Roles for Class III HD-Zip and KANADI genes in Arabidopsis root development

Abstract

Meristems within the plant body differ in their structure and the patterns and identities of organs they produce. Despite these differences, it is becoming apparent that shoot and root apical and vascular meristems share significant gene expression patterns. Class III HD-Zip genes are required for the formation of a functional shoot apical meristem. In addition, Class III HD-Zip and KANADI genes function in patterning lateral organs and vascular bundles produced from the shoot apical and vascular meristems, respectively. We utilize both gain- and loss-of-function mutants and gene expression patterns to analyze the function of Class III HD-Zip and KANADI genes in Arabidopsis roots. Here we show that both Class III HD-Zip and KANADI genes play roles in the ontogeny of lateral roots and suggest that Class III HD-Zip gene activity is required for meristematic activity in the pericycle analogous to its requirement in the shoot apical meristem.

Figure 1.

KANADI and Class III HD-Zip expression in Arabidopsis lateral roots. A to H, G to M, and O, Whole mounts showing GUS expression. F, G, N, P, Eight-micrometers wax sections showing GUS expression. A to H, PHB expression. A to C, PHB is detected throughout the lateral root primordia. A, At stage I, anticlinal divisions are observed (arrowheads). B, By stage II, periclinal divisions are evident (arrowheads). C, PHB is expressed throughout stage VI primordia. D, At emergence, PHB is seen to become restricted to the central region of the primordia. E and F, Postemergent primordia do not show PHB gene expression in the outer cell layers. G, This cross section near the root apex clearly illustrates PHB is expressed centrally. H, PHB gene expression in the lateral root apical meristem (RAM). The early expression patterns of both REV and ATHB8 are identical to PHB and therefore are not shown here. I, REV expression in the lateral root apex is not detected in the apical meristem, but is expressed in the vascular cambium. J, KAN4 expression first becomes apparent in Stage VII primordia. K, Postemergence, KAN4 expression is detected in the root cap and in the periphery at the base of the developing lateral root primordium. L to P, KAN1 is expressed in the periphery of lateral root apices. L, KAN1 is first detected in the apex of postemergent lateral roots. M, KAN1 expression in a top view of an adventitious root. N, Cross section near the root apex. O and P, KAN1 expression in the root apical meristem. Note that KAN2 expression is not shown as it exactly matches that of KAN1. KAN3::GUS lines show no expression in either lateral root primordia or root apices.

Figure 2.

Cross sections of roots of wild-type plants and genotypes with altered KANADI and Class III HD-Zip functions. All plants were sectioned 5 DAG except 35S::KAN1:VP16:GR grown continuously on dexamethasone, which was sectioned 7 DAG, as this was when the root was the same length as the other plants. All sections were made at the base of the root toward the hypocotyl to ensure that the vascular cylinder was at maturity. A and B, Landsberg erecta showing the variation seen between individual roots. A, Xylem poles, stained light blue, that do not connect across the center of the vascular cylinder. B, Xylem extending completely across the vascular cylinder. C to F, None of the genotypes with altered KANADI or Class III HD-Zip functions show any clear deviation in vascular development from wild type. Each shows the two clear xylem poles juxtaposed with two poles of smaller phloem cells. C, rev-10d. D, 35S::KAN1:VP16:GR grown continuously on dexamethasone. E, phb-6 phv-5 rev-9. F, kan1-2 kan2-1 kan3-1. Size bar represents 100 μm.

Figure 3.

Whole root systems of wild type and genotypes with altered KANADI and Class III HD-Zip functions 10 DAG with and without auxin treatment. A, C, E, G, K, M, Untreated plants. B, D, F, H, L, Plants treated with auxin for 70 h. I, 35S::KAN1:GR treated with dexamethasone for 94 h. J, 35S::KAN1:GR treated with dexamethasone for 94 h and auxin for 70 h. N, 35S::KAN1:VP16:GR treated continuously with dexamethasone. O, 35S::KAN1:VP16:GR treated continuously with dexamethasone and auxin for 70 h. P, Closer view of the root tip in O. Size bars represent 2 mm.