Maize Genetics Cooperation Newsletter 80. 2006.

BERGAMO, ITALY

Istituto Sperimentale per la Cerealicoltura

Characterization of gl1, a maize gene that affects cuticular wax accumulation

--Sturaro, M; Salamini, F; Schnelzer, E; Motto, M

The surfaces of land plants are covered with a cuticle secreted by epidermal cells, which plays several protective roles and consists of a reticulated cuticle membrane covered and interspersed by amorphous waxes (Kunst and Samuels, Progr. Lipid Res. 42:51-80, 2003).

The maize glossy1 (gl1) gene is one of several loci involved in epicuticular wax biosynthesis in seedling leaves. Due to the strong reduction of juvenile waxes, mutations at this locus confer a glossy phenotype to the first five to six leaves, in contrast to the dull appearance of their wild-type counterparts. Although sequence analysis predicts a metabolic function for gl1, its specific activity in wax biosynthesis has not been defined yet.

To gain insights into gl1 function, transcriptional analysis and microscopic inspection of mutant leaves were performed. From the expression profile it turned out that gl1 activity is not restricted to the juvenile developmental phase of the maize plant, but it is active also in adult leaves and anthers (Figure 1). Moreover, gl1 transcription is negatively affected by drought, although this stress condition promotes wax biosynthesis (Figure 2). These data suggest a broader role for gl1 than anticipated on the basis of the

Figure 1. Analysis of gl1 transcription. Northern analysis of gl1 (A) and cyGAPDH (B) expression in: (1) young WT leaf, (2) young gl1-Ref leaf, (3) root, (4) old WT leaf, (5) silk and (6) anther. Expression in the same tissues of gl1 (C) and cyGAPDH (D) as assayed by RT-PCR. Sizes in Kb are given on the right.

Figure 2. Effect of water stress on gl1 transcription. Time course of gl1 and cyGAPDH expression during a 5-day water stress period. RWC: Relative Water Content.

visual phenotype of mutant plants. Accordingly, ultrastructural analysis indicated a pleiotropic effect of the gl1-Ref mutation on juvenile epidermis development (Figure 3). In addition to the reduction in wax biosynthesis, scanning electron microscopy (SEM) analysis revealed alteration of leaf trichomes, namely decreased trichome size to half of the wild-type and increased trichome frequency. Transmission electron microscopy (TEM) analysis highlighted a strong reduction of cuticle membrane thickness in mutant seedlings. The effect of the gl1 mutation seems to be limited to the epidermal layer; in fact, as revealed by light microscopic inspection, the whole architecture of mutant seedling leaves is not altered.

Figure 3. Ultrastructural analysis of seedling leaves. SEM analysis of WT (A) and mutant (B) leaf surface. Inserts: close-up view of upper surface showing details of trichome morphology and density (all images are at 100x enlargement). TEM analysis of WT (C) and mutant (D) cuticle membrane.

gl1 is the putative orthologue of the Arabidopsis WAX2 gene given the strong homology of their sequences (62% identity at the protein level). The ultrastructural analysis of the gl1-ref mutant further supports this hypothesis; in fact, similarly to gl1, mutations of the WAX2 gene alter cuticle membrane synthesis, epicuticular wax production and trichome morphology. Research is in progress in this laboratory to further clarify gl1 functions.

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