Molecular Basis Of Jasmonate Action In Arabidopsis

Jasmonates (JAs) are a new class of plant hormones, which synthesize fromlinolenic acid via an octadecanoid pathway in planta. They play important role inplant growth and development as well as in plant defense. Jasmonates regulate theexpression of enormous genes that are involved in response to UV irradiation,wounding, insect attack and pathogen infection. Although the'effect of JA on plantis well-characterized, little is known about the mechanism whereby jasmonatesexert their biological functions. This study shed a little light on the understandingof the jasmonate signaling pathway, and the major results are shown as following fl. The SCFco,' ubiquitin-ligase complexes are required fOr jasmonateresponse in Arabidopsis(l) Arabidopsis COI is required for all the jasmonate-regulated response. Itencodes a protein containing leucine-rich repeats and a degenerate F-box motifThese structural features are characteristic of F-box proteins that function inubiquitin ligase complexes for the ubiquitylation of substrate-proteins targeted fordegradat1on. This study demonstrated that the Arabidopsis F-box protein COIlassociated with AtCUL1, AtRbxl and Skpl-like proteins ASKl and ASK2 toassemble SCFcoIl ubiquitin ligase complexes. Also, we found that the AtCULlcomponent of SCFcoIl complexes contained two species including AtCULl andmodified AtCULl.(2) We found that COIl assembled to two separate SCFcoIl complexes with eitherASKl or ASK2 through immunoprecipitation analysis with plant expressing Myc-tagged version of ASK2.(3) We further demonstrated that the AtCULl component of SCFco11 complexeswas modified via AXR1. Mutations AXR1 resulted in reduction of the modifiedAtCULl component of SCFcol1 complex.(4) The mutant alleles of AXRi (axr]-3 and axr]-12) showed moderate insensitivity -to JA-inhibitory root growth and mild reduction of JA-inducible gene expression ofAtVSP, Thi2.i and PDFi.2 compared with that of wild type. The evidence thatmutations in AXR1 reduced the abundance of modified AtCULl subunit of SCFcoIland lead to a reduction of JA response suggested that the AXR1-dependent RUBlmodification to AtCULl component of SCFco,l was important for JA signaling.(5) The genetics analysis between AXR1 and COI1 gene demonstrated that axr1 andcoiI doub1e mutations displayed a synergistic genetic interaction. The axrI-3/coi1-2 double mutants were completely male sterile, resistant to JA-inhibitory rootgrowth, and lacked the expression of JA-inducible genes. Whereas their parentmutant axr1-3 was male fertile, showed moderate insensitivity to JA-inhibitory rootgrowth and mild reduction of JA-inducible gene expression; parent mutant coi1-2 -was a leaky mutant allele of COI!, it exhibited reduced JA insensitivity and partialfertility. -Taken together, all these results suggested that SCFcoIl complex was requiredfor the JA response and that the AXR1-dependent RUBl modification of AtCULlsubunit of SCFcol' was important for JA signaling in Arabidopsis.2. Idcntification of an Arabidopsis mutaut cex1 exhibits constantaccumulation ofjasmonatc-regulated AtVSP Thi2.1 and PDF1.2.From 80,000 Arabidopsis M2 secdlings, two mutants cex1-1 and cex1(2 wereidentified. When grown on MS medium they exhibited stunt groWth phenotypesimilar to the wild type plants grown on the medium with methyl jasmonate.Northern blot analysis showed that the two mutants displayed a constant expressionof JA{inducible gene AtVSP Thi2. 1 and PDF].2. Genetic analysis of progeny fromthe crosses of WTxcex1-1 and WTxcex1-2 demonstrated that the phenotypes ofcex1-2 mutants were inherited as a single recessive Mendelian marker. Geneticanalysis of progeny from cross cex1-1 xcexI-2 showed that cex1-1 and cex!-2 wereallelic at the cex1 locus.