Fig. 6: DA1 is involved in cytokinin-induced accumulation of WUS protein and enlargement of the SAM.

a, b 6-BA treatment decreases the level of DA1 protein. a Total protein extracts were detected by anti-GFP and anti-ACTIN (loading control) antibodies b Quantification of GFP-DA1 protein levels in a. c, d Quantification of mRNA levels in 35 S:GFP-DA1 or Col-0 treated with or without 6-BA. e SAMs from pWUS:WUS-GFP transgenic plants (n = 14). f SAMs from pWUS:WUS-GFP transgenic plants treated with 6-BA (n = 13). g SAMs from pWUS:WUS-GFP; da1-1 transgenic plants(n = 9). h SAMs from pWUS:WUS-GFP; da1-1 transgenic plants treated with 6-BA (n = 13). i SAMs of 6-day-old Col-0 plants. j SAMs of 6-day-old Col-0 plants treated with 6-BA. k SAMs of 6-day-old da1-1 plants. l SAMs of 6-day-old da1-1 plants treated with 6-BA (n = 30). m The fluorescing cell number of SAMs in pWUS:WUS-GFP (n = 14), pWUS:WUS-GFP + 6-BA (n = 13), pWUS:WUS-GFP; da1-1 (n = 9), and pWUS:WUS-GFP; da1-1+6-BA (n = 13) plants. n Quantification of WUS-GFP mRNA levels. pWUS:WUS-GFP (C), pWUS:WUS-GFP+6-BA (C+6), pWUS:WUS-GFP; da1-1 (d) and pWUS:WUS-GFP; da1-1 + 6-BA (d+6). o The average area of SAMs of Col-0, Col-0 + 6-BA, da1-1 and da1-1 + 6-BA SAMs (n = 30). p A model for cytokinin-DA1-WUS module in controlling the shoot stem cell function and SAM size. DA1 represses WUS to regulate shoot stem cell function and SAM size, while Cytokinin represses DA1 level in the SAM. Cytokinin promotes the WUS accumulation partially through the repression of DA1. The da1-1 mutation caused the WUS accumulation, resulting in the large SAMs with increased stem cell population. Cytokinin also promotes WUS expression. Data in b, c, d and n are mean ± s.e.m with three biological repeats. P values are from two-sided Student’s t tests(b–d) and one-way ANOVA with Tukey’s multiple comparison test (n). Data in m and o are presented as mean values ± s.e.m relative to the mock value (100%). P values are from one-way ANOVA with Tukey’s multiple comparison test (P  <  0.05). GFP (green) fluorescence were shown. Scale bars, 20 μm (e–l). The experiments in e–m and o were repeated independently at least twice with similar results. Source data are provided as a Source Data file.