Differential regulation of jasmonate responses in multiple jaz mutants.

The phytohormones jasmonates (JAs) regulate diverse aspects of plant growth and defense responses. The JA-ZIM domain (JAZ) family of repressors are targeted by the JA receptor Coronatine Insensitive 1 for ubiquitination and subsequent degradation via the 26S proteasome. We previously investigated the functions of JAZs in JA responses by analyzing jaz mutants of the phylogenetic group I (jaz1/2/5/6), group II/III (jaz10/11/12), group IV/V (jaz3/4/7/9 and jaz3/4/7/8/9), and their high-order mutant jaz1/2/3/4/5/6/7/9/10/11/12. Here, we examined JA-regulated root growth, apical hook curvature, flowering time, and defense against the insect Spodoptera exigua in the intermediate jaz mutants jaz1/2/5/6/10/11/12, jaz1/2/3/4/5/6/7/9, and jaz3/4/7/8/9/10/11/12. This study shows that these jaz mutants differentially affect JA responses, suggesting the complexity of JA pathway in these multiple jaz mutants.

The phytohormones jasmonates (JAs) control plant growth and development, such as root growth, apical hook curvature, flowering, and stamen development, and affect plant resistance and defense responses against insect attacks.[1-5] The JA-ZIM (JAZ) domain proteins function as core repressors of JA signaling and responses.[6-8] In response to JA signals, the JAZ repressors are recruited and ubiquitinated by the JA receptor Coronatine Insensitive 1 (COI1), and are subsequently degraded by the 26S proteasome, leading to activation of JAZs-repressed transcriptional responses and JA responses.[6,7,9,10]

The thirteen Arabidopsis JAZ repressors are classed into phylogenetic group I (JAZ1/2/5/6), II (JAZ10), III (JAZ11/12), IV (JAZ3/4/9), and V (JAZ7/8/13).[6,7,11 –13] The jaz1–1/2-2/5-1/6-5 mutant of group I is partially insensitive to JA in root growth inhibition.[13] The jaz10–1/11–1/12–1 mutant of group II/III is hypersensitive to JA in root growth inhibition.[13] jaz1/2/5/6 and jaz10/11/12 are more susceptible to the Spodoptera exigua larvae than wild-type.[13] The jaz3-5/4-1/7-1/9-1 and jaz3-5/4-1/7-1/8-1/9-1 mutants of group IV/V flower later than wild-type.[13] The high-order jaz mutants targeting most of JAZs, including jaz1-1/2-2/3-5/4-1/5-1/6-5/7-1/9-1/10–1/11–1/12–1,[13] jaz1-2/2–3/3-4/4-1/5–1/6–4/7–1/9–4/10–1/13–1 (jazD),[12] and jaz1-2/2–3/3–4/4–1/5– 1/6–4/7–1/8–V/9–4/10–1/13–1 (jazU),[12] are hypersensitive to JA-inhibitory root growth and apical hook curvature, hyposensitive to ethylene (ET)-enhanced apical hook curvature, more resistant to insect herbivores, and flower later than wild-type.[12,13]

To further understand JAZ family in JA responses, we performed genetic crosses with our previously characterized jaz mutants,[13] and constructed intermediate jaz mutants, including jaz1-1/2-2/5-1/6-5/10–1/11–1/12–1 of group I/II/III, jaz1-1/2-2/3-5/4-1/5-1/6-5/7-1/9-1 targeting group I/IV/V(JAZ7), jaz3-5/4-1/7-1/8-1/9-1/10-1/11-1/12-1 for group II/III/IV/V(JAZ7/8), and analyzed their JA responses.

The primary root growth of wild-type was inhibited by methyl-jasmonate (MeJA), and the undecuple jaz1/2/3/4/5/6/7/9/10/11/12 mutant control exhibited shorter roots under either mock or MeJA treatment (Figure 1a).[13] The primary root growth of jaz1/2/5/6/10/11/12 and jaz1/2/3/4/5/6/7/9 was similar to that of wild-type, while the root of jaz3/4/7/8/9/10/11/12 was shorter than that of jaz1/2/3/4/5/6/7/9/10/11/12 under mock or MeJA treatment (Figure 1a).

Figure 1.

JA-regulated root growth and apical hook curvature in the multiple jaz mutants. (a) Primary root length of 11-day-old seedlings of the Col-0 wild-type, jaz1/2/5/6/10/11/12, jaz1/2/3/4/5/6/7/9, jaz3/4/7/8/9/10/11/12, and the jaz1/2/3/4/5/6/7/9/10/11/12 control grown on MS medium containing 0, 5 or 20 μM MeJA under a 16-h (20–22°C)/8-h (18–20°C) light/dark photoperiod. Error bars represent SE (n = 20). (b, c) Apical hook curvatures (b) and hook phenotypes (c) of 4-day-old dark grown etiolated seedlings of the Col-0 wild-type, jaz1/2/5/6/10/11/12, jaz1/2/3/4/5/6/7/9, jaz3/4/7/8/9/10/11/12, and the jaz1/2/3/4/5/6/7/9/10/11/12 control on MS medium supplied with mock, 5 μM MeJA, 10 μM ACC (1-aminocyclopropane-1-carboxylic acid, the ethylene biosynthesis precursor), or 5 μM MeJA plus 10 μM ACC. Data are means ±SE (n = 15). Letters indicate significant differences by Tukey’s HSD test (P < .05).

JA represses the apical hook curvature of etiolated Arabidopsis wild-type seedlings, while ET functions oppositely and antagonizes the JA action.[14] The jaz1/2/3/4/5/6/7/9/10/11/12 control displayed reduced hook curvature, was hypersensitive to JA-inhibited hook curvature and partially insensitive to ACC (ET precursor)-strengthened hook curvature (Figures 1b,Figures 1c).[13] The jaz1/2/5/6/10/11/12, jaz1/2/3/4/5/6/7/9 and jaz3/4/7/8/9/10/11/12 mutants were indistinguishable from wild-type in JA/ET-regulated apical hook curvature (Figures 1b,Figures 1c), indicating the high redundancy of JAZ members in JA/ET-regulated hook curvature.

jaz1/2/3/4/5/6/7/9/10/11/12 flowers later than wild-type, while coi1-1 flowers earlier (Figures 2a,Figures 2b).[13] Considering rosette leaf numbers at flowering or days to flowering (the appearance of a 1 cm-long bolt[15]), jaz1/2/5/6/10/11/12 flowered earlier than wild-type (Figures 2a,Figures 2b). On the other hand, jaz1/2/3/4/5/6/7/9 and jaz3/4/7/8/9/10/11/12 flowered later than wild-type, but earlier than jaz1/2/3/4/5/6/7/9/10/11/12 (Figures 2a,Figures 2b). jaz1/2/3/4/5/6/7/9 and jaz3/4/7/8/9/10/11/12 suppressed the early flowering phenotype of coi1-1, to the degree of coi1-1 jaz1/2/3/4/5/6/7/9/10/11/12, while jaz1/2/5/6/10/11/12 did not obviously affect the flowering time of coi1-1 (Figures 2c,Figures 2d).

Figure 2.

Flowering time in the multiple jaz mutants. Rosette leaf number at flowering (a, c) and days to flowering (b, d) of the Col-0 wild-type, jaz1/2/3/4/5/6/7/9, jaz3/4/7/8/9/10/11/12, and the jaz1/2/3/4/5/6/7/9/10/11/12 control (a, b), or Col-0, coi1-1, and the indicated high-order jaz mutants in coi1-1 background (c, d) grown under a 16-h (20–22°C)/8-h (18–20°C) light/dark photoperiod. Data are means ±SE (n = 36). Letters indicate significant differences by one-way ANOVA analysis with Tukey’s HSD post hoc test (P < .05).

jaz1/2/3/4/5/6/7/9/10/11/12 is more resistant than wild-type in defense against the pest Spodoptera exigua (Figures 3a,Figures 3b).[13]The S. exigua larvae fed with jaz1/2/3/4/5/6/7/9 and jaz3/4/7/8/9/10/11/12 gained less weight than those with wild-type, but more weight than those with jaz1/2/3/4/5/6/7/9/10/11/12 (Figures 3a,Figures 3b). On the other hand, S. exigua larvae reared with jaz1/2/5/6/10/11/12 gained more weight than those with wild-type (Figures 3a,Figures 3b), suggesting that jaz1/2/5/6/10/11/12 is more susceptible to S. exigua than wild-type.

Figure 3.

Multiple JAZs mutations differentially affect defense against insect attack. (a) and (b) Larval weights (a) and representative S. exigua larvae (b) after feeding for 14 days (a) or 7 days (b) on Col-0, jaz1/2/3/4/5/6/7/9, jaz3/4/7/8/9/10/11/12, and the jaz1/2/3/4/5/6/7/9/10/11/12 control. 7-day-old seedlings grown under a 16-h (20–22°C)/8-h (18–20°C) light/dark photoperiod were transplanted into soil and grown under a 10-h (20–22°C)/14-h (18–20°C) light/dark photoperiod. The newly hatched S. exigua larvae were reared on 6-week-old plants. Error bars represent SE (n = 15). Scale bar = 0.2 cm. Letters indicate significant differences by one-way ANOVA analysis with Tukey’s HSD post hoc test (P < .05).

In conclusion, these results showed that the three high-order jaz mutants exhibited different effects on JA responses, including root growth, apical hook curvature, flowering time, and defense against insects, and indicate the redundancy of JAZ members and complexity of JA pathway in high-order jaz mutants.