, 16).A group of basic leucine zipper transcription factors, including ABI5, serve as the downstream targets of SnRK2 and phosphorylation is correlated with ABI5 transcriptional activity and protein stability (17-19). ABI5 undergoes 26S proteasome-mediated degradation and ABA stabilizes the ABI5 protein (17). Two ABI5 interacting proteins, AFP (ABI5-interacting protein) and KEG (KEEP ON GOING, a RING-finger ubiquitin E3 ligase) are required for ABI5 degradation (20,21). Arabidopsis lacking either gene is hypersensitive to ABA and an abi5 mutation suppresses the ABA sensitivity of both the afp-1 and keg mutants (20,21). Recently it has been shown that ABI5 is sumoylated by the small ubiquitin-like modifier (SUMO) E3 ligase SIZ1 at amino acid ...

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