Plant Phytochromes and their Phosphorylation

Abstract

Extensive research over several decades in plant light signaling mediated by photoreceptors has identified the molecular mechanisms for how phytochromes regulate photomorphogenic development, which includes degradation of phytochrome-interacting factors (PIFs) and inactivation of COP1-SPA complexes with the accumulation of master transcription factors for photomorphogenesis, such as HY5. However, the initial biochemical mechanism for the function of phytochromes has not been fully elucidated. Plant phytochromes have long been known as phosphoproteins, and a few protein phosphatases that directly interact with and dephosphorylate phytochromes have been identified. However, there is no report thus far of a protein kinase that acts on phytochromes. On the other hand, plant phytochromes have been suggested as autophosphorylating serine/threonine protein kinases, proposing that the kinase activity might be important for their functions. Indeed, the autophosphorylation of phytochromes has been reported to play an important role in the regulation of plant light signaling. More recently, evidence that phytochromes function as protein kinases in plant light signaling has been provided using phytochrome mutants displaying reduced kinase activities. In this review, we highlight recent advances in the reversible phosphorylation of phytochromes and their functions as protein kinases in plant light signaling.

Keywords: autophosphorylation; light signaling; plant photoreceptors; protein kinase; reversible phosphorylation.

Figure 1

A representative domain structure of plant phytochromes. The N- and C-termini of protein are indicated, and a chromophore is covalently attached to a cysteine residue in the cGMP phosphodiesterase/adenylyl cyclase/FhlA (GAF) domain. The photosensory module (PSM) consists of N-terminal extension (NTE) and the photosensory core (PAS/GAF/PHY), and the output module (OPM) contain PAS-related domain (PRD) with a pair of PAS repeats (labeled as PAS-A and PAS-B) and histidine kinase-related domain (HKRD). The PSM and OPM are linked with each other by a hinge region, and the knot lasso motif in GAF domain and the hairpin motif in the phytochrome-specific (PHY) domain are indicated by the orange and green loops, respectively. The phosphorylation sites of plant phytochromes have been reported in the NTE and hinge regions (see below).

Figure 2

A simplified view of the phytochrome-mediated photomorphogenesis in A. thaliana. For simplicity, we use PIF3 as a representative of PIFs, HY5 as a representative of the master transcription factors of photomorphogenesis, and SPA1 as a representative of the SPA proteins. In the dark (left panel), phytochromes are biosynthesized as the inactive Pr forms, staying in the cytoplasm. Meanwhile, PIF3 proteins accumulate in the nucleus and regulate the expression of genes to prevent photomorphogenesis (shown as a blue T bar) by promoting skotomorphogenesis (shown as a red arrow). In addition, the COP1-SPA1 complex constantly degrade the expressed HY5 proteins via the ubiquitin 26S proteasome pathway to prevent photomorphogenesis (shown as a dotted black arrow). In the light (right panel), the photoactivated Pfr forms of phytochromes translocate into the nucleus (in the case of phyA, FHY1 and FHL are the facilitators for the import), where they can interact with downstream signaling components. They inactivate PIF3 by inducing protein degradation and the COP1-SPA1 complex by inducing its dissociation and subsequent nuclear exclusion of COP1, which all contribute to the accumulation of the master transcription factors for photomorphogenesis, such as HY5. Finally, HY5 induces the expression of light-responsive genes for photomorphogenic development. Movements of proteins are also shown as dotted arrows.

Figure 3

A proposed model to explain the molecular mechanisms for phyA signaling under a light condition. PrA, the Pr form of phyA; PfrA, the Pfr form of phyA; 26S, the 26S proteasome complex; P, phosphate; PK, unknown protein kinase(s). As a protein kinase, PfrA is autophosphorylated (AutoP) and phosphorylates substrate proteins such as PIF3. So far, PIF3 is also known to be phosphorylated by PPKs and BIN2, and COP1 is phosphorylated by PID. However, the protein kinase(s) that can phosphorylate phytochromes are unknown (labeled as “PK?”). In addition, it is not known how FHY1 and FHL can be phosphorylated (i.e., another label with “PK?”). Moreover, it is possible that PIF3 and COP1 might be phosphorylated further by unknown protein kinase(s). As shown in this model, phosphorylation (shown as red arrows) takes significant portions in the phytochrome signaling in plants. In summary, the function of phytochromes is to inactivate the negative regulators for photomorphogenesis such as PIF3 and COP1-SPA1 complex, which eventually express and stabilize HY5 to initiate photomorphogenic development. At the same time, phosphorylated PfrP is rapidly degraded via the ubiquitin 26S proteasome pathway (shown as green arrows) for an efficient desensitization of the phyA signal, which is necessary for responses to subsequent changes in fluctuating light environments.