Figure 4.

Dynamics of E. lathyris laticifer differentiation and activity in the whole plant and during leaf development. A, Major isoprenoids present in latex: CYC, cycloartenol; LAN, lanosterol; BUT, butyrospermol; 24M, 24-methylene-cycloartanol. B and C, GC-MS analysis of latex extracts (B) and whole-leaf extracts (C). D, Relative content of each of the four major latex isoprenoids in extracts derived from whole plants, stems, and leaves. E, Comparative analysis of triterpenoid content in stems, leaves, and roots with respect to its net accumulation in the whole developed plant. Bars represent mean ± sd, n = 9 independent plants. An ANOVA was conducted to assess significant differences in isoprenoid content, with a priori P < 0.05 level of significance; the letters above the bars indicate different homogeneous groups with statistically significant differences. F, Image of E. lathyris leaves at the four stages (1–4) of leaf expansion. G, Whole-mount staining of leaves and close-up of a sector of the leaf blade, showing the presence of laticifer cells at the four stages of leaf expansion shown in F. The insert cartoons serve to indicate the relative position of the selected leaves in the stem. H, LI recording for each stage of leaf expansion. I, Triterpenoid content in leaves at different stages of leaf expansion. Bars represent mean ± sd, n = 9 independent plants. J, Expression of the EH, PE, and SMT1 genes at different stages of leaf expansion. Relative expression was assayed by RT-qPCR on total RNA from leaves at the indicated stages. Data represent means ± sd (n = 3 biological replicates). Expression was normalized to the constitutive Histone H3 gene, then to expression attained at stage 1 of leaf expansion.