Rubisco evolution in C4 eudicots: an analysis of Amaranthaceae sensu lato

BACKGROUND Rubisco (ribulose-1,5-bisphosphate carboxylase/oxygenase) catalyses the key reaction in the photosynthetic assimilation of CO₂. In C₄ plants CO₂ is supplied to Rubisco by an auxiliary CO₂-concentrating pathway that helps to maximize the carboxylase activity of the enzyme while suppressing its oxygenase activity. As a consequence, C₄ Rubisco exhibits a higher maximum velocity but lower substrate specificity compared with the C₃ enzyme. Specific amino-acids in Rubisco are associated with C₄ photosynthesis in monocots, but it is not known whether selection has acted on Rubisco in a similar way in eudicots. METHODOLOGY/PRINCIPAL FINDINGS We investigated Rubisco evolution in Amaranthaceae sensu lato (including Chenopodiaceae), the third-largest family of C₄ plants, using phylogeny-based maximum likelihood and Bayesian methods to detect Darwinian selection on the chloroplast rbcL gene in a sample of 179 species. Two Rubisco residues, 281 and 309, were found to be under positive selection in C₄ Amaranthaceae with multiple parallel replacements of alanine by serine at position 281 and methionine by isoleucine at position 309. Remarkably, both amino-acids have been detected in other C₄ plant groups, such as C₄ monocots, illustrating a striking parallelism in molecular evolution. CONCLUSIONS/SIGNIFICANCE Our findings illustrate how simple genetic changes can contribute to the evolution of photosynthesis and strengthen the hypothesis that parallel amino-acid replacements are associated with adaptive changes in Rubisco.This research was funded by NERC (http://www.nerc.ac.uk/; grant number NE/H007741/1)

Widespread positive selection in the photosynthetic Rubisco enzyme

Background: Rubisco enzyme catalyzes the first step in net photosynthetic CO2 assimilation and photorespiratory carbon oxidation and is responsible for almost all carbon fixation on Earth. The large subunit of Rubisco is encoded by the chloroplast rbcL gene, which is widely used for reconstruction of plant phylogenies due to its conservative nature. Plant systematicists have mainly used rbcL paying little attention to its function, and the question whether it evolves under Darwinian selection has received little attention. The purpose of our study was to evaluate how common is positive selection in Rubisco among the phototrophs and where in the Rubisco structure does positive selection occur. Results: We searched for positive selection in rbcL sequences from over 3000 species representing all lineages of green plants and some lineages of other phototrophs, such as brown and red algae, diatoms, euglenids and cyanobacteria. Our molecular phylogenetic analysis found the presence of positive selection in rbcL of most analyzed land plants, but not in algae and cyanobacteria. The mapping of the positively selected residues on the Rubisco tertiary structure revealed that they are located in regions important for dimer-dimer, intradimer, large subunit-small subunit and Rubisco-Rubisco activase interactions, and that some of the positively selected residues are close to the active site. Conclusion: Our results demonstrate that despite its conservative nature, Rubisco evolves under positive selection in most lineages of land plants, and after billions of years of evolution Darwinian selection still fine-tunes its performance. Widespread positive selection in rbcL has to be taken into account when this gene is used for phylogenetic reconstructions. </p

Molecular Adaptation during Adaptive Radiation in the Hawaiian Endemic Genus Schiedea

BACKGROUND: “Explosive” adaptive radiations on islands remain one of the most puzzling evolutionary phenomena. The rate of phenotypic and ecological adaptations is extremely fast during such events, suggesting that many genes may be under fairly strong selection. However, no evidence for adaptation at the level of protein coding genes was found, so it has been suggested that selection may work mainly on regulatory elements. Here we report the first evidence that positive selection does operate at the level of protein coding genes during rapid adaptive radiations. We studied molecular adaptation in Hawaiian endemic plant genus Schiedea (Caryophyllaceae), which includes closely related species with a striking range of morphological and ecological forms, varying from rainforest vines to woody shrubs growing in desert-like conditions on cliffs. Given the remarkable difference in photosynthetic performance between Schiedea species from different habitats, we focused on the “photosynthetic” Rubisco enzyme, the efficiency of which is known to be a limiting step in plant photosynthesis. RESULTS: We demonstrate that the chloroplast rbcL gene, encoding the large subunit of Rubisco enzyme, evolved under strong positive selection in Schiedea. Adaptive amino acid changes occurred in functionally important regions of Rubisco that interact with Rubisco activase, a chaperone which promotes and maintains the catalytic activity of Rubisco. Interestingly, positive selection acting on the rbcL might have caused favorable cytotypes to spread across several Schiedea species. SIGNIFICANCE: We report the first evidence for adaptive changes at the DNA and protein sequence level that may have been associated with the evolution of photosynthetic performance and colonization of new habitats during a recent adaptive radiation in an island plant genus. This illustrates how small changes at the molecular level may change ecological species performance and helps us to understand the molecular bases of extremely fast rate of adaptation during island adaptive radiations

Predicting plant Rubisco kinetics from RbcL sequence data using machine learning

Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is responsible for the conversion of atmospheric CO2 to organic carbon during photosynthesis, and often acts as a rate limiting step in the later process. Screening the natural diversity of Rubisco kinetics is the main strategy used to find better Rubisco enzymes for crop engineering efforts. Here, we demonstrate the use of Gaussian processes (GPs), a family of Bayesian models, coupled with protein encoding schemes, for predicting Rubisco kinetics from Rubisco large subunit (RbcL) sequence data. GPs trained on published experimentally obtained Rubisco kinetic datasets were applied to over 9000 sequences encoding RbcL to predict Rubisco kinetic parameters. Notably, our predicted kinetic values were in agreement with known trends, e.g. higher carboxylation turnover rates (Kcat) for Rubisco enzymes from C4 or crassulacean acid metabolism (CAM) species, compared with those found in C3 species. This is the first study demonstrating machine learning approaches as a tool for screening and predicting Rubisco kinetics, which could be applied to other enzymes

Rubisco evolution in C₄ eudicots: an analysis of Amaranthaceae sensu lato.

BACKGROUND: Rubisco (ribulose-1,5-bisphosphate carboxylase/oxygenase) catalyses the key reaction in the photosynthetic assimilation of CO₂. In C₄ plants CO₂ is supplied to Rubisco by an auxiliary CO₂-concentrating pathway that helps to maximize the carboxylase activity of the enzyme while suppressing its oxygenase activity. As a consequence, C₄ Rubisco exhibits a higher maximum velocity but lower substrate specificity compared with the C₃ enzyme. Specific amino-acids in Rubisco are associated with C₄ photosynthesis in monocots, but it is not known whether selection has acted on Rubisco in a similar way in eudicots. METHODOLOGY/PRINCIPAL FINDINGS: We investigated Rubisco evolution in Amaranthaceae sensu lato (including Chenopodiaceae), the third-largest family of C₄ plants, using phylogeny-based maximum likelihood and Bayesian methods to detect Darwinian selection on the chloroplast rbcL gene in a sample of 179 species. Two Rubisco residues, 281 and 309, were found to be under positive selection in C₄ Amaranthaceae with multiple parallel replacements of alanine by serine at position 281 and methionine by isoleucine at position 309. Remarkably, both amino-acids have been detected in other C₄ plant groups, such as C₄ monocots, illustrating a striking parallelism in molecular evolution. CONCLUSIONS/SIGNIFICANCE: Our findings illustrate how simple genetic changes can contribute to the evolution of photosynthesis and strengthen the hypothesis that parallel amino-acid replacements are associated with adaptive changes in Rubisco

Coevolution of amino acid residues in the key photosynthetic enzyme Rubisco

<p>Abstract</p> <p>Background</p> <p>One of the key forces shaping proteins is coevolution of amino acid residues. Knowing which residues coevolve in a particular protein may facilitate our understanding of protein evolution, structure and function, and help to identify substitutions that may lead to desired changes in enzyme kinetics. Rubisco, the most abundant enzyme in biosphere, plays an essential role in the process of carbon fixation through photosynthesis, thus facilitating life on Earth. This makes Rubisco an important model system for studying the dynamics of protein fitness optimization on the evolutionary landscape. In this study we investigated the selective and coevolutionary forces acting on large subunit of land plants Rubisco using Markov models of codon substitution and clustering approaches applied to amino acid substitution histories.</p> <p>Results</p> <p>We found that both selection and coevolution shape Rubisco, and that positively selected and coevolving residues have their specifically favored amino acid composition and pairing preference. The mapping of these residues on the known Rubisco tertiary structures showed that the coevolving residues tend to be in closer proximity with each other compared to the background, while positively selected residues tend to be further away from each other. This study also reveals that the residues under positive selection or coevolutionary force are located within functionally important regions and that some residues are targets of both positive selection and coevolution at the same time.</p> <p>Conclusion</p> <p>Our results demonstrate that coevolution of residues is common in Rubisco of land plants and that there is an overlap between coevolving and positively selected residues. Knowledge of which Rubisco residues are coevolving and positively selected could be used for further work on structural modeling and identification of substitutions that may be changed in order to improve efficiency of this important enzyme in crops.</p

Positively selected amino acid replacements within the RuBisCO enzyme of oak trees are associated with ecological adaptations

Phylogenetic analysis by maximum likelihood (PAML) has become the standard approach to study positive selection at the molecular level, but other methods may provide complementary ways to identify amino acid replacements associated with particular conditions. Here, we compare results of the decision tree (DT) model method with ones of PAML using the key photosynthetic enzyme RuBisCO as a model system to study molecular adaptation to particular ecological conditions in oaks (Quercus). We sequenced the chloroplast rbcL gene encoding RuBisCO large subunit in 158 Quercus species, covering about a third of the global genus diversity. It has been hypothesized that RuBisCO has evolved differentially depending on the environmental conditions and leaf traits governing internal gas diffusion patterns. Here, we show, using PAML, that amino acid replacements at the residue positions 95, 145, 251, 262 and 328 of the RuBisCO large subunit have been the subject of positive selection along particular Quercus lineages associated with the leaf traits and climate characteristics. In parallel, the DT model identified amino acid replacements at sites 95, 219, 262 and 328 being associated with the leaf traits and climate characteristics, exhibiting partial overlap with the results obtained using PAML

The structure of Rubisco enzyme in two projections (A, B) and (C) the residues that belong to active site (coloured red) and that are involved in interactions with Rubisco activase (coloured green; after [15], [24], [26], [27]).

<p>Eight residues replaced in <i>Schiedea</i> (positions 23, 86, 145, 326, 354, 363, 367, 470) are coloured yellow.</p

Improving recombinant Rubisco biogenesis, plant photosynthesis and growth by coexpressing its ancillary RAF1 chaperone

Enabling improvements to crop yield and resource use by enhancing the catalysis of the photosynthetic CO2-fixing enzyme Rubisco has been a longstanding challenge. Efforts toward realization of this goal have been greatly assisted by advances in understanding the complexities of Rubisco\u27s biogenesis in plastids and the development of tailored chloroplast transformation tools. Here we generate transplastomic tobacco genotypes expressing Arabidopsis Rubisco large subunits (AtL), both on their own (producing tobAtL plants) and with a cognate Rubisco accumulation factor 1 (AtRAF1) chaperone (producing tobAtL-R1 plants) that has undergone parallel functional coevolution with AtL. We show AtRAF1 assembles as a dimer and is produced in tobAtL-R1 and Arabidopsis leaves at 10-15 nmol AtRAF1 monomers per square meter. Consistent with a postchaperonin large (L)-subunit assembly role, the AtRAF1 facilitated two to threefold improvements in the amount and biogenesis rate of hybrid L8 AS8 t Rubisco [comprising AtL and tobacco small (S) subunits] in tobAtL-R1 leaves compared with tobAtL, despite \u3ethreefold lower steady-state Rubisco mRNA levels in tobAtL-R1. Accompanying twofold increases in photosynthetic CO2-assimilation rate and plant growth were measured for tobAtL-R1 lines. These findings highlight the importance of ancillary protein complementarity during Rubisco biogenesis in plastids, the possible constraints this has imposed on Rubisco adaptive evolution, and the likely need for such interaction specificity to be considered when optimizing recombinant Rubisco bioengineering in plants