Fig. 4: ROWY1 colocalized with PIN2 and regulated its subcellular distribution.

a, b ROWY1 and PIN2 colocalized in root epidermal cells. a Coexpression of PIN2-mCherry (left) and ROWY1-YFP (middle) in the primary root. Merged image of PIN2-mCherry and ROWY1-YFP (right). Track 1: laser wavelength, 594 nm, 40.00%; 0.0% offset; detector gain, 650 V. Track 2: laser wavelength, 488 nm, 20.00%; 0.0% offset; detector gain, 650 V. The white line indicates the plot area. Scale bar, 50 μm. b The gray value was measured to indicate the fluorescence intensity. Fluorescence plots and measurements were performed via ImageJ. c–e PIN2 and ROWYs play a role in the same signaling pathway. c Representative images showing comparisons of 7-day-old Col-0, rowy123, eir1-4, and eir1-4 rowy123 seedlings. Scale bar, 1 cm. d The angles of the primary root tips of 7-day-old seedlings were measured and are shown as a polar histogram. n values are indicated. e The primary root length of 7-day-old seedlings was measured. The data are presented as the means ± SDs; n values are indicated. The experiments were repeated three times independently, with similar results obtained. f–j Loss of function of ROWYs impaired the subcellular distribution of PIN2. f Representative images showing PIN2::PIN2-Venus expression in the epidermal cells of Col-0 and rowy1-1 rowy2-2 rowy3-2. Four-day-old seedlings were grown constantly on MS media; laser wavelength, 488 nm, 10.00%; 0.0% offset; detector gain, 650 V; scale bar, 50 μm. g The mean value was measured to indicate the expression of PIN2 in epidermal cells; n values are indicated. P values were calculated with a two-tailed t test. h Representative images showing the epidermal cells of PIN2::PIN2-Venus. i Representative images showing the epidermal cells of PIN2::PIN2-Venus;rowy123. Insets represent enlargements of the boxed regions from the corresponding images. Laser wavelength, 488 nm, 10.00%; 0.0% offset; detector gain, 650 V. Scale bar, 20 μm. j The mean value was measured to indicate the intracellular signal; n values are indicated. P values were calculated with a two-tailed t test. The experiments were repeated four times independently, with similar results obtained.