Figure 1.
Schematic of a representative FLA containing two AGP domains and two fasciclin domains before and after post-translational modifications.
Deduced proteins include an N-terminal secretion signal, two fasciclin domains, two AGP domains and a C-terminal signal sequence for addition of a GPI-anchor. Mature FLAs are predicted to be extensively modified post-translationally with peptidyl proline (Pro) modified to hydroxyproline (Hyp), the addition of O-linked oligo/poly-saccharide chains to Hyp residues, and the addition of a C-terminal GPI-anchor.
Figure 2.
Characterisation of DNA insertion mutants for FLA1.
A Schematic representation of the T-DNA insertion locus (black triangle) of fla1-1, located in the intron of FLA1 identified by screening the Feldmann mutant lines (WS ecotype), and fla1-2, located in exon 1 (SALK insertion mutant, COL ecotype). B Phenotypic comparison of six-week-old WS and fla1-1 mutant Arabidopsis plants grown in soil. Appearance of key developmental stages (days) of seedlings on plates (C) and pots (D) according to Boyes et al. [32].
Figure 3.
Shoot induction phenotypes of fla1 mutants.
A-D Wild-type and fla1-1 mutant root explants after 4 d treatment on callus induction medium (CIM) then 14 d incubation on shoot induction medium (SIM). (A and B) Wild-type (WS) and (C and D) fla1-1 mutant root explants. (A and C). Root explants from the zone of maturation of the primary root where lateral roots were forming were incubated on CIM then transferred to SIM. (B and D) Four representative root explants (indicated on (A) and (C) by an asterisk) from the zone of maturation of wild-type (B) and fla1-1 mutant (D) roots after CIM and SIM treatment. The long arrows indicate the base of a new shoot and the arrowheads indicate the tip of a new root. 13 independent experiments comparing WS and fla1-1 were performed. (E) Wild type and (F) mutant root segments after only 1 d on CIM, followed by 14 d SIM. (G) Wild-type (WS) and fla1-1 mutant root explants after 4 d on CIM then 14 d SIM. (H) Wild-type (COL) and fla1-2 mutant root explants after 4 d on CIM then 14 d SIM. Experiments comparing COL and fla1-2 were repeated three times, and WS2 and fla1-1 were also compared in each of these three experiments. Scale bar is 1 mm.
Table 1.
Shoot and root regeneration of wild-type and fla1-1 mutants.
Figure 4.
Light microscopy of wild-type and fla1-1 root explants after CIM and SIM treatment.
Sections through a region of WS (A to E) and fla1-1 (F to J) root explants. Transverse sections through the differentiated zone of a 14 d wild-type (A) and fla1-1 mutant (F) root. Transverse sections through root explants incubated on CIM for 2 d (B and G) and 4 d (C and H). Wild-type (D and E) and fla1-1 mutant (I and J) root explants after 4 d CIM and 7 d (D and I) or 14 d (E and J) SIM treatment. Centres of radial organisation which are presumptive sites of meristem formation are indicated by arrows. Sections were stained with toluidine blue. Scale is 0.1 mm; P, pericycle.
Figure 5.
Comparison of root phenotypes of fla1 mutant alleles in different genetic backgrounds.
A Length of primary root (cm) of wild-type (WS), fla1-1, wild-type (COL) and fla1-2. Significance levels a and b (P<0.01) are based on Tukey's post test (1-way analysis of variance). B Number of lateral roots of wild-type (WS), fla1-1, wild-type (COL) and fla1-2. Number of seedlings analysed was n = 12, 14, 13, 14 for WS, fla1-1, COL, fla1-2 respectively. Significance levels a and c (P<0.01), others (P<0.05), are based on Tukey's post test (1-way analysis of variance). Error bars represent standard deviation of the mean.
Figure 6.
FLA1 promoter:GUS analysis of 14 d plate grown seedlings and flowering, soil grown Arabidopsis plants.
GUS activity was detected in the A petioles and roots. B petioles and base, stomata and leaf hairs (trichomes, indicated by a white arrow) of young leaves. C stomata throughout the leaf and at the hydathode at the leaf tip (indicated by a black arrow). D lateral roots. E primary root, but only in the early elongation zone of the root tip. F developing lateral roots in the vascular and pericycle cells. G developing anthers of closed flowers. H stamen filaments (but not anthers) of open flowers. I early embryos of fertilised stigma and style, dissected from an open flower. GUS expression was observed in vegetative portion of developing siliques (J) including the stomata (K) and not in embryos (L). Scale bar is 0.1 mm, except in (C) where it is 0.03 mm.
Figure 7.
FLA1 promoter:GUS analysis of Arabidopsis root explants after CIM and SIM treatment.
GUS activity was detected in the A lateral roots of 14 d Arabidopsis seedlings before incubation on CIM and SIM (see also Fig. 6). B some vascular and pericycle cells in root explant after 2 d CIM. C along the length of the root explant in the vasculature and de-differentiating pericycle cells after 3 d and 4 d (D) CIM treatment. E in callus and vascular tissue of root explant after 4 d CIM and 3 d SIM. (F and G) in the vascular tissue and pericycle cells of newly forming roots after 4 d CIM and 7 d SIM. H in the vascular tissue of newly formed roots of root explants after 4 d CIM and 14 d SIM. Scale bar is 0.1 mm.