Chlorophyll
a and chlorophyll
b are the major constituents of the photosynthetic apparatus in land plants and green algae. Chlorophyll
a is essential in photochemistry, while chlorophyll
b is ...apparently dispensable for their photosynthesis. Instead, chlorophyll
b is necessary for stabilizing the major light-harvesting chlorophyll-binding proteins. Chlorophyll
b is synthesized from chlorophyll
a and is catabolized after it is reconverted to chlorophyll
a. This interconversion system between chlorophyll
a and chlorophyll
b refers to the chlorophyll cycle. The chlorophyll
b levels are determined by the activity of the three enzymes participating in the chlorophyll cycle, namely, chlorophyllide
a oxygenase, chlorophyll
b reductase, and 7-hydroxymethyl-chlorophyll reductase. This article reviews the recent progress on the analysis of the chlorophyll cycle and its enzymes. In particular, we emphasize the impact of genetic modification of chlorophyll cycle enzymes on the construction and destruction of the photosynthetic machinery. These studies reveal that plants regulate the construction and destruction of a specific subset of light-harvesting complexes through the chlorophyll cycle. This article is part of a Special Issue entitled: Regulation of Electron Transport in Chloroplasts.
► Chlorophyll
b is essential in stabilizing light-harvesting complexes (LHC). ► Chlorophyll
b breakdown is requisite for the degradation of LHC. ► Interconversion of chlorophyll a and chlorophyll b refers to the chlorophyll cycle. ► Two out of three enzymes involved in the chlorophyll cycle have been identified. ► The regulatory mechanism of the chlorophyll cycle is partly understood.
Tetrapyrroles play vital roles in various biological processes, including photosynthesis and respiration. Higher plants contain four classes of tetrapyrroles, namely, chlorophyll, heme, siroheme, and ...phytochromobilin. All of the tetrapyrroles are derived from a common biosynthetic pathway. Here we review recent progress in the research of tetrapyrrole biosynthesis from a cellular biological view. The progress consists of biochemical, structural, and genetic analyses, which contribute to our understanding of how the flow and the synthesis of tetrapyrrole molecules are regulated and how the potentially toxic intermediates of tetrapyrrole synthesis are maintained at low levels. We also describe interactions of tetrapyrrole biosynthesis and other cellular processes including the stay-green events, the cell-death program, and the plastid-to-nucleus signal transduction. Finally, we present several reports on attempts for agricultural and horticultural applications in which the tetrapyrrole biosynthesis pathway was genetically modified.
Although seeds are a sink organ, chlorophyll synthesis and degradation occurs during embryogenesis and in a manner similar to that observed in photosynthetic leaves. Some mutants retain chlorophyll ...after seed maturation, and they are disturbed in seed storability. To elucidate the effects of chlorophyll retention on the seed storability of Arabidopsis (Arabidopsis thaliana), we examined the non-yellow coloring1 (nyc1)/nyc1-like (nol) mutants that do not degrade chlorophyll properly. Approximately 10 times more chlorophyll was retained in the dry seeds of the nyc1/nol mutant than in the wild-type seeds. The germination rates rapidly decreased during storage, with most of the mutant seeds failing to germinate after storage for 23 months, whereas 75% of the wild-type seeds germinated after 42 months. These results indicate that chlorophyll retention in the seeds affects seed longevity. Electron microscopic studies indicated that many small oil bodies appeared in the embryonic cotyledons of the nyc1/nol mutant; this finding indicates that the retention of chlorophyll affects the development of organelles in embryonic cells. A sequence analysis of the NYC1 promoter identified a potential abscisic acid (ABA)-responsive element. An electrophoretic mobility shift assay confirmed the binding of an ABA-responsive transcriptional factor to the NYC1 promoter DNA fragment, thus suggesting that NYC1 expression is regulated by ABA. Furthermore, NYC1 expression was repressed in the ABA-insensitive mutants during embryogenesis. These data indicate that chlorophyll degradation is induced by ABA during seed maturation to produce storable seeds.
The interconversion of chlorophyll a and chlorophyll b, referred to as the chlorophyll cycle, plays a crucial role in the processes of greening, acclimation to light intensity, and senescence. The ...chlorophyll cycle consists of three reactions: the conversions of chlorophyll a to chlorophyll b by chlorophyllide a oxygenase, chlorophyll b to 7-hydroxymethyl chlorophyll a by chlorophyll b reductase, and 7-hydroxymethyl chlorophyll a to chlorophyll a by 7-hydroxymethyl chlorophyll a reductase. We identified 7-hydroxymethyl chlorophyll a reductase, which is the last remaining unidentified enzyme of the chlorophyll cycle, from Arabidopsis thaliana by genetic and biochemical methods. Recombinant 7-hydroxymethyl chlorophyll a reductase converted 7-hydroxymethyl chlorophyll a to chlorophyll a using ferredoxin. Both sequence and biochemical analyses showed that 7-hydroxymethyl chlorophyll a reductase contains flavin adenine dinucleotide and an iron-sulfur center. In addition, a phylogenetic analysis elucidated the evolution of 7-hydroxymethyl chlorophyll a reductase from divinyl chlorophyllide vinyl reductase. A mutant lacking 7-hydroxymethyl chlorophyll a reductase was found to accumulate 7-hydroxymethyl chlorophyll a and pheophorbide a. Furthermore, this accumulation of pheophorbide a in the mutant was rescued by the inactivation of the chlorophyll b reductase gene. The downregulation of pheophorbide a oxygenase activity is discussed in relation to 7-hydroxymethyl chlorophyll a accumulation.
The light-harvesting chlorophyll a/b-protein complex of photosystem II (LHCII) is the most abundant membrane protein in green plants, and its degradation is a crucial process for the acclimation to ...high light conditions and for the recovery of nitrogen (N) and carbon (C) during senescence. However, the molecular mechanism of LHCII degradation is largely unknown. Here, we report that chlorophyll b reductase, which catalyzes the first step of chlorophyll b degradation, plays a central role in LHCII degradation. When the genes for chlorophyll b reductases NOL and NYC1 were disrupted in Arabidopsis thaliana, chlorophyll b and LHCII were not degraded during senescence, whereas other pigment complexes completely disappeared. When purified trimeric LHCII was incubated with recombinant chlorophyll b reductase (NOL), expressed in Escherichia coli, the chlorophyll b in LHCII was converted to 7-hydroxymethyl chlorophyll a. Accompanying this conversion, chlorophylls were released from LHCII apoproteins until all the chlorophyll molecules in LHCII dissociated from the complexes. Chlorophyll-depleted LHCII apoproteins did not dissociate into monomeric forms but remained in the trimeric form. Based on these results, we propose the novel hypothesis that chlorophyll b reductase catalyzes the initial step of LHCII degradation, and that trimeric LHCII is a substrate of LHCII degradation.
Abstract
The relationship between enzymes and substrates does not perfectly match the “lock and key” model, because enzymes act on molecules other than their true substrate in different catalytic ...reactions. Such biologically nonfunctional reactions are called “promiscuous activities.” Promiscuous activities are apparently useless, but they can be an important starting point for enzyme evolution. It has been hypothesized that enzymes with low promiscuous activity will show enhanced promiscuous activity under selection pressure and become new specialists through gene duplication. Although this is the prevailing scenario, there are two major problems: 1) it would not apply to prokaryotes because horizontal gene transfer is more significant than gene duplication and 2) there is no direct evidence that promiscuous activity is low without selection pressure. We propose a new scenario including various levels of promiscuous activity throughout a clade and horizontal gene transfer. STAY-GREEN (SGR), a chlorophyll a—Mg dechelating enzyme, has homologous genes in bacteria lacking chlorophyll. We found that some bacterial SGR homologs have much higher Mg-dechelating activities than those of green plant SGRs, while others have no activity, indicating that the level of promiscuous activity varies. A phylogenetic analysis suggests that a bacterial SGR homolog with high dechelating activity was horizontally transferred to a photosynthetic eukaryote. Some SGR homologs acted on various chlorophyll molecules that are not used as substrates by green plant SGRs, indicating that SGR acquired substrate specificity after transfer to eukaryotes. We propose that horizontal transfer of high promiscuous activity is one process of new enzyme acquisition.
The light-harvesting complex (LHC) constitutes the major light-harvesting antenna of photosynthetic eukaryotes. LHC contains a characteristic sequence motif, termed LHC motif, consisting of 25–30 ...mostly hydrophobic amino acids. This motif is shared by a number of transmembrane proteins from oxygenic photoautotrophs that are termed light-harvesting-like (LIL) proteins. To gain insights into the functions of LIL proteins and their LHC motifs, we functionally characterized a plant LIL protein, LIL3. This protein has been shown previously to stabilize geranylgeranyl reductase (GGR), a key enzyme in phytol biosynthesis. It is hypothesized that LIL3 functions to anchor GGR to membranes. First, we conjugated the transmembrane domain of LIL3 or that of ascorbate peroxidase to GGR and expressed these chimeric proteins in an Arabidopsis mutant lacking LIL3 protein. As a result, the transgenic plants restored phytol-synthesizing activity. These results indicate that GGR is active as long as it is anchored to membranes, even in the absence of LIL3. Subsequently, we addressed the question why the LHC motif is conserved in the LIL3 sequences. We modified the transmembrane domain of LIL3, which contains the LHC motif, by substituting its conserved amino acids (Glu-171, Asn-174, and Asp-189) with alanine. As a result, the Arabidopsis transgenic plants partly recovered the phytol-biosynthesizing activity. However, in these transgenic plants, the LIL3-GGR complexes were partially dissociated. Collectively, these results indicate that the LHC motif of LIL3 is involved in the complex formation of LIL3 and GGR, which might contribute to the GGR reaction.
The light-harvesting complex (LHC) motif is an amino acid consensus sequence found in various thylakoid proteins.
Modification and replacement of the transmembrane domain encompassing the LHC motif of a plant protein retains its membrane-anchoring function but impairs its protein-protein interaction.
This domain functions in membrane anchoring and complex formation.
This study provides new insights regarding the function of the LHC motif.
In the chlorophyll biosynthesis pathway, the 8-vinyl group of the chlorophyll precursor is reduced to an ethyl group by 8-vinyl reductase. Two isozymes of 8-vinyl reductase have been described in ...oxygenic photosynthetic organisms: one encoded by
BciA
and another by
BciB
. Only BciB contains an Fe-S cluster and most cyanobacteria harbor this form; whereas a few contain
BciA
. Given this disparity in distribution, cyanobacterial BciA has remained largely overlooked, which has limited understanding of chlorophyll biosynthesis in these microorganisms. Here, we reveal that cyanobacterial
BciA
encodes a functional 8-vinyl reductase, as evidenced by measuring the in vitro activity of recombinant
Synechococcus
and
Acaryochloris
BciA. Genomic comparison revealed that
BciB
had been replaced by
BciA
during evolution of the marine cyanobacterium
Synechococcus
, and coincided with replacement of Fe-superoxide dismutase (SOD) with Ni-SOD. These findings imply that the acquisition of
BciA
confers an adaptive advantage to cyanobacteria living in low-iron oceanic environments.
The plastid plays a vital role in various cellular activities within plant cells including photosynthesis and other metabolic pathways. It is believed that the functional status of the plastid is ...somehow monitored by the nucleus to optimize the expression of genes encoding plastid proteins. The currently dominant model for plastid-derived signaling ("plastid signaling") proposes that Mg-protoporphyrin IX (MgProto) is a negative signal that represses the expression of a wide range of nuclear genes encoding plastid-localized proteins when plastid development is inhibited. In this study, we have re-evaluated this hypothesis by quantifying the steady-state levels of MgProto (as well as its neighboring intermediates protoporphyrin IX and Mg-Proto monomethyl ester MgProtoMe) in Arabidopsis plants with altered plastid signaling responses as monitored by expression of the Lhcb1, RBCS, HEMA1, BAM3 and CA1 genes. In addition, we have examined the correlation between gene expression and MgProto (MgProtoMe) in a range of mutants and conditions in which the steady-state levels of MgProto (MgProtoMe) have been modified. Overall we found that there was no correlation between the steady-state levels of MgProto (MgProtoMe) and Lhcb1 expression or with any of the other genes tested. Taking these results together, we propose that the current model on plastid signaling must be revised.
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