Fig 1.
High far-red (FR) light irradiation reduces sensitivity of Arabidopsis seedlings to methyl jasmonate (MeJA) and a negative feedback regulation of FIN219 levels in response to MeJA.
A. Hypocotyl response of wild-type Col, fin219-2, and FIN219 inducible transgenic line (PGR219) to different concentrations of MeJA under low FR light. Seedlings were grown under low FR light (2 μmol m-2 s-1) for 3 days with 0, 25, 50, 75 and 100 μM MeJA. The hypocotyl length in response to MeJA was converted to inhibition percentage of that without MeJA. n = 35. Different lowercase letters represent significant differences by Tukey’s studentized range test at P< 0.05. B. Hypocotyl response of wild-type Col, fin219-2, and PGR219 to different concentrations of MeJA under high FR light. Treatment was as in (A) except for high FR light (10 μmol m-2 s-1). Data are mean±SEM. n = 35. C. Gel blot analysis of FIN219 protein level in wild-type Col in response to various concentrations of MeJA under low and high FR light. Seedlings were grown under low and high FR light with different concentrations of MeJA for 3 days. Total protein, 70 μg, was loaded in each lane and probed with FIN219 monoclonal and JAZ1 polyclonal antibodies. Coomassie blue staining was a loading control. D. Gel blot analysis of FIN219 protein level in PGR219 in response to various concentrations of MeJA under low and high FR light conditions. Seedlings of the PGR219 transgenic line were used for detecting FIN219 levels as shown in (C).
Fig 2.
Microarray assay of genes with significant differences in transcript abundance between wild type and fin219-2 in response to FR light with and without MeJA.
Venn diagram of number of genes with significant difference (≥ 2-fold) in transcript abundance between fin219-2 and the wild type without (-MeJA) or with 50 μM MeJA (+MeJA) under low FR light (2 μmol m-2 s-1).
Fig 3.
Relative ratio of Gene Ontology (GO) groups among gene expression profiles affected by FIN219 levels under low FR light without or with MeJA treatment.
Relative ratio of GO groups among gene expression profiles in fin219-2 versus wild-type Col with at least 2-fold difference in expression under low FR light (2 μmol m-2 s-1) (A) and with 50 μM MeJA under low FR light (B). Relative ratio calculated as gene number in each GO group / total affected genes (2,249 in fin219/Col and 3,063 in fin219/Col with MeJA) * 100%.
Table 1.
Number and percentage of the components involved in multiple hormone biosynthesis and signaling pathways changed in loss-of-function (fin219-2) or gain-of-function (PGR219) FIN219/JAR1 in Arabidopsis seedlings with or without 50 μM methyl jasmonate (MeJA).
Data are derived from microarray assay shown in Fig 3. up, upregulated; down, downregulated.
Fig 4.
The fin219-2 mutant shows altered expression of a number of transcription factors (TFs) under low FR light with or without MeJA.
Hierarchical clustering analysis of the basic helix-loop-helix (bHLH) TFs in fin219-2 under low FR light without or with 50 μM MeJA. Seedlings of wild type and fin219-2 mutant were grown under low FR light without or with 50 μM MeJA for 3 days and subjected to microarray assay. Red asterisk marks the genes At2g40200 (bHLH51) and At4g29930 (bHLH27) that were downregulated in fin219-2 under both low FR light and MeJA.
Fig 5.
Validation of selected bHLH TFs in fin219-2 mutant.
(A)-(I) Quantitative real-time PCR analysis of bHLH TF expression in seedlings of wild type and fin219-2 grown without or with 50 μM MeJA under low FR light for 3 days. Data are mean±SEM from 3 biological replicates. Different lowercase letters represent significant differences by Tukey’s studentized range test at P< 0.05.
Fig 6.
Loss-of-function mutants of selected bHLH TFs show altered hypocotyl responses to exogenous MeJA.
A-E. Hypocotyl elongation of wild-type Col-0 (Col), fin219-1, fin219-2 and selected bHLH TF mutants without or with MeJA under FR light. Seedlings were grown in low FR light (2 μmol m-2 s-1) without MeJA (-MeJA) or with 50 μM MeJA (+MeJA) for 3 days. Scale bar is 5 mm. F-G. Quantitative analysis of hypocotyl lengths of Col, fin219-2 and selected bHLH TF mutants without MeJA (F) or with MeJA (G) under low FR light. Different lowercase letters represent significant differences by Tukey’s studentized range test at P< 0.05. H. Quantitative analysis of MeJA-mediated inhibition of hypocotyl elongation shown in (A-E). Data are mean±SEM from 3 biological replicates. Different lowercase letters represent significant differences by Tukey’s studentized range test at P< 0.05.
Fig 7.
A model to illustrate FIN219/JAR1 functions in the integration of FR light and JA signaling.
FR light perceived by the photoreceptor phyA regulates gene expression likely via FIN219/JAR1-mediated positive or negative modulation of TFs, including bHLH TFs. FIN219/JAR1 as a JA-conjugating enzyme is mainly responsible for the production of JA-Ile, an active form of JA hormone involved in regulating various physiological responses, and crosstalks positively and negatively with other hormones such as ABA, ethylene, auxin and JA, for regulation of hypocotyl elongation and root development. Regular arrow, positive effect; inverted T, negative effect.