PMC

Cytokinin promotes an increase in meristem size in a CLV3-dependent manner. (A–D) Scanning electron images of wild type mock-treated (A), wild-type cytokinin-treated (B), clv3-2 mutant mock-treated (C), and clv3-2 mutant cytokinin-treated (D) SAMs. (E) Schematic of rates of proliferation along the 1D column of cells for wild type. (F) Simulated proliferation rates along the 1D column of cells for wild type (blue), wild-type cytokinin-treated (green), clv3 mutant (red), and cytokinin-treated clv3 mutant (black). (G) Simulated cell numbers for wild type, clv3 mutant, cytokinin-treated wild type, and cytokinin-treated clv3 mutant. [Scale bars: 200 μm (A–D).]

Model tests for two cases: first, cytokinin-induced rescue of inner whorl floral organ development requires WUS function; and second, cytokinin affects WUS apical–basal positioning. (A and B) Sepal, petal, stamen, and carpel number for mock-treated and cytokinin-treated wus-1 (n = 55, 81) (A) and wus-6 (n = 353, 182) (B) flowers. (C) Simulations of CLV3 (green) and WUS (red) abundance within a column of cells along the apical–basal axis of the SAM (apical cell, 0; basal-most cell, 30) in wild type (upper plot), wus-6 mutant (center plot), and the wus-6 mutant treated with cytokinin (lower plot). (D and E) Simulations of CLV3 concentration along a column of cells (D) and WUS concentration along the same column of cells (E) (cytokinin perturbation simulations; blue, mock; red, cytokinin-treated). (F and G) Longitudinal view of WUS_pro::WUS-2xYFP (red) and TCS_pro::GFP (green) in mock-treated SAM. (H and I) Longitudinal view of WUS_pro::WUS-2xYFP (red) and TCS_pro::GFP (green) in cytokinin-treated SAM. (Scale bars: 50 μm.)

Discriminating feedback models between WUS and cytokinin biosynthesis validated by experimental observations. Simulation results for WUS, CLV3, phosphorylated type B ARR (Bp), and cytokinin concentration as a function of cell number, along the apical–basal axis. Wild type (A) and clv3 loss of function (B) corresponding to the model where we include WUS positive feedback on cytokinin synthesis. Wild type (C) and clv3 loss of function (D) corresponding to the model where we include WUS negative feedback on cytokinin synthesis (SI Appendix). (E and F) TCS_pro::GFP reporter expression in clv3-2 mutant mock-treated SAM and in clv3-2 mutant cytokinin-treated SAM. (G and H) LOG4_pro::2xYpet-N7 reporter expression in clv3-2 mutant mock-treated SAM and in clv3-2 mutant cytokinin-treated SAM. FM4-64 membrane dye marks cell membranes in red (E–H). (I) The schematic shows that cell divisions are controlled antagonistically by CLV3 and cytokinin and that cytokinin biosynthesis is negatively regulated by WUS. This is in addition to the feedback between CLV3 and WUS, as well as WUS induction by cytokinin. (Scale bars: 50 μm.)

Distribution of cytokinin synthesis, perception, and signaling relative to WUS within the SAM and a dynamic model for pattering the apical–basal axis of the SAM during growth. (A) Longitudinal view of LOG4_pro::2xYpet-N7 reporter expression (green) and cell membranes (red; FM4-64) in the SAM and floral meristem. (B) Longitudinal distribution of WUS_pro::DsRed-N7 (red) within the apical half of the AHK4_pro::GFP (green) domain. (C and D) Longitudinal view of WUS_pro::WUS-2xVenus (red) (C) and TCS_pro::GFP (green) (D) in the SAM and floral meristem. PIN1_pro::PIN1-CFP (blue) marks cell membranes. [Scale bars: 50 μm (A–D)]. (E) Schematic of the regulatory interactions between cytokinin/WUS and WUS/CLV within the SAM. (F) WUS expression determined by CLV3 suppression [f_sup(CLV3(D))] and activation by cytokinin [f_act(cyt(D))] results in a maximum (f_max) for the WUS distribution [f_WUS(D)]. (G) Initial pattern of expression of key components in single column of 30 cells spanning the apical–basal axis of the SAM. First column, CLV3 (green) and WUS (magenta); second column, type A ARR (teal) and phosphorylated type B ARR (red). Interspaced regions are in dark blue. (H) Patterns of components after a growth simulation labeled as in G. In H, the larger plot shows all 120 cells after growth, whereas the smaller plots show only the apical-most 30 cells for comparison with patterns in G.