Salicylate accumulation inhibits growth at chilling temperature in Arabidopsis

Abstract

The growth of Arabidopsis plants in chilling conditions could be related to their levels of salicylic acid (SA). Plants with the SA hydroxylase NahG transgene grew at similar rates to Col-0 wild types at 23 degrees C, and growth of both genotypes was slowed by transfer to 5 degrees C. However, at 5 degrees C, NahG plants displayed relative growth rates about one-third greater than Col-0, so that by 2 months NahG plants were typically 2.7-fold larger. This resulted primarily from greater cell expansion in NahG rosette leaves. Specific leaf areas and leaf area ratios remained similar in both genotypes. Net assimilation rates were similar in both genotypes at 23 degrees C, but higher in NahG at 5 degrees C. Chlorophyll fluorescence measurements revealed no PSII photodamage in chilled leaves of either genotype. Col-0 shoots at 5 degrees C accumulated SA, particularly in glucosylated form. SA in NahG shoots showed similar tendencies at 5 degrees C, but at greatly depleted levels. Catechol was not detected as a metabolite of the NahG transgene product. We also examined growth and SA levels in SA signaling and metabolism mutants at 5 degrees C. The partially SA-insensitive npr1 mutant displayed growth intermediate between NahG and Col-0, while the SA-deficient eds5 mutant behaved like NahG. In contrast, the cpr1 mutant at 5 degrees C accumulated very high levels of SA and its growth was much more inhibited than wild type. At both temperatures, cpr1 was the only SA-responsive genotype in which oxidative damage (measured as thiobarbituric acid-reactive substances) was significantly different from wild type.

Figure 1.

Temperature effects on Col-0 and NahG growth. A, Mean whole-plant biomass during incubation at 5°C or during the same period at 23°C (n = 4). B, Phenotypes of plants after 49 d at 5°C.

Figure 2.

Leaf growth in Col-0 and NahG at low temperature. A, Mean total leaf areas of rosettes. B, Mean numbers of rosette leaves per plant during incubation at 5°C (n = 4). C, Mean areas of five individual fully expanded rosette leaves from plants kept 76 d at 5°C. D, From each of these leaves, cross-sectional areas of 10 epidermal cells were averaged and overall means (n = 5) are shown. E, Mean numbers of cells per leaf were thereby calculated. Bars are se.

Figure 3.

LARs during growth of Col-0 and NahG at low temperature, calculated as plant leaf area ÷ plant biomass. Values are means of four replicates.

Figure 4.

Net assimilation rates and PSII efficiency in Col-0 and NahG plants. A, NARs during 6–21 d at 23°C, or 8–35 d at 5°C. Each value is a mean of RGR ÷ LAR at five time points in each period. B, Mean F_v/F_m ratios of mature rosette leaves from plants (n = 10) kept 12 d at 23°C or 42 d at 5°C. Bars are se.

Figure 5.

Salicylate accumulation in Col-0 and NahG shoots at low temperature. A, Equal ages. Mean free and glucosyl SA per g shoot fresh weight in plants kept 12 d at 23°C or 5°C. Triplicates of 36 (5°C) or 2 (23°C) shoots (approximately 0.45 g) were extracted. B, Equal stages. Mean free and glucosyl SA per g shoot fresh weight in plants grown to stage 1.08 during 5 d at 23°C or 20 d at 5°C. Six replicates of 18 shoots (approximately 0.45 g) were extracted. Bars are se.

Figure 6.

Low-temperature growth of genotypes in relation to SA levels. A, Shoot phenotypes after 36 d at 5°C. Bar = 1 cm. B, Mean shoot biomass after 14 d at 23°C or 52 d at 5°C (n = 8). Bars are se. C, Mean free and glucosyl SA per g shoot fresh weight in plants kept 42 d at 5°C. Four replicates of six shoots were extracted. Bars are se.

Figure 7.

Oxidative damage in genotypes at low temperature. Mean MDA yield from TBARS assays of plants kept 8 d at 23°C or 36 d at 5°C. Six replicates of three shoots were assayed. Bars are se.