γ-Radiation Induces Leaf Trichome Formation in Arabidopsis^{<a href="#FN1" rid="FN1" class=" fn">1</a>}

Abstract

Massive doses of ionizing radiation have been shown to induce physiological changes in plants, such as enhancement of respiration, increase in ethylene production (Young, 1965; Abdel-Kader et al., 1968; Lee et al., 1968; Akamine and Goo, 1971; Romani, 1984), induction of enzyme activities (particularly for phenolic metabolisms; Riov et al., 1970; Pendharkar and Nair, 1975; Frylink et al., 1987), and accumulation of Suc (Hayashi and Aoki, 1985) and specific protein species (Ferullo et al., 1994). Cellular macromolecular components such as cell walls, membranes, and DNA are also markedly affected by ionizing radiation (Casarett, 1968). These effects are considered a consequence of both the direct interactions between the ionizing radiation and the macromolecular structures and the indirect action of AOS generated by water radiolysis.

We used Arabidopsis to elucidate the mechanisms of stress recognition, signal transduction, and the initiation of plant self-protective programs at the molecular level because this plant provides certain key advantages for genetic and molecular studies. We observed that Arabidopsis displayed particular responses within several days after irradiation with γ-rays (1–3 kGy) from cobalt-60 (1.17 and 1.33 MeV). These responses included accumulation of anthocyanin in the aerial part of plant, induction of new trichome formation on the adaxial surface of mature leaves, radial expansion of root cell layers, and elongation of root hairs. In this paper we characterize γ-radiation-induced trichome formation.

Arabidopsis trichomes are single-cell-originated epidermal hairs that serve as an appropriate model for plant cell differentiation and cell elongation. The trichomes are present on the surfaces of the leaves, stems, and sepals and on the margins of leaves and sepals. Plant growth does not require trichomes, because plants that lack trichomes appear to be fully viable and fertile (Koornneef et al., 1983). We have taken a genetic approach to studying trichome development. More than 70 trichome mutants representing 21 different genes were isolated (Lee-Chen and Steinitz-Sears, 1967; Feenstra, 1978; Koornneef et al., 1982; Haughn and Somerville, 1988; Huelskamp et al., 1994; Marks and Esch, 1994). Genetic and molecular biological studies have revealed that the GL1 (GLABROUS1) and TTG (TRANSPARENT TESTA GLABROUS) genes are required for the initiation of trichome development.

The GL1 gene has been cloned and shown to encode a putative myb-class transcription factor (Oppenheimer et al., 1991; Larkin et al., 1994b). GL1 transcripts are present at a low level throughout the protoderm, with much higher levels of expression in developing trichomes and presumptive trichome precursor cells (Larkin et al., 1993). Mutations in TTG also affect anthocyanin synthesis, integument development (Koornneef, 1981), and root-hair patterning (Galway et al., 1994). The observation that expression of the maize R gene in ttg mutant plants complemented ttg mutation functionally (Lloyd et al., 1992) suggested that TTG encodes a homolog of the maize R gene or the gene that regulates the expression of an R homolog. However, recent cloning of the TTG gene revealed no sequence homology to the R gene (A.R. Walker and J.C. Gray, personal communication).

The ectopic expression of GL1 with the cauliflower mosaic virus 35S promoter does not lead to increased trichome formation, nor does it bypass the requirement for TTG, whereas the ectopic expression of both GL1 and R induced an increase in the trichome number (Larkin et al., 1994a). These results suggest that the GL1 and TTG gene products cooperate in promoting trichome initiation. The GL2 (GLABRA2) gene is necessary for subsequent phases of trichome morphogenesis, such as cell expansion, branching, and maturation of the trichome cell wall (Koornneef et al., 1982; Marks and Esch, 1994; Rerie et al., 1994). The GL2 gene has been also cloned and shown to have sequence similarity to homeodomain proteins (Rerie et al., 1994). Detailed expression analysis using anti-GL2 antibodies and the GUS reporter gene fused to the GL2 promoter revealed that GL2 expression persists in mature trichomes. A requirement of GL1 for GL2 expression in vivo has been suggested (Szymanski et al., 1998). An examination of the distribution of trichomes early in development disclosed that trichomes are initiated adjacent to other trichomes much less frequently than would be expected by chance.

Genetic and molecular biological studies have revealed the key genes related to the developmental process of trichome formation and have started the elucidation of the functions of these genes and their interactions. However, stress-induced trichome formation has never been found. In this paper we describe, for the first time to our knowledge, the induction of new trichomes after γ-irradiation. We also examined the conditions of new trichome formation thus induced and found that the AOS generated by water radiolysis may also contribute to the induction of new trichomes.

Data are means ± se of 100 plants.

Data are the means ± se of 100 plants.

ACKNOWLEDGMENTS

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