Accumulation of peroxidase-related reactive oxygen species in trichoblasts correlates with root hair initiation in barley

Root hairs are an important model in studies of cell differentiation and development in higher plants. The function of NADPH oxidase-related reactive oxygen species (ROS) in root hair development has been reported extensively in studies on Arabidopsis. In this study, we investigated the mechanism of the initiation of root hair formation, mediated by the peroxidase-dependent production of the highly reactive hydroxyl radical in barley (Hordeum vulgare L.). The distribution of ROS, including the hydroxyl radical (radical dotOH) and superoxide (O2radical dot−) was assessed using hydroxyphenyl fluorescein and nitroblue tetrazolium chloride, respectively, in the roots of wild-type plants and two root-hair mutants: root-hairless (rhl1.a) and with root hair growth blocked at the primordium stage (rhp1.b). Peroxidase-dependent radical dotOH accumulation was linked to root hair initiation and growth in plants where root hair formation was at least initiated, whereas radical dotOH was not detectable in the epidermis of the root-hairless mutant rhl1.a. O2radical dot− distribution in the roots of rhl1.a and rhp1.b mutants was not impaired and did not influence the root hair phenotype. Peroxidase inhibitor treatments of wild-type roots dramatically reduced the ability of growing roots to form root hairs and thus phenocopied the root-hairless phenotype. Expression of two candidate peroxidase genes, HvPRX45 and HvPRX2, was analyzed and their possible role in root hair-specific production of hydroxyl radicals was discussed. We propose a model of a two-step, coordinated ROS formation process in root hair cells that involves root hair-specific peroxidase(s) and root hair-specific NADPH oxidase necessary for a proper root hair formation in barley.