Extended Data Figure 7. Auxin is required for cell division, expansion and differentiation.

a, Protoxylem differentiation is inhibited in the axr3-1 mutant. First protoxylem is highlighted with an arrow. Higher up in the root protoxylem differentiation is often sporadic (inset, arrows indicate a stretch of a single protoxylem element). b, Twenty-four hour induction of pG1090:XVE>>axr3-1-RFP with 17β-oestradiol (est) inhibits xylem differentiation and root hair outgrowth. Left inset, a single protoxylem vessel highlighted with two arrows. Right inset, the arrow highlights the beginning of a more continuous protoxylem strand, which was probably already present before the induction of axr3-1-RFP. Yellow bars in b, c show the areas in the root in which visible protoxylem are present. Yellow arrow in b, c marks the first root hair. Est, 5 μM 17β-oestradiol. c, Twenty-hour treatment of 4-day old wild-type root (Col) with 25 μM auxinole (auxin antagonist) inhibits xylem differentiation, cell expansion and root hair outgrowth. Xylem is typically differentiated as short, sporadic stretches. Left inset, arrow shows the end of a stretch of a xylem strand comprising approximately six protoxylem elements; right inset, higher up in the root two continuous protoxylem strands appear (arrows). These strands were probably already present before auxinole application. The middle inset shows the presence of short cells (marked with arrowheads) high up in the root, indicating that cell expansion was defective. The cell length typically varied along the root. d, Bar plot shows that the final length of cortex cells is shorter when auxin signalling is inhibited by auxinole (n = 35 cells in 6 roots) or inducible pG1090:XVE>>axr3-1-RFP (n = 47 cells in 6 roots) when compared with differentiation zone cells in the control roots (n = 51 cells in 7 roots and n = 38 cells in 7 roots, respectively) located at a similar distance from the root tip. Asterisks, Mann–Whitney U test (P < 0.001). Error bars show s.d. e–j, The consequence of auxin application or inhibition of auxin signalling on marker gene expression. e, f, Auxin application (5 μM IAA) rapidly leads to expression of the root hair differentiation marker pEXP7:GUS (ref. ³³) (e) and the xylem differentiation marker S18 (ref. ³²) (f) in the elongation zone. Note: high auxin levels (such as 5 μM IAA) are inhibitory for root hair elongation. Therefore, even though the root hair marker pEXP7:GUS rapidly spreads into the elongation zone, root hair elongation is less pronounced there. e–g, Twenty-four hour induction of pG1090:XVE>>axr3-1-RFP (est) inhibits the expression of root hair differentiation marker pEXP7:GUS (e) and the xylem differentiation marker S18 (ref. ³²) (f), as well as cell division marker CYCB1;1-GUS²² (g). Note: both the signal intensity and the number of CYCB1;1–GUS-expressing cells are decreased in g. Twenty-four hour induction of XVE>>axr3-1 leads either to disappearance of S18 fluorescence (no expression in f), or S18 was present in short fragments (fragmented). h,i, Expression of PLT2–YFP continued to mark the shortened meristem after auxin signalling was inhibited by induction of pG1090:XVE>>axr3-1-RFP (est) (h) or by treatment of the seedlings with 25 μM auxinole (axnl) (i). j, Auxinole rapidly inhibits cell division as indicated by reduction of the number of CYCB1;1–GFP-expressing cells after auxinole treatment.