Fig. 4: Thermosensitive ARF accumulation alters ARF nuclear accumulation.

a Graphic of typical temperature-dependent phase diagram for upper critical saturation temperature (UCST) behavior. C₁ and C₂ correspond to protein critical saturation concentrations. In this example, the saturation concentration of the protein is higher with higher temperature (ie, the protein is more soluble at elevated temperature in UCST systems). b Confocal microscopy images of 4-day-old root tips of seedlings expressing ARF7:ARF7-mVenus after a 4-hour incubation at 22 °C or 37 °C. To the right are closeup views of nuclear signal. c Mean (±SD) fluorescence intensity of root tip nuclei in roots of seedlings pARF7: ARF7-mVenus¹¹ after 4 h incubation at 22 °C and 37 °C (n = 148 nuclei at 22 °C and 143 nuclei 37 °C). ****P = 3.02E-13 using two-tailed unpaired t-test. Arbitrary units are used. d Mean (±SD) fluorescence intensity of root tip nuclei of of 4-day-old seedlings expressing pUBQ10:YFP-ARF19 after 4 h incubation at 22 °C or 28 °C (n = 24 nuclei at 22 °C and 40 nuclei 37 °C). ****P = 8.62E-10 using two-tailed unpaired t-test. Arbitrary units are used. e Time course fluorescence microscopy images of hypocotyl cells of 4-day-old seedlings expressing pUBQ10:YFP-ARF19. Seedlings were mounted on a heated stage at 22 °C that rapidly transitioned to 32 °C. Images were taken at the indicated time points and, in one example cell, the nucleus pointed out with a puce carat and condensates pointed out with orange carats. f Mean (±SD) fluorescence intensity in nuclei at the start of treatment and after 31 min of treatment. (n = 88 nuclei at 0 min and 84 nuclei at 31 min from 5 individual seedlings). ****P = 2.51E-18 using two-tailed unpaired t-test. Arbitrary units are used. g Graphic demonstrating effectors and reporter in the FrankenARF7 system²². h Box plots of median and interquartile range of mScarlet signal found in mNeon positive cells; whiskers represent maximum and minimum values (n = 1340 for the control at 22 °C, 3554 for the control at 32 °C, 2210 for FrARF7 at 22 °C, and 1736 for FrARF7 at 32 °C). P = 1.05E-06 for the FrARF effector and P = 6.37E-10 for the FrARF reporter using two-tailed unpaired t-test. Box plots of median and interquartile range of mNeon signal in transformed protoplasts; whiskers represent maximum and minimum values.