Fig. 3: Thermostability is encoded in the ARF DBD and MR. | Nature Communications

Fig. 3: Thermostability is encoded in the ARF DBD and MR.

From: AUXIN RESPONSE FACTOR thermostability

Fig. 3: Thermostability is encoded in the ARF DBD and MR.The alternative text for this image may have been generated using AI.

a Graphic demonstrating examined ARF19 protein truncations. b Immunoblot analysis with anti-YFP antibody of seedlings carrying the indicated full length (FL) or ARF19 truncation incubated for 4 hours at the 22 °C or 37 °C. Two independent transformants shown for each truncation. Asterisk indicates a non-specific band, which displays decreased accumulation at elevated temperature. c Graphic depicting in vivo mutagenesis screen. d Alignment of ARF7 and ARF19 homologs with sites of variation identified. e Model of ARF19 aligned to ARF5 DBD (4LDU51) with ARF19W330L and ARF19R297W variant effects on contact sites. f Mean ( + SE) ratiometric signal of protoplasts expressing ARF19, ARF19W330L, or ARF19R297W ratiometric reporters (n = 5677 for ARF19 at 22 °C, 5904 for ARF19 at 32 °C, 11513 for ARF19W330L at 22 °C, 7373 for ARF19W330L at 32 °C, 7079 for ARF19R297W at 22 °C, 6649 for ARF19R297W at 32 °C, 7877 for empty vector at 22 °C, and 5197 for empty vector at 32 °C,), letters indicate a significance of p < 0.0001, 2-way ANOVA with Tukey’s HSD test. g Immunoblot analysis with anti-GFP antibody of 5-day-old arf7 arf19 seedlings carrying UBQ10:YFP-ARF19, UBQ10:YFP-ARF19W330L, or UBQ10:YFP-ARF19R297W incubated for 4 hours at 22 °C or 32 °C. h Mean (±SE) hypocotyl lengths of 5-day-old seedlings of wildtype (Col-0), arf7 arf19, and arf7 arf19 carrying UBQ10:YFP-ARF19, UBQ10:YFP-ARF19W330L, or UBQ10:YFP-ARF19R297W grown at 22 °C or 32 °C (n = 58 for wild type at 22 °C, 59 for wild type at 32 °C, 107 for arf7 arf19at 22 °C, 109 for arf7 arf19 at 32 °C, 212 for arf7 arf19 carrying UBQ10:YFP-ARF19 at 22 °C, 107 for arf7 arf19 carrying UBQ10:YFP-ARF19 at 32 °C, 134 for arf7 arf19 carrying UBQ10:YFP-ARF19W330L at 22 °C, 212 for arf7 arf19 carrying UBQ10:YFP-ARF19W330L at 32 °C, 118 for arf7 arf19 carrying UBQ10:YFP-ARF19R297W at 22 °C, and 93 for arf7 arf19 carrying UBQ10:YFP-ARF19R297W at 32 °C). Groups not sharing a letter are significantly different from one another (Two-factor ANOVA on the log transformed data; Tukey’s multiple comparison tests, p < 0.05). i Mean (±SE) increase in hypocotyl length of 5-day-old seedlings of wildtype (Col-0), arf7 arf19, and arf7 arf19 carrying UBQ10:YFP-ARF19, UBQ10:YFP-ARF19W330L, or UBQ10:YFP-ARF19R297W grown at 22 °C or 32 °C, calculated from the data in (h). Groups not sharing a letter are significantly different from one another (One-factor ANOVA on the log-transformed data, Tukey’s multiple comparison tests, p < 0.05).

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