Isolation and Initial Characterization of Arabidopsis Mutants That Are Deficient in Phytochrome A

Abstract

Phytochrome, a red/far-red-light photoreceptor protein of plants, is encoded by a small gene family. Phytochrome A (PHYA), the product of the PHYA gene, is the predominant molecular species of phytochrome in etiolated tissue and has been best charaderized biochemically. To define a role for PHYA, we isolated new mutants, designated frel (far-red elongated), in Arabidopsis thaliana that were specifically deficient in PHYA spectral activity and protein accumulation. These mutants were identified on the basis of their long hypocotyl phenotype under continuous far-red light. Although the frel mutants lacked the hypocotyl response to continuous far-red light, their responses to continuous white light and to end-of-day far-red-light treatments were normal. Thus, PHYA appears to play only a minor role in the regulation of hypocotyl elongation under natural conditions. In contrast, the frel mutation affected greening; a frel mutant was less able than the wild type to deetiolate after growth in the dark. However, the potentiation effect of a red-light pulse on accumulation of chlorophyll was not changed significantly in the frel mutants. Thus, the function of PHYA might be highly specialized and restricted to certain phases of Arabidopsis development.

Light-Induced Nuclear Translocation of Endogenous Pea Phytochrome A Visualized by Immunocytochemical Procedures

“…The one exception to date is the report of the nuclear localization of PHYB (Sakamoto and Nagatani, 1996) in which a PHYB-deficient mutant was used as a negative control. In the earlier studies, the antibodies most likely reacted mainly to PHYA (Nagatani, 1997), because this is the predominant species of phytochrome in dark-grown plants (Somers et al, 1991;Nagatani et al, 1993;Weller et al, 1997). However, artifactual staining can often occur during the immunostaining process, and study of a mutant specifically deficient in the antigen in question is crucial to evaluate the specificity of the antibodies.…”

“…Phytochrome extraction buffer (Nagatani et al, 1993) was added to the powder (2 mL per gram of tissue) and allowed to sit for 15 min at room temperature. After centrifugation (27.6g for 20 min at 4ЊC), the supernatant was retained, and saturated ammonium sulfate solution was added (2:3 v/v) before 30 min of incubation on ice.…”

An Arabidopsis Mutant Hypersensitive to Red and Far-Red Light Signals

“…Crude extracts were prepared from 7-day-old etiolated and lightgrown seedlings, as described by Nagatani et al (1993). Proteins were separated by 7.5% acrylamide SDS-PAGE and blotted onto nitrocellulose membranes.…”

“…To determine whether a specific phytochrome pathway was modified by the psi2 mutation, we constructed double mutants by crossing psi2 with three phytochrome-deficient mutants: the phyB-1 mutant (Somers et al, 1991), which lacks the PhyB apoprotein, the phyA-201 mutant, which lacks phytochrome A (Nagatani et al, 1993), and hy2, a mutant in the chromophore of any phytochrome type (Koornneef et al, 1980). A double mutant was also constructed between psi2 and fhy1, a mutant that is impaired in a branch of the phytochrome A signaling pathway (Whitelam et al, 1993; Barnes et al, 1996).…”

Two Small Spatially Distinct Regions of Phytochrome B Are Required for Efficient Signaling Rates.

“…Contrasting photosensory functions of the phyA and phyB holoproteins have been defined through analysis of responses to constitutive overexpression of Quail, 1989, 1991;Kay et al, 1989;Keller et al, 1989;Cherry et al, 1991;Wagner et al, 1991;McCormac et al, 1993) and mutations in (Nagatani et al, 1991(Nagatani et al, , 1993Somers et al, 1991;Dehesh et al, 1993;Parks and Quail, 1993;Reed et al, 1993;Whitelam et al, 1993;Wagner and Quail, 1995;Xu et al, 1995) hypocotyl elongation, hook opening, and cotyledon expansion) in continuous far-red light (FRc), whereas phyB is primarily responsible for deetiolation in continuous red light (Rc). phyA is most abundant in dark-grown seedlings (m50-fold phyB levels), whereas, due to light-induced phyA degradation, phyB is more abundant in Rc (approximately twofold phyA) (Somers et al, 1991;.…”