BASL and EPF2 act independently to regulate asymmetric divisions during stomatal development

Abstract

The initiation of stomatal development in the developing Arabidopsis epidermis is characterized by an asymmetric 'entry' division in which a small cell, known as a meristemoid, and a larger daughter cell is formed. The meristemoid may undergo further asymmetric divisions, regenerating a meristemoid each time, before differentiating into a guard mother cell which divides symmetrically to form a pair of guard cells surrounding a stomatal pore. Recently EPF2 and BASL have emerged as regulators of these asymmetric divisions and here we present results indicating that these two factors operate independently to control stomatal development.

Figure 1

BASL & EPF2 act independently. Cleared Normarski images of abaxial epidermis of (A) Col-0, (B) basl-4, (C) epf2-1, (D) basl-4;epf2-1 leaves approximately 2 cm in length. gc, pair of guard cells; pc, pavement cell; m, meristemoids. *indicates meristemoids that have undergone an abnormal symmetric division. Scale bar = 30 µm. (E) Densities of stomatal and (F) stomatal clusters of abaxial surface of fully expanded leaves. Error Bars are SEM. Photographs of mature plants of Col-0 (G) and basl-4;epf2-1 (H). Scale bar = 3 cm. Plant transformation, plant material, GUS staining and microscopy were as described in. basl-4 (SALK_086936) was obtained from NASC and verified by PCR with primers 5′-CTC GTG ACA AA G AAA CAC AAA CA -3′, 5′-GAA TCT ACA ACA TT G GAA CCC TAA A-3′. Crosses between basl-4 and epf2-1 were verified by PCR.

Figure 2

Additional small epidermal cells in basl-4;epf2-1 are stomatal lineage cells. Col-0, epf2-1, basl-4 and basl-4;epf2-1 mutants expressing the pTMM:GUS-GFP transcriptional reporter histochemically stained for GUS (A–D) or examined under UV fluorescence (E–H). Staining and fluorescence are not from the same plant. Scale bar (A–D) = 40 µm, (E–H) = 20 µm. Three independent lines of basl-4;epf2-1 expressing pTMM:GUS-GFP were obtained and all showed similar staining or GFP fluorescence pattern.