LOV domain-containing F-box proteins: light-dependent protein degradation modules in Arabidopsis

Abstract

Plants constantly survey the surrounding environment using several sets of photoreceptors. They can sense changes in the quantity (=intensity) and quality (=wavelength) of light and use this information to adjust their physiological responses, growth, and developmental patterns. In addition to the classical photoreceptors, such as phytochromes, cryptochromes, and phototropins, ZEITLUPE (ZTL), FLAVIN-BINDING, KELCH REPEAT, F-BOX 1 (FKF1), and LOV KELCH PROTEIN 2 (LKP2) proteins have been recently identified as blue-light photoreceptors that are important for regulation of the circadian clock and photoperiodic flowering. The ZTL/FKF1/LKP2 protein family possesses a unique combination of domains: a blue-light-absorbing LOV (Light, Oxygen, or Voltage) domain along with domains involved in protein degradation. Here, we summarize recent advances in our understanding of the function of the Arabidopsis ZTL/FKF1/LKP2 proteins. We summarize the distinct photochemical properties of their LOV domains and discuss the molecular mechanisms by which the ZTL/FKF1/LKP2 proteins regulate the circadian clock and photoperiodic flowering by controlling blue-light-dependent protein degradation.

Figure 1.

The Domain Structures of ZTL/FKF1/LKP2 and Related Proteins, Amino Acid Sequence Alignments of LOV Domains, and Photochemical Properties of Phototropin and FKF1 LOV Domains. (A) Schematic illustration of functional domains of ZTL/FKF1/LKP2 family proteins, phototropin proteins, and PAS/LOV proteins. LOV domain bound to an FMN molecule functions as a blue-light-sensing domain. The ZTL/FKF1/LKP2 family proteins possess one LOV domain at the N-terminus region followed by an F-box domain and six Kelch repeats in the C-terminal region. The phototropins contain two FMN-binding LOV domains in their N-terminal region (LOV1 and LOV2) and a serine/threonine kinase domain at the C-terminus. The PAS/LOV proteins (PLP or also called LLP) contain two LOV domains; however, the cysteine residue essential for the cysteinyl adduct formation within the first LOV domain is not always conserved in some plant species, including Arabidopsis (Kasahara et al., 2010), indicating that there is no light-induced photocycle of the LOV domain. The LOV2 domains of PLP proteins usually contain the conserved cysteine and show the blue-light-induced photocycle in vitro. (B) Sequence alignment of several LOV domains. The alignment includes Arabidopsis thaliana ZTL, LKP2, FKF1, phot1-LOV1, phot1-LOV2, phot2-LOV1, and phot2-LOV2. Identical and similar amino acids are marked on top of the alignment by asterisks (*) and (: or .), respectively. The conserved cysteines for FMN binding are highlighted in orange. The characteristic loop regions of ZTL/FKF1/LKP2 LOV are gray-shaded. The predicted secondary structure elements are shown on top of the alignment. Arrows and boxes indicate β-strands and α-helices, respectively. (C) Schematic representation of LOV-domain photochemistry. In darkness, the FMN chromophore is non-covalently bound in the LOV domain (L450). Light triggers the production of a reactive triplet-state flavin (L660) that leads to formation of a covalent bond between the FMN and a conserved cysteine residue in the LOV domain (S390). The photoreaction process of the phot-LOV protein (right blue arrow circuit) is fully reversible in the dark. In contrast, the FKF1–LOV protein (a black dotted arrow) shows a slow dark reversion rate. The dark reversion rate (half-lives; t_1/2) of each protein is shown.

Figure 2.

Tentative Structure Changes of FKF1 Triggered by Blue-Light Absorption. (A) Proposed structural changes of phot1. Two phot1 molecules dimerize through their LOV1 domains. The LOV2 domain is important for light-dependent function. Once phot1 absorbs blue light, the Jα-helix located adjacent to the LOV2 domain detaches from the LOV2 domain. This structural change is thought to activate the C-terminal kinase. The kinase phosphorylates N-terminal regions as well as substrate proteins to send phosphorylation signals. (B) FKF1 also forms at least a homodimer with anti-parallel configuration in vitro. In the dark, FKF1 may be incorporated in the SCF^FKF1 complex (which consists of FKF1 F-box protein, ASK, Cullin, and Rbx). After blue-light exposure, FKF1 interacts with GI through the LOV domain. Light may cause a similar structural change to phot1 to enable FKF1 to bind to GI. GI also binds to CDF1, a substrate of FKF1 for ubiquitination-dependent protein degradation. FKF1 binds to CDF1 through the Kelch repeat domain and is involved in protein degradation.

Figure 3.

Light-Mediated Proteolysis Controlled by ZTL, FKF1, and LKP2 in the Regulation of Flowering Time and the Circadian Clock. (A) The function of FKF1 in the control of flowering time. GI forms protein complexes with CDFs on the CO promoter. In the afternoon, FKF1 is recruited to the CO promoter region through blue-light-dependent interaction with GI. FKF1 degrades CDFs, facilitating expression of CO. Since CDF2 is stabilized further in the ztl fkf1 lkp2 triple mutants than in the fkf1 mutants, the ZTL–GI and LKP2–GI complexes may also contribute to the degradation of some CDF proteins. CO and GI bind to the FT promoter and activate FT transcription, which induces flowering. FKF1 may also bind to the FT promoter; however, the function of this interaction is unknown. (B) The function of ZTL in regulation of the circadian clock. Under blue light, ZTL interacts with GI. ZTL also forms a protein complex with HSP90. The interaction of ZTL with GI and HSP90 stabilizes ZTL protein. The ZTL–GI complex formation may sequester ZTL from interaction with TOC1 or PRR5. This enables TOC1 and PRR5 proteins to accumulate in the late afternoon. The FKF1–GI and LKP2–GI complex may also have a similar role (left side). In the dark, ZTL (FKF1 and LKP2) interact with TOC1 and PRR5 proteins and degrade them through a proteasome pathway (right side).