scholarly journals Role of IncP-1β Plasmids pWDL7::rfpand pNB8c in Chloroaniline Catabolism as Determined by Genomic and Functional Analyses

2011 ◽  
Vol 78 (3) ◽  
pp. 828-838 ◽  
Author(s):  
J. E. Król ◽  
J. T. Penrod ◽  
H. McCaslin ◽  
L. M. Rogers ◽  
H. Yano ◽  
...  
Keyword(s):  
Single Copy ◽  
Degradation Pathway ◽  
Bacterial Degradation ◽  
Ring Cleavage ◽  
Close Relative ◽  
Broad Host Range ◽  
Content Type ◽  
Aromatic Ring Cleavage ◽  
Functional Analyses

ABSTRACTBroad-host-range catabolic plasmids play an important role in bacterial degradation of man-made compounds. To gain insight into the role of these plasmids in chloroaniline degradation, we determined the first complete nucleotide sequences of an IncP-1 chloroaniline degradation plasmid, pWDL7::rfpand its close relative pNB8c, as well as the expression pattern, function, and bioaugmentation potential of the putative 3-chloroaniline (3-CA) oxidation genes. Based on phylogenetic analysis of backbone proteins, both plasmids are members of a distinct clade within the IncP-1β subgroup. The plasmids are almost identical, but whereas pWDL7::rfpcarries a duplicate inverted catabolic transposon, Tn6063, containing a putative 3-CA oxidation gene cluster,dcaQTA1A2BR, pNB8c contains only a single copy of the transposon. No genes for an aromatic ring cleavage pathway were detected on either plasmid, suggesting that only the upper 3-CA degradation pathway was present. ThedcaA1A2Bgene products expressed from a high-copy-number vector were shown to convert 3-CA to 4-chlorocatechol inEscherichia coli. Slight differences in thedcapromoter region between the plasmids and lack of induction of transcription of the pNB8cdcagenes by 3-CA may explain previous findings that pNB8C does not confer 3-CA transformation. Bioaugmentation of activated sludge with pWDL7::rfpaccelerated removal of 3-CA, but only in the presence of an additional carbon source. Successful bioaugmentation requires complementation of the upper pathway genes with chlorocatechol cleavage genes in indigenous bacteria. The genome sequences of these plasmids thus help explain the molecular basis of their catabolic activities.

scholarly journals Steroid Degradation in Comamonas testosteroni TA441: Identification of the Entire β-Oxidation Cycle of the Cleaved B Ring

2019 ◽  
Vol 85 (20) ◽  
Author(s):  
Masae Horinouchi ◽  
Hiroyuki Koshino ◽  
Michal Malon ◽  
Hiroshi Hirota ◽  
Toshiaki Hayashi
Keyword(s):  
Gene Clusters ◽  
Degradation Process ◽  
Degradation Pathway ◽  
Ring Cleavage ◽  
Comamonas Testosteroni ◽  
Content Type ◽  
Coa Transferase ◽  
Degrading Bacteria ◽  
Globally Distributed

ABSTRACT Comamonas testosteroni TA441 degrades steroids via aromatization of the A ring, followed by degradation of 9,17-dioxo-1,2,3,4,10,19-hexanorandrostan-5-oic acid, mainly by β-oxidation. In this study, we revealed that 7β,9α-dihydroxy-17-oxo-1,2,3,4,10,19-hexanorandrostanoic acid-coenzyme A (CoA) ester is dehydrogenated by (3S)-3-hydroxylacyl CoA-dehydrogenase, encoded by scdE (ORF27), and then the resultant 9α-hydroxy-7,17-dioxo-1,2,3,4,10,19-hexanorandrostan-5-oic acid-CoA ester is converted by 3-ketoacyl-CoA transferase, encoded by scdF (ORF23). With these results, the whole cycle of β-oxidation on the side chain at C-8 of 9,17-dioxo-1,2,3,4,10,19-hexanorandrostan-5-oic acid is clarified; 9-hydroxy-17-oxo-1,2,3,4,10,19-hexanorandrostan-5-oic acid-CoA ester is dehydrogenated at C-6 by ScdC1C2, followed by hydration by ScdD. 7β,9α-Dihydroxy-17-oxo-1,2,3,4,10,19-hexanorandrostanoic acid-CoA ester then is dehydrogenated by ScdE to be converted to 9α-hydroxy-17-oxo-1,2,3,4,5,6,10,19-octanorandrostan-7-oic acid-CoA ester and acetyl-CoA by ScdF. ScdF is an ortholog of FadA6 in Mycobacterium tuberculosis H37Rv, which was reported as a 3-ketoacyl-CoA transferase involved in C ring cleavage. We also obtained results suggesting that ScdF is also involved in C ring cleavage, but further investigation is required for confirmation. ORF25 and ORF26, located between scdF and scdE, encode enzymes belonging to the amidase superfamily. Disrupting either ORF25 or ORF26 did not affect steroid degradation. Among the bacteria having gene clusters similar to those of tesB to tesR, some have both ORF25- and ORF26-like proteins or only an ORF26-like protein, but others do not have either ORF25- or ORF26-like proteins. ORF25 and ORF26 are not crucial for steroid degradation, yet they might provide clues to elucidate the evolution of bacterial steroid degradation clusters. IMPORTANCE Studies on bacterial steroid degradation were initiated more than 50 years ago primarily to obtain materials for steroid drugs. Steroid-degrading bacteria are globally distributed, and the role of bacterial steroid degradation in the environment as well as in relation to human health is attracting attention. The overall aerobic degradation of the four basic steroidal rings has been proposed; however, there is still much to be revealed to understand the complete degradation pathway. This study aims to uncover the whole steroid degradation process in Comamonas testosteroni TA441 as a model of steroid-degrading bacteria. C. testosteroni is one of the most studied representative steroid-degrading bacteria and is suitable for exploring the degradation pathway, because the involvement of degradation-related genes can be determined by gene disruption. Here, we elucidated the entire β-oxidation cycle of the cleaved B ring. This cycle is essential for the following C and D ring cleavage.

scholarly journals Proteogenomic Elucidation of the Initial Steps in the Benzene Degradation Pathway of a Novel Halophile, Arhodomonas sp. Strain Rozel, Isolated from a Hypersaline Environment

2012 ◽  
Vol 78 (20) ◽  
pp. 7309-7316 ◽  
Author(s):  
Sonal Dalvi ◽  
Sei Azetsu ◽  
Marianna A. Patrauchan ◽  
Deniz F. Aktas ◽  
Babu Z. Fathepure
Keyword(s):  
Mass Spectrometry ◽  
Tricarboxylic Acid Cycle ◽  
Draft Genome ◽  
Degradation Pathway ◽  
Phenol Hydroxylase ◽  
Ring Cleavage ◽  
Content Type ◽  
Benzene Degradation ◽  
Chromatography Mass Spectrometry

ABSTRACTLately, there has been a special interest in understanding the role of halophilic and halotolerant organisms for their ability to degrade hydrocarbons. The focus of this study was to investigate the genes and enzymes involved in the initial steps of the benzene degradation pathway in halophiles. The extremely halophilic bacteriaArhodomonassp. strain Seminole andArhodomonassp. strain Rozel, which degrade benzene and toluene as the sole carbon source at high salinity (0.5 to 4 M NaCl), were isolated from enrichments developed from contaminated hypersaline environments. To obtain insights into the physiology of this novel group of organisms, a draft genome sequence of the Seminole strain was obtained. A cluster of 13 genes predicted to be functional in the hydrocarbon degradation pathway was identified from the sequence. Two-dimensional (2D) gel electrophoresis and liquid chromatography-mass spectrometry were used to corroborate the role of the predicted open reading frames (ORFs). ORFs 1080 and 1082 were identified as components of a multicomponent phenol hydroxylase complex, and ORF 1086 was identified as catechol 2,3-dioxygenase (2,3-CAT). Based on this analysis, it was hypothesized that benzene is converted to phenol and then to catechol by phenol hydroxylase components. The resulting catechol undergoes ring cleavage via the meta pathway by 2,3-CAT to form 2-hydroxymuconic semialdehyde, which enters the tricarboxylic acid cycle. To substantiate these findings, the Rozel strain was grown on deuterated benzene, and gas chromatography-mass spectrometry detected deuterated phenol as the initial intermediate of benzene degradation. These studies establish the initial steps of the benzene degradation pathway in halophiles.

scholarly journals Estrogen Degraders and Estrogen Degradation Pathway Identified in an Activated Sludge

2018 ◽  
Vol 84 (10) ◽  
Author(s):  
Yi-Lung Chen ◽  
Han-Yi Fu ◽  
Tzong-Huei Lee ◽  
Chao-Jen Shih ◽  
Lina Huang ◽  
...  
Keyword(s):  
Wastewater Treatment ◽  
Activated Sludge ◽  
Wastewater Treatment Plants ◽  
Sewage Treatment Plant ◽  
Degradation Pathway ◽  
Bacterial Degradation ◽  
Ring Cleavage ◽  
Main Process ◽  
Content Type ◽  
Degrading Bacteria

ABSTRACTThe environmental release and fate of estrogens are becoming an increasing public concern. Bacterial degradation has been considered the main process for eliminating estrogens from wastewater treatment plants. Various bacterial isolates are reportedly capable of aerobic estrogen degradation, and several estrogen degradation pathways have been proposed in proteobacteria and actinobacteria. However, the ecophysiological relevance of estrogen-degrading bacteria in the environment is unclear. In this study, we investigated the estrogen degradation pathway and corresponding degraders in activated sludge collected from the Dihua Sewage Treatment Plant, Taipei, Taiwan. Cultivation-dependent and cultivation-independent methods were used to assess estrogen biodegradation in the collected activated sludge. Estrogen metabolite profile analysis revealed the production of pyridinestrone acid and two A/B-ring cleavage products in activated sludge incubated with estrone (1 mM), which are characteristic of the 4,5-secopathway. PCR-based functional assays detected sequences closely related to alphaproteobacterialoecC, a key gene of the 4,5-secopathway. Metagenomic analysis suggested thatNovosphingobiumspp. are major estrogen degraders in estrone-amended activated sludge.Novosphingobiumsp. strain SLCC, an estrone-degrading alphaproteobacterium, was isolated from the examined activated sludge. The general physiology and metabolism of this strain were characterized. Pyridinestrone acid and the A/B-ring cleavage products were detected in estrone-grown strain SLCC cultures. The production of pyridinestrone acid was also observed during the aerobic incubation of strain SLCC with 3.7 nM (1 μg/liter) estrone. This concentration is close to that detected in many natural and engineered aquatic ecosystems. The presented data suggest the ecophysiological relevance ofNovosphingobiumspp. in activated sludge.IMPORTANCEEstrogens, which persistently contaminate surface water worldwide, have been classified as endocrine disruptors and human carcinogens. We contribute new knowledge on the major estrogen biodegradation pathway and estrogen degraders in wastewater treatment plants. This study considerably advances the understanding of environmental estrogen biodegradation, which is instrumental for the efficient elimination of these hazardous pollutants. Moreover, this study substantially improves the understanding of microbial estrogen degradation in the environment.

scholarly journals Biodegradation of the Allelopathic Chemical Pterostilbene by a Sphingobium sp. Strain from the Peanut Rhizosphere

2018 ◽  
Vol 85 (5) ◽  
Author(s):  
Ri-Qing Yu ◽  
Zohre Kurt ◽  
Fei He ◽  
Jim C. Spain
Keyword(s):  
Pathogenic Fungi ◽  
Bacterial Degradation ◽  
Ecological Role ◽  
Microbial Infection ◽  
Catabolic Pathway ◽  
Content Type ◽  
Dead End ◽  
Initial Cleavage ◽  
Bacterial Interference

ABSTRACT Many plants produce allelopathic chemicals, such as stilbenes, to inhibit pathogenic fungi. The degradation of allelopathic compounds by bacteria associated with the plants would limit their effectiveness, but little is known about the extent of biodegradation or the bacteria involved. Screening of tissues and rhizosphere of peanut (Arachis hypogaea) plants revealed substantial enrichment of bacteria able to grow on resveratrol and pterostilbene, the most common stilbenes produced by the plants. Investigation of the catabolic pathway in Sphingobium sp. strain JS1018, isolated from the rhizosphere, indicated that the initial cleavage of pterostilbene was catalyzed by a carotenoid cleavage oxygenase (CCO), which led to the transient accumulation of 4-hydroxybenzaldehyde and 3,5-dimethoxybenzaldehyde. 4-Hydroxybenzaldehyde was subsequently used for the growth of the isolate, while 3,5-dimethoxybenzaldehyde was further converted to a dead-end metabolite with a molecular weight of 414 (C24H31O6). The gene that encodes the initial oxygenase was identified in the genome of strain JS1018, and its function was confirmed by heterologous expression in Escherichia coli. This study reveals the biodegradation pathway of pterostilbene by plant-associated bacteria. The prevalence of such bacteria in the rhizosphere and plant tissues suggests a potential role of bacterial interference in plant allelopathy. IMPORTANCE Pterostilbene, an analog of resveratrol, is a stilbene allelochemical produced by plants to inhibit microbial infection. As a potent antioxidant, pterostilbene acts more effectively than resveratrol as an antifungal agent. Bacterial degradation of this plant natural product would affect the allelopathic efficacy and fate of pterostilbene and thus its ecological role. This study explores the isolation and abundance of bacteria that degrade resveratrol and pterostilbene in peanut tissues and rhizosphere, the catabolic pathway for pterostilbene, and the molecular basis for the initial cleavage of pterostilbene. If plant allelopathy is an important process in agriculture and management of invasive plants, the ecological role of bacteria that degrade the allelopathic chemicals must be equally important.

scholarly journals Functional Redundancy of Linuron Degradation in Microbial Communities in Agricultural Soil and Biopurification Systems

2016 ◽  
Vol 82 (9) ◽  
pp. 2843-2853 ◽  
Author(s):  
Benjamin Horemans ◽  
Karolien Bers ◽  
Erick Ruiz Romero ◽  
Eva Pose Juan ◽  
Vincent Dunon ◽  
...  
Keyword(s):  
Agricultural Soil ◽  
Bacterial Consortium ◽  
Bacterial Degradation ◽  
Catabolic Gene ◽  
Bacterial Populations ◽  
Content Type ◽  
Respective Contribution ◽  
On Farm ◽  
Biopurification Systems

ABSTRACTThe abundance oflibA, encoding a hydrolase that initiates linuron degradation in the linuron-metabolizingVariovoraxsp. strain SRS16, was previously found to correlate well with linuron mineralization, but not in all tested environments. Recently, an alternative linuron hydrolase, HylA, was identified inVariovoraxsp. strain WDL1, a strain that initiates linuron degradation in a linuron-mineralizing commensal bacterial consortium. The discovery of alternative linuron hydrolases poses questions about the respective contribution and competitive character ofhylA- andlibA-carrying bacteria as well as the role of linuron-mineralizing consortia versus single strains in linuron-exposed settings. Therefore, dynamics ofhylAas well asdcaQas a marker for downstream catabolic functions involved in linuron mineralization, in response to linuron treatment in agricultural soil and on-farm biopurification systems (BPS), were compared with previously reportedlibAdynamics. The results suggest that (i) organisms containing eitherlibAorhylAcontribute simultaneously to linuron biodegradation in the same environment, albeit to various extents, (ii) environmental linuron mineralization depends on multispecies bacterial food webs, and (iii) initiation of linuron mineralization can be governed by currently unidentified enzymes.IMPORTANCEA limited set of different isofunctional catabolic gene functions is known for the bacterial degradation of the phenylurea herbicide linuron, but the role of this redundancy in linuron degradation in environmental settings is not known. In this study, the simultaneous involvement of bacteria carrying one of two isofunctional linuron hydrolysis genes in the degradation of linuron was shown in agricultural soil and on-farm biopurification systems, as was the involvement of other bacterial populations that mineralize the downstream metabolites of linuron hydrolysis. This study illustrates the importance of the synergistic metabolism of pesticides in environmental settings.

scholarly journals Novel Pathway for the Degradation of 2-Chloro-4-Nitrobenzoic Acid by Acinetobacter sp. Strain RKJ12

2011 ◽  
Vol 77 (18) ◽  
pp. 6606-6613 ◽  
Author(s):  
Dhan Prakash ◽  
Ravi Kumar ◽  
R. K. Jain ◽  
B. N. Tiwary
Keyword(s):  
Salicylic Acid ◽  
Tricarboxylic Acid Cycle ◽  
Sole Source ◽  
Degradation Pathway ◽  
Ring Cleavage ◽  
Muconic Acid ◽  
Content Type ◽  
Nitrobenzoic Acid ◽  
Catabolic Genes ◽  
Catechol 1,2 Dioxygenase

ABSTRACTThe organismAcinetobactersp. RKJ12 is capable of utilizing 2-chloro-4-nitrobenzoic acid (2C4NBA) as a sole source of carbon, nitrogen, and energy. In the degradation of 2C4NBA by strain RKJ12, various metabolites were isolated and identified by a combination of chromatographic, spectroscopic, and enzymatic activities, revealing a novel assimilation pathway involving both oxidative and reductive catabolic mechanisms. The metabolism of 2C4NBA was initiated by oxidativeorthodehalogenation, leading to the formation of 2-hydroxy-4-nitrobenzoic acid (2H4NBA), which subsequently was metabolized into 2,4-dihydroxybenzoic acid (2,4-DHBA) by a mono-oxygenase with the concomitant release of chloride and nitrite ions. Stoichiometric analysis indicated the consumption of 1 mol O2per conversion of 2C4NBA to 2,4-DHBA, ruling out the possibility of two oxidative reactions. Experiments with labeled H218O and18O2indicated the involvement of mono-oxygenase-catalyzed initial hydrolytic dechlorination and oxidative denitration mechanisms. The further degradation of 2,4-DHBA then proceeds via reductive dehydroxylation involving the formation of salicylic acid. In the lower pathway, the organism transformed salicylic acid into catechol, which was mineralized by theorthoring cleavage catechol-1,2-dioxygenase tocis, cis-muconic acid, ultimately forming tricarboxylic acid cycle intermediates. Furthermore, the studies carried out on a 2C4NBA−derivative and a 2C4NBA+transconjugant demonstrated that the catabolic genes for the 2C4NBA degradation pathway possibly reside on the ∼55-kb transmissible plasmid present in RKJ12.

scholarly journals Construction of a Broad-Host-Range Tn7-Based Vector for Single-Copy PBAD-Controlled Gene Expression in Gram-Negative Bacteria

2012 ◽  
Vol 79 (2) ◽  
pp. 718-721 ◽  
Author(s):  
F. Heath Damron ◽  
Elizabeth S. McKenney ◽  
Herbert P. Schweizer ◽  
Joanna B. Goldberg
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Host Range ◽  
Single Copy ◽  
Broad Host Range ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Content Type ◽  
Delivery Vector ◽  
Inducible Gene

ABSTRACTWe describe a mini-Tn7-based broad-host-range expression cassette for arabinose-inducible gene expression from the PBADpromoter. This delivery vector, pTJ1, can integrate a single copy of a gene into the chromosome of Gram-negative bacteria for diverse genetic applications, of which several are discussed, usingPseudomonas aeruginosaas the model host.

scholarly journals The LuxS-Dependent Quorum-Sensing System Regulates Early Biofilm Formation by Streptococcus pneumoniae Strain D39

2011 ◽  
Vol 79 (10) ◽  
pp. 4050-4060 ◽  
Author(s):  
Jorge E. Vidal ◽  
Herbert P. Ludewick ◽  
Rebekah M. Kunkel ◽  
Dorothea Zähner ◽  
Keith P. Klugman
Keyword(s):  
Streptococcus Pneumoniae ◽  
Quorum Sensing ◽  
Biofilm Formation ◽  
Single Copy ◽  
Middle Ear Mucosa ◽  
Transcript Levels ◽  
Content Type ◽  
Sensing System ◽  
Quorum Sensing System

ABSTRACTStreptococcus pneumoniaeis the leading cause of death in children worldwide and forms highly organized biofilms in the nasopharynx, lungs, and middle ear mucosa. TheluxS-controlled quorum-sensing (QS) system has recently been implicated in virulence and persistence in the nasopharynx, but its role in biofilms has not been studied. Here we show that this QS system plays a major role in the control ofS. pneumoniaebiofilm formation. Our results demonstrate that theluxSgene is contained by invasive isolates and normal-flora strains in a region that contains genes involved in division and cell wall biosynthesis. TheluxSgene was maximally transcribed, as a monocistronic message, in the early mid-log phase of growth, and this coincides with the appearance of early biofilms. Demonstrating the role of the LuxS system in regulatingS. pneumoniaebiofilms, at 24 h postinoculation, two different D39ΔluxSmutants produced ∼80% less biofilm biomass than wild-type (WT) strain D39 did. Complementation of these strains withluxS, either in a plasmid or integrated as a single copy in the genome, restored their biofilm level to that of the WT. Moreover, a soluble factor secreted by WT strain D39 or purified AI-2 restored the biofilm phenotype of D39ΔluxS. Our results also demonstrate that during the early mid-log phase of growth, LuxS regulates the transcript levels oflytA, which encodes an autolysin previously implicated in biofilms, and also the transcript levels ofply, which encodes the pneumococcal pneumolysin. In conclusion, theluxS-controlled QS system is a key regulator of early biofilm formation byS. pneumoniaestrain D39.

scholarly journals Glycerol-3-Phosphate Acyltransferase Contributes to Triacylglycerol Biosynthesis, Lipid Droplet Formation, and Host Invasion in Metarhizium robertsii

2013 ◽  
Vol 79 (24) ◽  
pp. 7646-7653 ◽  
Author(s):  
Qiang Gao ◽  
Yanfang Shang ◽  
Wei Huang ◽  
Chengshu Wang
Keyword(s):  
Pathogenic Fungus ◽  
Single Copy ◽  
Fungal Genome ◽  
Model Organisms ◽  
Glycerol Phosphate ◽  
Fungal Virulence ◽  
Metarhizium Robertsii ◽  
Content Type ◽  
Rate Limiting Step

ABSTRACTEnzymes involved in the triacylglycerol (TAG) biosynthesis have been well studied in the model organisms of yeasts and animals. Among these, the isoforms of glycerol-3-phosphate acyltransferase (GPAT) redundantly catalyze the first and rate-limiting step in glycerolipid synthesis. Here, we report the functions of mrGAT, a GPAT ortholog, in an insect-pathogenic fungus,Metarhizium robertsii. Unlike in yeasts and animals, a single copy of the mrGAT gene is present in the fungal genome and the gene deletion mutant is viable. Compared to the wild type and the gene-rescued mutant, the ΔmrGATmutant demonstrated reduced abilities to produce conidia and synthesize TAG, glycerol, and total lipids. More importantly, we found that mrGAT is localized to the endoplasmic reticulum and directly linked to the formation of lipid droplets (LDs) in fungal cells. Insect bioassay results showed thatmrGATis required for full fungal virulence by aiding fungal penetration of host cuticles. Data from this study not only advance our understanding of GPAT functions in fungi but also suggest that filamentous fungi such asM. robertsiican serve as a good model to elucidate the role of the glycerol phosphate pathway in fungal physiology, particularly to determine the mechanistic connection of GPAT to LD formation.

scholarly journals Coordinated Regulation of the Size and Number of Polyhydroxybutyrate Granules by Core and Accessory Phasins in the Facultative Microsymbiont Sinorhizobium fredii NGR234

2019 ◽  
Vol 85 (19) ◽  
Author(s):  
Yan-Wei Sun ◽  
Yan Li ◽  
Yue Hu ◽  
Wen-Xin Chen ◽  
Chang-Fu Tian
Keyword(s):  
Broad Host Range ◽  
Metabolic Profiles ◽  
Free Living ◽  
Sinorhizobium Fredii ◽  
Content Type ◽  
Cellular Processes ◽  
Size Number ◽  
Associated Proteins ◽  
Auxiliary Gene

ABSTRACT The exact roles of various granule-associated proteins (GAPs) of polyhydroxybutyrate (PHB) are poorly investigated, particularly for bacteria associated with plants. In this study, four structural GAPs, named phasins PhaP1 to PhaP4, were identified and demonstrated as true phasins colocalized with PHB granules in Sinorhizobium fredii NGR234, a facultative microsymbiont of Vigna unguiculata and many other legumes. The conserved PhaP2 dominated in regulation of granule size under both free-living and symbiotic conditions. PhaP1, another conserved phasin, made a higher contribution than accessory phasins PhaP4 and PhaP3 to PHB biosynthesis at stationary phase. PhaP3, with limited phyletic distribution on the symbiosis plasmid of Sinorhizobium, was more important than PhaP1 in regulating PHB biosynthesis in V. unguiculata nodules. Under the test conditions, no significant symbiotic defects were observed for mutants lacking individual or multiple phaP genes. The mutant lacking two PHB synthases showed impaired symbiotic performance, while mutations in individual PHB synthases or a PHB depolymerase yielded no symbiotic defects. This phenomenon is not related to either the number or size of PHB granules in test mutants within nodules. Distinct metabolic profiles and cocktail pools of GAPs of different phaP mutants imply that core and accessory phasins can be differentially involved in regulating other cellular processes in the facultative microsymbiont S. fredii NGR234. IMPORTANCE Polyhydroxybutyrate (PHB) granules are a store of carbon and energy in bacteria and archaea and play an important role in stress adaptation. Recent studies have highlighted distinct roles of several granule-associated proteins (GAPs) in regulating the size, number, and localization of PHB granules in free-living bacteria, though our knowledge of the role of GAPs in bacteria associated with plants is still limited. Here we report distinct roles of core and accessory phasins associated with PHB granules of Sinorhizobium fredii NGR234, a broad-host-range microsymbiont of diverse legumes. Core phasins PhaP2 and PhaP1 are conserved major phasins in free-living cells. PhaP2 and accessory phasin PhaP3, encoded by an auxiliary gene on the symbiosis plasmid, are major phasins in nitrogen-fixing bacteroids in cowpea nodules. GAPs and metabolic profiles can vary in different phaP mutants. Contrasting symbiotic performances between mutants lacking PHB synthases, depolymerase, or phasins were revealed.

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