Figure 1.

Glc-TOR signaling is compromised in prr579 mutant. (A) Venn digram showing overlap between genes activated by TOR signaling (9) and genes down regulated in prr579 mutant (39). (B) Compared to the WT (Col-0), primary root growth was reduced in prr579 after sugar-induced reactivation of TOR-signaling. Three-day-old seedlings grown in liquid sugar-free 1/2 MS were treated with 15 mM Glc or Suc for 3 days in weak light (22°C, LD). Red arrows indicate the hypocotyl and root junctions. DAG, days after germination; D, day; Mock, sugar-free; Glc, glucose; Suc, sucrose; (scale bar: 0.5 cm). (C) Quantitative analysis of primary root length in (B). Data represent mean ± s.e.m. (n = 18). The experiment was carried out with three biological replicates with similar results. The lengths of primary roots were measured by using ImageJ. (D, E) Root meristem proliferation was not fully activated in prr579 mutant after activation of TOR signaling by sugars. (D) Differential interference contrast (DIC) imaging of root meristems (scale bars: 50 μm). The blue arrow indicates the root quiescent cells, and the red arrow indicates the transition between meristem zone and elongation zone. (E) Quantitative analysis of root meristem length in (D). Data represent mean ± s.e.m. (n = 13). The lengths of root meristems were measured by using ImageJ. (F) The prr579 mutant was defective in S-phase entry of cell cycle as evidenced by reduced 5-ethynyl-2′-deoxyuridine staining (EdU) signals in situ (scale bars: 50 μm). (G) Time-course expression of TOR in Col-0 and prr579. The total RNA was isolated from the roots of 6-day-old seedlings grown in half-strength liquid MS medium without sugar in 12-h weak light (13 μmol m⁻¹ s⁻¹)/12-h dark cycles at 22°C. The gene expression level was normalized by the geometric mean of ACT2 and TUB4expression. Data represent mean ± s.e.m. of three biological replicates. (H) Expression of S-phase marker genes was reduced in prr579 mutant. For RNA extraction, the roots were harvested from the seedlings treated with 15 mM Glc or Suc for 3 days after growing in liquid sugar-free 1/2 MS for 3 days in weak light (22°C, LD) at ZT8. The gene expression level was normalized by ACT2 expression. The bar values represent the gene expression levels in sugar treatments relative to the corresponding expression levels in sugar-free condition. Data represent mean ± s.e.m. of three biological repeats. Different letters in (C) and (E) represent significant difference at P < 0.01 by one-way ANOVA using SPSS software. Uppercase letters compare with each other in 15 mM Glc treatment condition, and lowercase letters compare with each other in 15 mM Suc treatment condition. The asterisk in (G) and (H) indicates significant difference as * P < 0.05, ** P < 0.01, and *** P < 0.001 by t-test.