Heat promotes hypocotyl elongation through auxin-mediated transcriptional modulation of SlUVI4 and SlCCS52B.
We increased incubation temperature during the emergence of tomato seedlings, and observed that 33°C treatment significantly promoted hypocotyl elongation of cultivar ‘Heinz 1706’ by 1.6- and 1.3-fold at NMS and MS growth medium compared to the normal growth temperature 25°C, but did not affect the hypocotyl thickness (Figure 6A, B, C). Cell cycle analysis revealed that 33°C treatment improved endoreduplication in hypocotyl cells compared to 25°C resulting in increases of EI values from 0.85 and 0.94 at 25°C to 0.99 and 1.00 at 33°C on NMS and MS growth medium, respectively (Figure 6D). Heat induced promotion of endoreduplication was more pronounced at NMS medium than MS medium, which was in accord with the rate of improvement on hypocotyl elongation. We also measured hypocotyl growth and cell cycle progression of tomato cultivars ‘Moneymaker’ and ‘Alisa Craig’, and observed the same phenomena on hypocotyl elongation and endoreduplication after high temperature treatment except a slight decrease of hypocotyl diameter of ‘Alisa Craig’ grown on MS medium at 33°C comparted to that at 25°C (Figure S7). qRT-PCR analysis revealed that 33°C treatment significantly inhibited the transcription ofSlUVI4 , but did not affect SlCCS52B transcription. The transcription of SlCCS52A1 was not affected by 33°C treatment, while the transcript abundance of SlCCS52A2 had a slight increase suggesting a weak role in the regulation of endoreduplication (Figure 6E).
To investigate functions of plant hormones in heat treatment, we firstly applied different concentrations of auxin (IAA), brassinosteroids (BRs), gibberellins (GAs) and ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) in growth medium to analyze hypocotyl growth. We observed slight enhancement of hypocotyl elongation responding to low concentration of IAA (Figure 7A-C; Figure S8A, B). BR and GA did not have consistent effects on hypocotyl growth (Figure S8C-F). Higher concentration of ACC had more significant effects on hypocotyl elongation and thickening (Figure S8G, H). Cell cycle analysis in hypocotyls revealed no significant difference between the control and 0.1nM IAA treatment (Figure 7D). qRT-PCR analysis showed thatSlUVI4 transcriptions were significantly reduced, andSlCCS52B transcription had a slight reduction without statistical significance (Figure 7E), suggesting that reduced SlUVI4transcription and steady SlCCS52B transcription might be required to maintain endoreduplication in auxin induced hypocotyl elongation. Both SlCCS52A1 and SlCCS52A2 transcription were significantly reduced suggesting that they might not be as important asSlCCS52B in hypocotyl elongation. All the data above suggested that elevated temperature promoted hypocotyl elongation possibly due to auxin-mediated repression of SlUVI4 transcription and maintenance of SlCCS52B transcription.