Oxygenic photosynthetic organisms possess sophisticated mechanisms to adapt to their environment. Dependent upon light as an energy source, these organisms must cope with too little and too much light—an environmental factor that is especially challenging for highly pigmented species living in an aerobic environment. Plants therefore possess light receptor systems to recognize and respond to changes in light quality, fluence rate, direction and duration in their environment (Briggs and Spudich eds. (2005) Handbook of Photosensory Receptors (Weinheim: Wiley VCH)). Among the many physiological processes under light control include seed germination, seedling growth, synthesis of the photosynthetic apparatus, timing of flowering, shade avoidance, and senescence. Such light-regulated growth and developmental responses are collectively known as photomorphogenesis (Schäfer and Nagy eds. (2005) Photomorphogenesis in Plants and Bacteria: Function and Signal Transduction Mechanisms (3rd Edition), 3rd Edition (Dordrecht, The Netherlands: Springer)). Phytochrome was the first of the photomorphogenetic photoreceptor families to be identified in plants over 50 years ago (Butler et al. (1959) Proc. Natl. Acad. Sci. USA 45: 1703-1708; Rockwell et al. (2006) Annu. Rev. Plant Biol., 57: 837-858).
Synthesized in the red light-absorbing Pr form, plant phytochromes are regulated by red light (R) absorption that initiates the photochemical interconversion to the far red (FR) light-absorbing Pfr form. FR promotes the reverse conversion of Pfr to Pr—a process that typically abolishes the R-dependent activation of the photoreceptor (Schäfer and Nagy eds. (2005) Photomorphogenesis in Plants and Bacteria: Function and Signal Transduction Mechanisms (3rd Edition), 3rd Edition (Dordrecht, The Netherlands: Springer)). This R/FR photoreversibility is conferred by a linear tetrapyrrole (bilin) prosthetic group that is covalently attached to the phytochrome apoprotein (Rockwell et al. (2006) Annu. Rev. Plant Biol., 57: 837-858). Supporting evidence for the central dogma of phytochrome action, that is that Pfr is the active form, has been accumulating for years. Much of this evidence reflects the strong correlation between the amount of Pfr produced by a given fluence of light and the magnitude of the biological response. While the central dogma appears to hold true for R/FR-reversible low-fluence responses (LFR) and for R-dependent very low fluence responses (VLFR) (Shinomura et al. (1996) Proc. Natl. Acad. Sci. USA 93: 8129-8133), FR high irradiance responses (FR-HIR) do not conform to this simple view of phytochrome action (Furuya, and Schäfer (1996) Trends in Plant Sci., 1: 301-307; Shinomura et al. (2000) Plant Physiol. 122: 147-156). Such data indicate that Pr, Pfr, photocycled-Pr and possibly intermediates produced during Pfr to Pr photoconversion may all function to transduce the light signal depending on the phytochrome.
In flowering plants, phytochromes are encoded by a small family of genes that have arisen by repeated gene duplication of a eukaryote phytochrome progenitor during the course of evolution (Mathews et al. (1995) Annal. Missouri Botanical Garden 82: 296-321). In the model plant Arabidopsis thaliana, the phytochrome family consists of five genes—denoted PHYA-E (Clack et al. (1994) Plant Mol. Biol. 25: 413-427), while monocots species (eg. rice or maize) appear only to possess representatives of the PHYA-C families (Mathews and Sharrock (1997) Plant Cell Environ. 20: 666-671; Sawers et al. (2005) Trends in Plant Science 10: 138-143). Plant phytochromes can be classified into two groups based upon their light-stability. Phytochromes encoded by the PHYA gene family are responsible for the light-labile pool, while PHYB-E genes encode the light-stable phytochromes (Furuya (1993) Ann. Rev. Plant Physiol. Plant Mol. Biol. 44: 617-645). The pronounced light-lability of phyA holoproteins is due to two processes—light-dependent transcriptional repression of the PHYA gene (Quail (1991) Ann. Rev. Genet. 25: 389-409) and light-dependent phyA protein turnover (Clough and Vierstra (1997) Plant Cell Environ. 20: 713-721). By contrast, the steady state levels of the phyB-E photoreceptors are not significantly regulated by the light enviroment (Sharrock and Clack (2002) Plant Physiol. 130: 442-456). Plant phytochromes are dimeric, with phyA predominantly occuring as homodimers while phyC-E polypeptides forming heterodimers with phyB (Sharrock and Clack (2004) Proc. Natl. Acad. Sci. USA 101: 11500-11505). PhyA has been established to be responsible for both VLFR and FR-HIR responses, while the light-stable phytochromes mediate the R/FR photoreversible LFR. Since phyA and phyB are the predominant forms of phytochrome found in light-grown plants (Hirschfeld et al. (1998) Genetics 149: 523-535), null mutants in these genes are deficient in VLFR/FR-HIR and LFR responses, respectively (Shinomura et al. (1996) Proc. Natl. Acad. Sci. USA 93: 8129-8133).
Accumulating to high levels in plants grown for prolonged periods in darkness, the light-labile phyA class of photoreceptors function to regulate seed germination and seedling development under very low light fluence rates (Casal et al. (1997) Environ. 20: 813-819). Such conditions are encountered for seeds that germinate and develop underground or in the FR-enriched shade of a forest canopy. Dark-grown seedlings accumulate elevated levels of the Pr from of phyA. When such seedlings are exposed to light, phyA is rapidly degraded (Nagy and Schäfer (2002) Annu Rev Plant Biol 53: 329-355). The rate of phyA degradation in oat seedlings increases 100-fold upon light exposure—from a half-life of over 100 h for the Pr form to less than 1 h after photoconversion to Pfr (Clough and Vierstra (1997) Plant Cell Environ. 20: 713-721). Light-dependent phyA turnover is preceeded by the formation of sequestered areas of phytochromes or SAPs in the cytoplasm (MacKenzie et al. (1974) Plant Physiol. 53: Abstract No. 5; Speth et al. (1987) Planta 171: 332-338). Since SAPs formed under R and white light (W) and can be detected in phyA preparations in vitro, the hypothesis that SAPs represent self-aggregated Pfr:Pfr homodimers has been proposed (Hofmann et al. (1991) Planta 183: 265-273; Hofmann et al. (1990) Planta 180: 372-377). This assertion has received support from more recent studies with Arabidopsis and tobacco seedlings showing that phyA must be translocated to the nucleus to initiate signal transduction (Nagy and Schäfer (2002) Annu Rev Plant Biol 53: 329-355; Chen et al. (2004) Ann. Rev. Genet. 38: 87-117). In this model, phyA that remains sequestered in the cytosol is eventually degraded and therefore does not participate in signaling. Pr:Pfr heterodimers of phyA that move to the nucleus presumably account for the VLFR and FR-HIR responses. PhyB signaling also involves light-dependent translocation to the nucleus; however, in contrast with phyA, this process can be reversed by FR illumination. Thus it is phyB that is the photoreceptor that is primarily responsible for sensing the R/FR ratio that triggers shade avoidance (Smith and Whitelam (1997) Plant Cell Environ. 20: 840-844; Morelli and Ruberti (2002) Trends in Plant Science 7: 399-404; Franklin and Whitelam (2005) Annal. Botany (London)).
Phytochrome-mediated shade avoidance responses observed in high density plantings leads to increased resource partitioning towards elongation growth, reduced lateral branching and early flowering at the expense of seed/fruit crop yield (Sawers et al. (2005) Trends in Plant Science 10: 138-143; Smith (1995) Ann. Rev. Plant Physiol. Plant Mol. Biol. 46: 289-315). While traditional plant breeding has successfully been used to reduce shade avoidance responses, this process is time-consuming in order to introduce only the necessary gene(s) into the desired cultivar. Genetic engineering would be a more rapid/direct way to accomplish this goal assuming that the desired genes are known. In this regard, transgenic expression of PHYA has already proven successful to reduce shade avoidance losses in agronomically important crop species (Smith (1992) Photochem. Photobiol. 56: 815-822; Smith: H. (1994) Sem. Cell Biol. 5: 315-325). This approach requires elevated expression of PHYA to produce a sufficient amount of phyA holoprotein to counteract the shade avoidance responses mediated by endogenous light-stable phytochromes (Robson et al. (1996) Nature Biotechnology 14: 995-998; Kong et al. (2004) Molecular Breeding 14: 35-45; Garg et al. (2005) Planta: 1-10).
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