Title: Carboxysome Mispositioning Alters Growth, Morphology, and Rubisco Level of the Cyanobacterium Synechococcus elongatus PCC 7942
ABSTRACT Cyanobacteria are the prokaryotic group of phytoplankton responsible for a significant fraction of global CO 2 fixation. Like plants, cyanobacteria use the enzyme ribulose 1,5-bisphosphate carboxylase/oxidase (Rubisco) to fix CO 2 into organic carbon molecules via the Calvin-Benson-Bassham cycle. Unlike plants, cyanobacteria evolved a carbon-concentrating organelle called the carboxysome—a proteinaceous compartment that encapsulates and concentrates Rubisco along with its CO 2 substrate. In the rod-shaped cyanobacterium Synechococcus elongatus PCC 7942, we recently identified the McdAB system responsible for uniformly distributing carboxysomes along the cell length. It remains unknown what role carboxysome positioning plays with respect to cellular physiology. Here, we show that a failure to distribute carboxysomes leads to slower cell growth, cell elongation, asymmetric cell division, and elevated levels of cellular Rubisco. Unexpectedly, we also report that even wild-type S. elongatus undergoes cell elongation and asymmetric cell division when grown at the cool, but environmentally relevant, growth temperature of 20°C or when switched from a high- to ambient-CO 2 environment. The findings suggest that carboxysome positioning by the McdAB system functions to maintain the carbon fixation efficiency of Rubisco by preventing carboxysome aggregation, which is particularly important under growth conditions where rod-shaped cyanobacteria adopt a filamentous morphology. IMPORTANCE Photosynthetic cyanobacteria are responsible for almost half of global CO 2 fixation. Due to eutrophication, rising temperatures, and increasing atmospheric CO 2 concentrations, cyanobacteria have gained notoriety for their ability to form massive blooms in both freshwater and marine ecosystems across the globe. Like plants, cyanobacteria use the most abundant enzyme on Earth, Rubisco, to provide the sole source of organic carbon required for its photosynthetic growth. Unlike plants, cyanobacteria have evolved a carbon-concentrating organelle called the carboxysome that encapsulates and concentrates Rubisco with its CO 2 substrate to significantly increase carbon fixation efficiency and cell growth. We recently identified the positioning system that distributes carboxysomes in cyanobacteria. However, the physiological consequence of carboxysome mispositioning in the absence of this distribution system remains unknown. Here, we find that carboxysome mispositioning triggers changes in cell growth and morphology as well as elevated levels of cellular Rubisco. more »« less
MacCready, Joshua S.; Tran, Lisa; Basalla, Joseph L.; Hakim, Pusparanee; Vecchiarelli, Anthony G.(
, Molecular Microbiology)
Abstract
Carboxysomes are protein‐based organelles essential for carbon fixation in cyanobacteria and proteobacteria. Previously, we showed that the cyanobacterial nucleoid is used to equally space out β‐carboxysomes across cell lengths by a two‐component system (McdAB) in the model cyanobacteriumSynechococcus elongatusPCC 7942. More recently, we found that McdAB systems are widespread among β‐cyanobacteria, which possess β‐carboxysomes, but are absent in α‐cyanobacteria, which possess structurally and phyletically distinct α‐carboxysomes. Cyanobacterial α‐carboxysomes are thought to have arisen in proteobacteria and then horizontally transferred into cyanobacteria, which suggests that α‐carboxysomes in proteobacteria may also lack the McdAB system. Here, using the model chemoautotrophic proteobacteriumHalothiobacillus neapolitanus, we show that a McdAB system distinct from that of β‐cyanobacteria operates to position α‐carboxysomes across cell lengths. We further show that this system is widespread among α‐carboxysome‐containing proteobacteria and that cyanobacteria likely inherited an α‐carboxysome operon from a proteobacterium lacking themcdABlocus. These results demonstrate that McdAB is a cross‐phylum two‐component system necessary for positioning both α‐ and β‐carboxysomes. The findings have further implications for understanding the positioning of other protein‐based bacterial organelles involved in diverse metabolic processes.
Plain language summary
Cyanobacteria are well known to fix atmospheric CO2into sugars using the enzyme Rubisco. Less appreciated are the carbon‐fixing abilities of proteobacteria with diverse metabolisms. Bacterial Rubisco is housed within organelles called carboxysomes that increase enzymatic efficiency. Here we show that proteobacterial carboxysomes are distributed in the cell by two proteins, McdA and McdB. McdA on the nucleoid interacts with McdB on carboxysomes to equidistantly space carboxysomes from one another, ensuring metabolic homeostasis and a proper inheritance of carboxysomes following cell division. This study illuminates how widespread carboxysome positioning systems are among diverse bacteria. Carboxysomes significantly contribute to global carbon fixation; therefore, understanding the spatial organization mechanism shared across the bacterial world is of great interest.
Basalla, Joseph L; Mak, Claudia A; Byrne, Jordan A; Ghalmi, Maria; Hoang, Y; Vecchiarelli, Anthony G(
, eLife)
Across bacteria, protein-based organelles called bacterial microcompartments (BMCs) encapsulate key enzymes to regulate their activities. The model BMC is the carboxysome that encapsulates enzymes for CO2fixation to increase efficiency and is found in many autotrophic bacteria, such as cyanobacteria. Despite their importance in the global carbon cycle, little is known about how carboxysomes are spatially regulated. We recently identified the two-factor system required for the maintenance of carboxysome distribution (McdAB). McdA drives the equal spacing of carboxysomes via interactions with McdB, which associates with carboxysomes. McdA is a ParA/MinD ATPase, a protein family well studied in positioning diverse cellular structures in bacteria. However, the adaptor proteins like McdB that connect these ATPases to their cargos are extremely diverse. In fact, McdB represents a completely unstudied class of proteins. Despite the diversity, many adaptor proteins undergo phase separation, but functional roles remain unclear. Here, we define the domain architecture of McdB from the model cyanobacteriumSynechococcus elongatusPCC 7942, and dissect its mode of biomolecular condensate formation. We identify an N-terminal intrinsically disordered region (IDR) that modulates condensate solubility, a central coiled-coil dimerizing domain that drives condensate formation, and a C-terminal domain that trimerizes McdB dimers and provides increased valency for condensate formation. We then identify critical basic residues in the IDR, which we mutate to glutamines to solubilize condensates. Finally, we find that a condensate-defective mutant of McdB has altered association with carboxysomes and influences carboxysome enzyme content. The results have broad implications for understanding spatial organization of BMCs and the molecular grammar of protein condensates.
MacCready, Joshua S; Basalla, Joseph L; Vecchiarelli, Anthony G; Echave, Julian(
, Molecular Biology and Evolution)
Abstract Carboxysomes are protein-based organelles that are essential for allowing cyanobacteria to fix CO2. Previously, we identified a two-component system, McdAB, responsible for equidistantly positioning carboxysomes in the model cyanobacterium Synechococcus elongatus PCC 7942 (MacCready JS, Hakim P, Young EJ, Hu L, Liu J, Osteryoung KW, Vecchiarelli AG, Ducat DC. 2018. Protein gradients on the nucleoid position the carbon-fixing organelles of cyanobacteria. eLife 7:pii:e39723). McdA, a ParA-type ATPase, nonspecifically binds the nucleoid in the presence of ATP. McdB, a novel factor that directly binds carboxysomes, displaces McdA from the nucleoid. Removal of McdA from the nucleoid in the vicinity of carboxysomes by McdB causes a global break in McdA symmetry, and carboxysome motion occurs via a Brownian-ratchet-based mechanism toward the highest concentration of McdA. Despite the importance for cyanobacteria to properly position their carboxysomes, whether the McdAB system is widespread among cyanobacteria remains an open question. Here, we show that the McdAB system is widespread among β-cyanobacteria, often clustering with carboxysome-related components, and is absent in α-cyanobacteria. Moreover, we show that two distinct McdAB systems exist in β-cyanobacteria, with Type 2 systems being the most ancestral and abundant, and Type 1 systems, like that of S. elongatus, possibly being acquired more recently. Lastly, all McdB proteins share the sequence signatures of a protein capable of undergoing liquid–liquid phase separation. Indeed, we find that representatives of both McdB types undergo liquid–liquid phase separation in vitro, the first example of a ParA-type ATPase partner protein to exhibit this behavior. Our results have broader implications for understanding carboxysome evolution, biogenesis, homeostasis, and positioning in cyanobacteria.
Carboxysomes, responsible for a substantial fraction of CO 2 fixation on Earth, are proteinaceous microcompartments found in many autotrophic members of domain Bacteria , primarily from the phyla Proteobacteria and Cyanobacteria . Carboxysomes facilitate CO 2 fixation by the Calvin-Benson-Bassham (CBB) cycle, particularly under conditions where the CO 2 concentration is variable or low, or O 2 is abundant. These microcompartments are composed of an icosahedral shell containing the enzymes ribulose 1,5-carboxylase/oxygenase (RubisCO) and carbonic anhydrase. They function as part of a CO 2 concentrating mechanism, in which cells accumulate HCO 3 − in the cytoplasm via active transport, HCO 3 − enters the carboxysomes through pores in the carboxysomal shell proteins, and carboxysomal carbonic anhydrase facilitates the conversion of HCO 3 − to CO 2 , which RubisCO fixes. Two forms of carboxysomes have been described: α-carboxysomes and β-carboxysomes, which arose independently from ancestral microcompartments. The α-carboxysomes present in Proteobacteria and some Cyanobacteria have shells comprised of four types of proteins [CsoS1 hexamers, CsoS4 pentamers, CsoS2 assembly proteins, and α-carboxysomal carbonic anhydrase (CsoSCA)], and contain form IA RubisCO (CbbL and CbbS). In the majority of cases, these components are encoded in the genome near each other in a gene locus, and transcribed together as an operon. Interestingly, genome sequencing has revealed some α-carboxysome loci that are missing genes encoding one or more of these components. Some loci lack the genes encoding RubisCO, others lack a gene encoding carbonic anhydrase, some loci are missing shell protein genes, and in some organisms, genes homologous to those encoding the carboxysome-associated carbonic anhydrase are the only carboxysome-related genes present in the genome. Given that RubisCO, assembly factors, carbonic anhydrase, and shell proteins are all essential for carboxysome function, these absences are quite intriguing. In this review, we provide an overview of the most recent studies of the structural components of carboxysomes, describe the genomic context and taxonomic distribution of atypical carboxysome loci, and propose functions for these variants. We suggest that these atypical loci are JEEPs, which have modified functions based on the presence of Just Enough Essential Parts.
Hanson, Maureen R.; Lin, Myat T.; Carmo‐Silva, A. Elizabete; Parry, Martin A. J.(
, The Plant Journal)
Summary
Photosynthesis in C3 plants is limited by features of the carbon‐fixing enzyme Rubisco, which exhibits a low turnover rate and can react with O2instead ofCO2, leading to photorespiration. In cyanobacteria, bacterial microcompartments, known as carboxysomes, improve the efficiency of photosynthesis by concentratingCO2near the enzyme Rubisco. Cyanobacterial Rubisco enzymes are faster than those of C3 plants, though they have lower specificity towardCO2than the land plant enzyme. Replacement of land plant Rubisco by faster bacterial variants with lowerCO2specificity will improve photosynthesis only if a microcompartment capable of concentratingCO2can also be installed into the chloroplast. We review current information about cyanobacterial microcompartments and carbon‐concentrating mechanisms, plant transformation strategies, replacement of Rubisco in a model C3 plant with cyanobacterial Rubisco and progress toward synthesizing a carboxysome in chloroplasts.
Rillema, Rees, Hoang, Y, MacCready, Joshua S., and Vecchiarelli, Anthony G. Carboxysome Mispositioning Alters Growth, Morphology, and Rubisco Level of the Cyanobacterium Synechococcus elongatus PCC 7942. Retrieved from https://par.nsf.gov/biblio/10298476. mBio 12.4 Web. doi:10.1128/mBio.02696-20.
Rillema, Rees, Hoang, Y, MacCready, Joshua S., & Vecchiarelli, Anthony G. Carboxysome Mispositioning Alters Growth, Morphology, and Rubisco Level of the Cyanobacterium Synechococcus elongatus PCC 7942. mBio, 12 (4). Retrieved from https://par.nsf.gov/biblio/10298476. https://doi.org/10.1128/mBio.02696-20
Rillema, Rees, Hoang, Y, MacCready, Joshua S., and Vecchiarelli, Anthony G.
"Carboxysome Mispositioning Alters Growth, Morphology, and Rubisco Level of the Cyanobacterium Synechococcus elongatus PCC 7942". mBio 12 (4). Country unknown/Code not available. https://doi.org/10.1128/mBio.02696-20.https://par.nsf.gov/biblio/10298476.
@article{osti_10298476,
place = {Country unknown/Code not available},
title = {Carboxysome Mispositioning Alters Growth, Morphology, and Rubisco Level of the Cyanobacterium Synechococcus elongatus PCC 7942},
url = {https://par.nsf.gov/biblio/10298476},
DOI = {10.1128/mBio.02696-20},
abstractNote = {ABSTRACT Cyanobacteria are the prokaryotic group of phytoplankton responsible for a significant fraction of global CO 2 fixation. Like plants, cyanobacteria use the enzyme ribulose 1,5-bisphosphate carboxylase/oxidase (Rubisco) to fix CO 2 into organic carbon molecules via the Calvin-Benson-Bassham cycle. Unlike plants, cyanobacteria evolved a carbon-concentrating organelle called the carboxysome—a proteinaceous compartment that encapsulates and concentrates Rubisco along with its CO 2 substrate. In the rod-shaped cyanobacterium Synechococcus elongatus PCC 7942, we recently identified the McdAB system responsible for uniformly distributing carboxysomes along the cell length. It remains unknown what role carboxysome positioning plays with respect to cellular physiology. Here, we show that a failure to distribute carboxysomes leads to slower cell growth, cell elongation, asymmetric cell division, and elevated levels of cellular Rubisco. Unexpectedly, we also report that even wild-type S. elongatus undergoes cell elongation and asymmetric cell division when grown at the cool, but environmentally relevant, growth temperature of 20°C or when switched from a high- to ambient-CO 2 environment. The findings suggest that carboxysome positioning by the McdAB system functions to maintain the carbon fixation efficiency of Rubisco by preventing carboxysome aggregation, which is particularly important under growth conditions where rod-shaped cyanobacteria adopt a filamentous morphology. IMPORTANCE Photosynthetic cyanobacteria are responsible for almost half of global CO 2 fixation. Due to eutrophication, rising temperatures, and increasing atmospheric CO 2 concentrations, cyanobacteria have gained notoriety for their ability to form massive blooms in both freshwater and marine ecosystems across the globe. Like plants, cyanobacteria use the most abundant enzyme on Earth, Rubisco, to provide the sole source of organic carbon required for its photosynthetic growth. Unlike plants, cyanobacteria have evolved a carbon-concentrating organelle called the carboxysome that encapsulates and concentrates Rubisco with its CO 2 substrate to significantly increase carbon fixation efficiency and cell growth. We recently identified the positioning system that distributes carboxysomes in cyanobacteria. However, the physiological consequence of carboxysome mispositioning in the absence of this distribution system remains unknown. Here, we find that carboxysome mispositioning triggers changes in cell growth and morphology as well as elevated levels of cellular Rubisco.},
journal = {mBio},
volume = {12},
number = {4},
author = {Rillema, Rees and Hoang, Y and MacCready, Joshua S. and Vecchiarelli, Anthony G.},
editor = {Komeili, Arash}
}
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