Drought tolerance is important for grain crops, including rice (Oryza sativa); for example, rice cultivated under intermittent irrigation produces less methane gas compared to rice grown in anaerobic ...paddy field conditions, but these plants require greater drought tolerance. Moreover, the roles of rice circadian‐clock genes in drought tolerance remain largely unknown. Here, we show that the mutation of LOV KELCH REPEAT PROTEIN 2 (OsLKP2) enhanced drought tolerance by increasing cuticular wax biosynthesis. Among ZEITLUPE family genes, OsLKP2 expression specifically increased under dehydration stress. OsLKP2 knockdown (oslkp2‐1) and knockout (oslkp2‐2) mutants exhibited enhanced drought tolerance. Cuticular waxes inhibit non‐stomatal water loss. Under drought conditions, total wax loads on the leaf surface increased by approximately 10% in oslkp2‐1 and oslkp2‐2 compared to the wild type, and the transcript levels of cuticular wax biosynthesis genes were upregulated in the oslkp2 mutants. Yeast two‐hybrid, bimolecular fluorescence complementation, and coimmunoprecipitation assays revealed that OsLKP2 interacts with GIGANTEA (OsGI) in the nucleus. The osgi mutants also showed enhanced tolerance to drought stress, with a high density of wax crystals on their leaf surface. These results demonstrate that the OsLKP2‐OsGI interaction negatively regulates wax accumulation on leaf surfaces, thereby decreasing rice resilience to drought stress.
Summary statement
Loss of LOV KELCH REPEAT PROTEIN 2 (OsLKP2) function results in drought tolerance in rice due to an elevated total wax load on leaf surfaces through upregulated expression of wax‐related genes. In the nucleus, OsLKP2 interacts with GIGANTEA, whose null mutant also shows enhanced drought tolerance.
In the marine α-proteobacterium
more than 40 genes of the aerobic anoxygenic photosynthesis are regulated in a light-dependent manner. A genome-wide screen of 5,605 clones from a
transposon library ...for loss of pigmentation and changes in bacteriochlorophyll absorbance identified 179 mutant clones. The gene encoding the LOV-domain containing protein Dshi_1135 was identified by its colorless phenotype. The mutant phenotype was complemented by the expression of a Dshi_1135-strep fusion protein in trans. The recombinantly produced and chromatographically purified Dshi_1135 protein was able to undergo a blue light-induced photocycle mediated by bound FMN. Transcriptome analyses revealed an essential role for Dshi_1135 in the light-dependent expression of the photosynthetic gene cluster. Interactomic studies identified the repressor protein PpsR as an interaction partner of Dshi_1135. The physical contact between PpsR and the Dshi_1135 protein was verified
using the bacterial adenylate cyclase-based two-hybrid system. In addition, the antirepressor function of the Dshi_1135 protein was demonstrated
testing of a
reporter gene fusion in a heterologous
-based host system. We therefore propose to rename the Dshi_1135 protein to LdaP (light-dependent antirepressor of PpsR). Using the bacterial two-hybrid system, it was also shown that cobalamin (B
) is essential for the interaction of the antirepressor PpaA with PpsR. A regulatory model for the photosynthetic gene cluster in
was derived, including the repressor PpsR, the light-dependent antirepressor LdaP and the B
-dependent antirepressor PpaA.
Photoreceptor flavoproteins of the LOV, BLUF, and cryptochrome families are ubiquitous among the three domains of life and are configured as UVA/blue-light systems not only in plants-their original ...arena-but also in prokaryotes and microscopic algae. Here, we review these proteins' structure and function, their biological roles, and their evolution and impact in the living world, and underline their growing application in biotechnologies. We present novel developments such as the interplay of light and redox stimuli, emerging enzymatic and biological functions, lessons on evolution from picoalgae, metagenomics analysis, and optogenetics applications.
Methods to acutely manipulate protein interactions at the subcellular level are powerful tools in cell biology. Several blue-light-dependent optical dimerization tools have been developed. In these ...systems one protein component of the dimer (the bait) is directed to a specific subcellular location, while the other component (the prey) is fused to the protein of interest. Upon illumination, binding of the prey to the bait results in its subcellular redistribution. Here, we compared and quantified the extent of light-dependent dimer occurrence in small, subcellular volumes controlled by three such tools: Cry2/CIB1, iLID, and Magnets. We show that both the location of the photoreceptor protein(s) in the dimer pair and its (their) switch-off kinetics determine the subcellular volume where dimer formation occurs and the amount of protein recruited in the illuminated volume. Efficient spatial confinement of dimer to the area of illumination is achieved when the photosensitive component of the dimerization pair is tethered to the membrane of intracellular compartments and when on and off kinetics are extremely fast, as achieved with iLID or Magnets. Magnets and the iLID variants with the fastest switch-off kinetics induce and maintain protein dimerization in the smallest volume, although this comes at the expense of the total amount of dimer. These findings highlight the distinct features of different optical dimerization systems and will be useful guides in the choice of tools for specific applications.
Bacteria and fungi of the plant microbiota can be phytopathogens, parasites or symbionts that establish mutually advantageous relationships with plants. They are often rich in photoreceptors for ...UVA–Visible light, and in many cases, they exhibit light regulation of growth patterns, infectivity or virulence, reproductive traits, and production of pigments and of metabolites. In addition to the light-driven effects, often demonstrated via the generation of photoreceptor gene knock-outs, microbial photoreceptors can exert effects also in the dark. Interestingly, some fungi switch their attitude towards plants in dependence of illumination or dark conditions in as much as they may be symbiotic or pathogenic. This review summarizes the current knowledge about the roles of light and photoreceptors in plant-associated bacteria and fungi aiming at the identification of common traits and general working ideas. Still, reports on light-driven infection of plants are often restricted to the description of macroscopically observable phenomena, whereas detailed information on the molecular level, e.g., protein–protein interaction during signal transduction or induction mechanisms of infectivity/virulence initiation remains sparse. As it becomes apparent from still only few molecular studies, photoreceptors, often from the red- and the blue light sensitive groups interact and mutually modulate their individual effects. The topic is of great relevance, even in economic terms, referring to plant-pathogen or plant-symbionts interactions, considering the increasing usage of artificial illumination in greenhouses, the possible light-regulation of the synthesis of plant-growth stimulating substances or herbicides by certain symbionts, and the biocontrol of pests by selected fungi and bacteria in a sustainable agriculture.
In Arabidopsis, GIGANTEA (GI), together with the blue-light receptors ZTL, LKP2, and FKF1, regulates degradation of the core clock protein TOC1 and the flowering repressor CDFs, thereby controlling ...circadian oscillation and flowering. Despite the significance of GI in diverse plant physiology, its molecular function is not much understood because of technical problems in protein preparation and a lack of structural information. Here, we report the purification of the GI monomer and the crystal structure of the GI/LKP2 complex. The crystal structure reveals that residues 1–813 of GI possess an elongated rigid structure formed by stacking hydrophobic α-helices and that the LOV domain of LKP2 binds to the middle region of the GI (residues 563–789). Interaction analysis further shows that LOV homodimers are converted to monomers by GI binding. Our results provide structural insights into the regulation of the circadian clock and photoperiodic flowering by GI and ZTL/LKP2/FKF1.
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•GI possesses an elongated rigid structure formed by stacking hydrophobic α-helices•GI forms a heterodimer with the LOV domains of ZTL, LKP2, and FKF1•The LOV domain binds to the middle region of GI at residues 563–789•LOV homodimer is disrupted by GI binding
GIGANTEA (GI) and blue-light receptors contribute to plant circadian rhythm and flowering. Kwon et al. describes the crystal structure of GI in complex with the LOV domain of LKP2 from Arabidopsis thaliana. This reveals the overall structure of GI and its interactions with the LOV domains.
Light-oxygen-voltage (LOV) domains are widespread photosensory modules that can be used in fluorescence microscopy, optogenetics and controlled production of reactive oxygen species. All of the ...currently known LOV domains have absorption maxima in the range of ~440 to ~450 nm, and it is not clear whether they can be shifted significantly using mutations. Here, we have generated a panel of LOV domain variants by mutating the key chromophore-proximal glutamine aminoacid of a thermostable flavin based fluorescent protein CagFbFP (Gln148) to asparagine, aspartate, glutamate, histidine, lysine and arginine. Absorption spectra of all of the mutants are blue-shifted, with the maximal shift of 8 nm observed for the Q148H variant. While CagFbFP and its Q148N/D/E variants are not sensitive to pH, Q148H/K/R reveal a moderate red shift induced byacidic pH. To gain further insight, we determined high resolution crystal structures of all of the mutants studied at the resolutions from 1.07 Å for Q148D to 1.63 Å for Q148R. Whereas in some of the variants, the aminoacid 148 remains in the vicinity of the flavin, in Q148K, Q148R and partially Q148D, the C-terminus of the protein unlatches and the side chain of the residue 148 is reoriented away from the chromophore. Our results explain the absence of color shifts from replacing Gln148 with charged aminoacids and pave the way for rational design of color-shifted flavin based fluorescent proteins.
Phototropins are blue-light receptors controlling a range of responses that serve to optimize the photosynthetic efficiency of plants. These include phototropism, light-induced stomatal opening, and ...chloroplast movements in response to changes in light intensity. Since the isolation of the Arabidopsis PHOT1 gene in 1997, phototropins have been identified in ferns and mosses where their physiological functions appear to be conserved. Arabidopsis contains two phototropins, phot1 and phot2, that exhibit overlapping functions in addition to having unique physiological roles. Phototropins are light-activated serine/threonine protein kinases. Light sensing by the phototropins is mediated by a repeated motif at the N-terminal region of the protein known as the LOV domain. Photoexcitation of the LOV domain results in receptor autophosphorylation and an initiation of phototropin signaling. Here we summarize the photochemical and biochemical events underlying phototropin activation in addition to the current knowledge of the molecular mechanisms associated with photoreceptor signaling.
The mechanism of light-triggered conformational change and signaling in light-oxygen-voltage (LOV) domains remains elusive in spite of extensive investigation and their use in optogenetic studies. ...The LOV2 domain of Avenasativa phototropin 1 (AsLOV2), a member of the Per-Arnt-Sim (PAS) family, contains a flavin mononucleotide chromophore that forms a covalent bond with a cysteine upon illumination. This event leads to the release of the carboxy-terminal Jα helix, the biological output signal. Using mutational analysis, circular dichroism, and NMR, we find that the largely ignored amino-terminal helix is a control element in AsLOV2's light-activated conformational change. We further identify a direct amino-to-carboxy-terminal “input–output” signaling pathway. These findings provide a framework to rationalize the LOV domain architecture, as well as the signaling mechanisms in both isolated and tandem arrangements of PAS domains. This knowledge can be applied in engineering LOV-based photoswitches, opening up new design strategies and improving existing ones.
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► We investigate the mechanism of light-triggered conformational changes in AsLOV2. ► Spectroscopy indicates that the N-terminal helix unfolds upon illumination. ► This event triggers the unfolding of the C-terminal Jα helix. ► Results can be used in other LOV-based photoswitches and new designs strategies. ► An N- to C-terminal signaling mechanism is possible in other PAS domains.
Ferromagnesian chondrules present a remarkable dichotomy between reduced (type I) and oxidized (type II) varieties. How these formed, and how they may be related remains contentious. Many type II ...chondrules, especially in carbonaceous chondrites, contain forsteritic grains in disequilibrium with FeO-rich host olivine grains, which must be relicts of precursor material. In this study, we analyzed the oxygen isotopic composition of magnesian relict and host olivine grains in type II chondrules in CO and CR chondrites. The analyzed Mg-rich relicts are generally more 16O-rich than ferroan olivine (mostly host) grains and plot in the range (in term of chemistry and isotopic composition) of type I chondrules in carbonaceous chondrites. Remarkably, they tend to cluster around the dominant Δ17O peaks of the type I chondrules in their host chondrites, viz. –6 ‰ and –2 ‰ for CO and CR, respectively. With the occurrence of relatively intact type I chondrules within some type II chondrules, this corroborates that local type I chondrules were among the precursors of type II chondrules, and that chondrule formation occurred within the accretion reservoir of the eventual chondrites. This supports the nebular brand of chondrule-forming scenarios. Since not all previous generations of chondrules (or other precursor objects) have been recycled, chondrule formation events must also have been extremely localized.
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