Arabidopsis NAC transcription factor JUNGBRUNNEN1 affects thermomemory-associated genes and enhances heat stress tolerance in primed and unprimed conditions

Abstract

We recently reported that the NAC transcription factor JUNGBRUNNEN1 (JUB1; ANAC042) extends longevity and increases tolerance to heat stress in Arabidopsis thaliana when overexpressed, while the opposite is observed in jub1-1 knock-down lines. Here we extend our previous findings by demonstrating that JUB1 also positively regulates plant survival under heat stress when plants were treated by a prior moderate (and non-lethal) temperature regime (so-called priming). We further find that JUB1 shows thermomemory-related expression, similar to two other genes previously reported to be important for thermopriming, i.e., HSFA2, encoding a heat shock factor, and HSA32, encoding a heat shock protein. Our analysis also identifies ASCORBATE PEROXIDASE2 (APX2) and the heat shock protein genes HSP18.2 and HSP21 as thermomemory-expressed genes, revealing them as new candidates for studies to decode the molecular processes controlling thermopriming.

Keywords: ANAC042; Abiotic stress; JUB1; heat stress tolerance; hydrogen peroxide; molecular memory; reactive oxygen species; thermopriming.

Figure 1. Plant survival under unprimed and primed conditions. (A) Temperature profile. The priming stimulus was given at 37°C for 1.5 h. The triggering heat stimulus at 45°C was applied for 45 min two days after the priming stimulus. In between, plants were kept at 23°C (recovery or memory phase). Note, that the priming stimulus was omitted for unprimed seedlings. (B) Seedlings of jub1-1, wild-type (WT) and JUB1ox lines photographed at day 7 after the triggering heat stimulus. (C) Percentage of seedlings that survived the triggering heat stimulus in unprimed and primed conditions (7 d after the triggering stimulus). Note, that unprimed and primed JUB1ox seedlings survived considerably better than the jub1-1 and WT seedlings. Importantly, unprimed JUB1ox seedlings had a better survival rate than primed jub1-1 seedlings (p < 0.01; Student’s t-test), indicating that JUB1ox seedlings are already primed even in the absence of stress. Notably, however, even in these plants, priming further enhanced the heat stress tolerance. Mean of three experiments ± SE is given. Pair-wise comparisons between unprimed and primed samples (a vs. b, c vs. d, and e vs. f), between unprimed samples of different genotypes (a vs. c, and c vs. e), and between primed samples of different genotypes (b vs. d, and d vs. f) revealed that differences are statistically significant (p < 0.005; Student’s t-test).

Figure 2. Expression analysis. (A) Heatmap representation of the expression of selected heat stress-associated genes during the thermomemory phase. Gene expression analysis was performed using qRT-PCR. Expression values obtained for wild-type (WT) seedlings at 2 h, 4 h and 28 h after thermopriming at 37°C were compared with those of unprimed control plants harvested at identical time points. Blue shades indicate expression increase (log2-fold basis) relative to unprimed samples. The experiment was performed in three replicates. (B) Expression of heat memory-associated genes in 5-d-old seedlings of jub1-1, WT and JUB1ox lines at unprimed condition. The Y axis indicates log2 fold change expression compared with WT. Note the enhanced expression of HSFA2, HSP21 and HSP18.2 in the JUB1ox line compared with WT and jub1-1 seedlings. (C) Gene expression in 5-d-old WT and jub1-1 seedlings 4 h after the priming heat stimulus (37°C). Data in (B) and (C) are means of two biological replications. AGI codes: JUB1 (At2g43000); HSFA2 (At2g26150); APX2 (At3g09640); HSA32 (At4g21320); HSP21 (At4g27670); HSP17.6II (At5g12020); HSP17.6A (At5g12030); HSP15.7-CI (At5g37670); HSP18.2 (At5g59720); HSP26.5-P (At1g52560).