scholarly journals A Pair of Iron-Responsive Genes Encoding Protein Kinases with a Ser/Thr Kinase Domain and a His Kinase Domain Are Regulated by NtcA in the Cyanobacterium Anabaena sp. Strain PCC 7120

2006 ◽  
Vol 188 (13) ◽  
pp. 4822-4829 ◽  
Author(s):  
Yong Cheng ◽  
Jian-Hong Li ◽  
Lei Shi ◽  
Li Wang ◽  
Amel Latifi ◽  
...  
Keyword(s):  
Iron Acquisition ◽  
Iron Chelator ◽  
Kinase Domain ◽  
Iron Limitation ◽  
Filamentous Cyanobacterium ◽  
Wild Type ◽  
Putative Promoter Region ◽  
Iron Deprivation ◽  
Genes Encoding ◽  
Pcc 7120

ABSTRACT The filamentous cyanobacterium Anabaena sp. strain PCC 7120 can fix N2 when combined nitrogen is not available in the growth medium. It has a family of 13 genes encoding proteins with both a Ser/Thr kinase domain and a His kinase domain. The function of these enzymes is unknown. Two of them are encoded by pkn41 (alr0709) and pkn42 (alr0710). These two genes are separated by only 72 bp on the chromosome, and our results indicate that they are cotranscribed. The expression of pkn41 and pkn42 is induced by iron deprivation irrespective of the nature of the nitrogen source. Mutants inactivating either pkn41, pkn42, or both grow similarly to the wild type under normal conditions, but their growth is impaired either in the presence of an iron chelator or under conditions of nitrogen fixation and iron limitation, two situations where the demand for iron is particularly strong. Consistent with these results, these mutants display lower iron content than the wild type and a higher level of expression for nifJ1 and nifJ2, which encode pyruvate:ferredoxin oxidoreductases. Both nifJ1 and nifJ2 are known to be induced by iron limitation. NtcA, a global regulatory factor for different metabolic pathways, binds to the putative promoter region of pkn41, and the induction of pkn41 in response to iron limitation no longer occurs in an ntcA mutant. Our results suggest that ntcA not only regulates the expression of genes involved in nitrogen and carbon metabolism but also coordinates iron acquisition and nitrogen metabolism by activating the expression of pkn41 and pkn42.

scholarly journals Two Genes Encoding Protein Kinases of the HstK Family Are Involved in Synthesis of the Minor Heterocyst-Specific Glycolipid in the Cyanobacterium Anabaena sp. Strain PCC 7120

2007 ◽  
Vol 189 (14) ◽  
pp. 5075-5081 ◽  
Author(s):  
Lei Shi ◽  
Jian-Hong Li ◽  
Yong Cheng ◽  
Li Wang ◽  
Wen-Li Chen ◽  
...  
Keyword(s):  
Nitrogenase Activity ◽  
Early Stage ◽  
Kinase Domain ◽  
Filamentous Cyanobacterium ◽  
Regulatory Pathways ◽  
Genes Encoding ◽  
Putative Transcription Factor ◽  
Oxygen Sensitive ◽  
Cyanobacterium Anabaena ◽  
Pcc 7120

ABSTRACT The filamentous cyanobacterium Anabaena sp. strain PCC 7120 can fix N2 under oxic conditions, and the activity of nitrogen fixation occurs exclusively in heterocysts, cells differentiated from vegetative cells in response to a limitation of a combined-nitrogen source in the growth medium. At the late stages of heterocyst differentiation, an envelope layer composed of two glycolipids is formed to limit the entry of oxygen so that the oxygen-sensitive nitrogenase can function. The genome of Anabaena sp. strain PCC 7120 possesses a family of 13 genes (the hstK family), all encoding proteins with a putative Ser/Thr kinase domain at their N termini and a His-kinase domain at their C termini. In this study, we showed that the double mutant D4.3 strain, in which two members of this gene family, pkn44 (all1625) and pkn30 (all3691), were both inactivated, failed to fix N2 in the presence of oxygen (Fox−). In an environment without oxygen, a low level of nitrogenase activity was detectable (Fix+). Heterocyst development in the mutant D4.3 was delayed by 24 h and arrested at a relatively early stage without the formation of the glycolipid layer (Hgl−). Only the minor species of the two heterocyst-specific glycolipids (HGLs) was missing in the mutant. We propose that DevH, a putative transcription factor, coordinates the synthesis of both HGLs, while Pkn44/Pkn30 and the previously characterized PrpJ may represent two distinct regulatory pathways involved in the synthesis of the minor HGL and the major HGL, respectively.

scholarly journals PlmA, a New Member of the GntR Family, Has Plasmid Maintenance Functions in Anabaena sp. Strain PCC 7120

2003 ◽  
Vol 185 (15) ◽  
pp. 4315-4325 ◽  
Author(s):  
Martin H. Lee ◽  
Michael Scherer ◽  
Sébastien Rigali ◽  
James W. Golden
Keyword(s):  
Plasmid Copy Number ◽  
Filamentous Cyanobacterium ◽  
Wild Type ◽  
Plasmid Copy ◽  
Reading Frame ◽  
Plasmid Maintenance ◽  
Cyanobacterial Species ◽  
A Genome ◽  
Pcc 7120 ◽  
Gntr Family

ABSTRACT The filamentous cyanobacterium Anabaena (Nostoc) sp. strain PCC 7120 maintains a genome that is divided into a 6.4-Mb chromosome, three large plasmids of more that 100 kb, two medium-sized plasmids of 55 and 40 kb, and a 5.5-kb plasmid. Plasmid copy number can be dynamic in some cyanobacterial species, and the genes that regulate this process have not been characterized. Here we show that mutations in an open reading frame, all1076, reduce the numbers of copies per chromosome of several plasmids. In a mutant strain, plasmids pCC7120δ and pCC7120ζ are both reduced to less than 50% of their wild-type levels. The exogenous pDU1-based plasmid pAM1691 is reduced to less than 25% of its wild-type level, and the plasmid is rapidly lost. The peptide encoded by all1076 shows similarity to members of the GntR family of transcriptional regulators. Phylogenetic analysis reveals a new domain topology within the GntR family. PlmA homologs, all coming from cyanobacterial species, form a new subfamily that is distinct from the previously identified subfamilies. The all1076 locus, named plmA, regulates plasmid maintenance functions in Anabaena sp. strain PCC 7120.

scholarly journals c-di-GMP Homeostasis Is Critical for Heterocyst Development in Anabaena sp. PCC 7120

2021 ◽  
Vol 12 ◽  
Author(s):  
Min Huang ◽  
Ju-Yuan Zhang ◽  
Xiaoli Zeng ◽  
Cheng-Cai Zhang
Keyword(s):  
Nitrogen Starvation ◽  
Filamentous Cyanobacterium ◽  
Nitrogen Deprivation ◽  
Heterocyst Differentiation ◽  
Combined Nitrogen ◽  
Heterocyst Frequency ◽  
Genes Encoding ◽  
Anabaena Sp ◽  
Opposing Effects ◽  
Pcc 7120

c-di-GMP is a ubiquitous bacterial signal regulating various physiological process. Anabaena PCC 7120 (Anabaena) is a filamentous cyanobacterium able to form regularly-spaced heterocysts for nitrogen fixation, in response to combined-nitrogen deprivation in 24h. Anabaena possesses 16 genes encoding proteins for c-di-GMP metabolism, and their functions are poorly characterized, except all2874 (cdgS) whose deletion causes a decrease in heterocyst frequency 48h after nitrogen starvation. We demonstrated here that c-di-GMP levels increased significantly in Anabaena after combined-nitrogen starvation. By inactivating each of the 16 genes, we found that the deletion of all1175 (cdgSH) led to an increase of heterocyst frequency 24h after nitrogen stepdown. A double mutant ΔcdgSHΔcdgS had an additive effect over the single mutants in regulating heterocyst frequency, indicating that the two genes acted at different time points for heterocyst spacing. Biochemical and genetic data further showed that the functions of CdgSH and CdgS in the setup or maintenance of heterocyst frequency depended on their opposing effects on the intracellular levels of c-di-GMP. Finally, we demonstrated that heterocyst differentiation was completely inhibited when c-di-GMP levels became too high or too low. Together, these results indicate that the homeostasis of c-di-GMP level is important for heterocyst differentiation in Anabaena.

scholarly journals Ferric reductase A is essential for effective iron acquisition in Paracoccus denitrificans

Microbiology ◽  
2009 ◽  
Vol 155 (4) ◽  
pp. 1294-1301 ◽  
Author(s):  
Vojtěch Sedláček ◽  
Rob J. M. van Spanning ◽  
Igor Kučera
Keyword(s):  
Paracoccus Denitrificans ◽  
Iron Acquisition ◽  
Brain Heart Infusion ◽  
Normal Growth ◽  
Kanamycin Resistance ◽  
Broad Host Range ◽  
Ferric Reductase ◽  
Wild Type ◽  
Putative Promoter Region ◽  
Broad Host Range Plasmid

Based on N-terminal sequences obtained from the purified cytoplasmic ferric reductases FerA and FerB, their corresponding genes were identified in the published genome sequence of Paracoccus denitrificans Pd1222. The ferA and ferB genes were cloned and individually inactivated by insertion of a kanamycin resistance marker, and then returned to P. denitrificans for exchange with their wild-type copies. The resulting ferA and ferB mutant strains showed normal growth in brain heart infusion broth. Unlike the ferB mutant, the strain lacking FerA did not grow on succinate minimal medium with ferric 2,3-dihydroxybenzoate as the iron source, and grew only poorly in the presence of ferric sulfate, chloride, citrate, NTA, EDTA and EGTA. Moreover, the ferA mutant strain was unable to produce catechols, which are normally detectable in supernatants from iron-limited wild-type cultures. Complementation of the ferA mutation using a derivative of the conjugative broad-host-range plasmid pEG400 that contained the whole ferA gene and its putative promoter region largely restored the wild-type phenotype. Partial, though significant, restoration could also be achieved with 1 mM chorismate added to the growth medium. The purified FerA protein acted as an NADH : FMN oxidoreductase and catalysed the FMN-mediated reductive release of iron from the ferric complex of parabactin, the major catecholate siderophore of P. denitrificans. The deduced amino acid sequence of the FerA protein has closest similarity to flavin reductases that form part of the flavin-dependent two-component monooxygenases. Taken together, our results demonstrate an essential role of reduced flavins in the utilization of exogenous ferric iron. These flavins not only provide the electrons for Fe(III) reduction but most probably also affect the rate of siderophore production.

Campylobacter jejuni gene expression in response to iron limitation and the role of Fur

Microbiology ◽  
2005 ◽  
Vol 151 (1) ◽  
pp. 243-257 ◽  
Author(s):  
Kathryn Holmes ◽  
Francis Mulholland ◽  
Bruce M. Pearson ◽  
Carmen Pin ◽  
Johanna McNicholl-Kennedy ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Iron Transport ◽  
Iron Homeostasis ◽  
Iron Acquisition ◽  
Fumarate Hydratase ◽  
Iron Limitation ◽  
Transport Systems ◽  
Wild Type ◽  
Mobility Shift

Campylobacter jejuni is a zoonotic pathogen and the most common cause of bacterial foodborne diarrhoeal illness worldwide. To establish intestinal colonization prior to either a commensal or pathogenic interaction with the host, C. jejuni will encounter iron-limited niches where there is likely to be intense competition from the host and normal microbiota for iron. To gain a better understanding of iron homeostasis and the role of ferric uptake regulator (Fur) in iron acquisition in C. jejuni, a proteomic and transcriptome analysis of wild-type and fur mutant strains in iron-rich and iron-limited growth conditions was carried out. All of the proposed iron-transport systems for haemin, ferric iron and enterochelin, as well as the putative iron-transport genes p19, Cj1658, Cj0177, Cj0178 and cfrA, were expressed at higher levels in the wild-type strain under iron limitation and in the fur mutant in iron-rich conditions, suggesting that they were regulated by Fur. Genes encoding a previously uncharacterized ABC transport system (Cj1660–Cj1663) also appeared to be Fur regulated, supporting a role for these genes in iron uptake. Several promoters containing consensus Fur boxes that were identified in a previous bioinformatics search appeared not to be regulated by iron or Fur, indicating that the Fur box consensus needs experimental refinement. Binding of purified Fur to the promoters upstream of the p19, CfrA and CeuB operons was verified using an electrophoretic mobility shift assay (EMSA). These results also implicated Fur as having a role in the regulation of several genes, including fumarate hydratase, that showed decreased expression in response to iron limitation. The known PerR promoters were also derepressed in the C. jejuni Fur mutant, suggesting that they might be co-regulated in response to iron and peroxide stress. These results provide new insights into the effects of iron on metabolism and oxidative stress response as well as the regulatory role of Fur.

scholarly journals Reductive Iron Assimilation and Intracellular Siderophores Assist Extracellular Siderophore-Driven Iron Homeostasis and Virulence

2014 ◽  
Vol 27 (8) ◽  
pp. 793-808 ◽  
Author(s):  
Bradford J. Condon ◽  
Shinichi Oide ◽  
Donna M. Gibson ◽  
Stuart B. Krasnoff ◽  
B. Gillian Turgeon
Keyword(s):  
Iron Homeostasis ◽  
Iron Acquisition ◽  
Wild Type ◽  
Peptide Synthetases ◽  
Asexual Sporulation ◽  
High Affinity ◽  
Iron Assimilation ◽  
Genes Encoding ◽  
Essential Nutrient

Iron is an essential nutrient and prudent iron acquisition and management are key traits of a successful pathogen. Fungi use nonribosomally synthesized secreted iron chelators (siderophores) or reductive iron assimilation (RIA) mechanisms to acquire iron in a high affinity manner. Previous studies with the maize pathogen Cochliobolus heterostrophus identified two genes, NPS2 and NPS6, encoding different nonribosomal peptide synthetases responsible for biosynthesis of intra- and extracellular siderophores, respectively. Deletion of NPS6 results in loss of extracellular siderophore biosynthesis, attenuated virulence, hypersensitivity to oxidative and iron-depletion stress, and reduced asexual sporulation, while nps2 mutants are phenotypically wild type in all of these traits but defective in sexual spore development when NPS2 is missing from both mating partners. Here, it is reported that nps2nps6 mutants have more severe phenotypes than both nps2 and nps6 single mutants. In contrast, mutants lacking the FTR1 or FET3 genes encoding the permease and ferroxidase components, respectively, of the alternate RIA system, are like wild type in all of the above phenotypes. However, without supplemental iron, combinatorial nps6ftr1 and nps2nps6ftr1 mutants are less virulent, are reduced in growth, and are less able to combat oxidative stress and to sporulate asexually, compared with nps6 mutants alone. These findings demonstrate that, while the role of RIA in metabolism and virulence is overshadowed by that of extracellular siderophores as a high-affinity iron acquisition mechanism in C. heterostrophus, it functions as a critical backup for the fungus.

scholarly journals A TolC-Like Protein Is Required for Heterocyst Development in Anabaena sp. Strain PCC 7120

2007 ◽  
Vol 189 (21) ◽  
pp. 7887-7895 ◽  
Author(s):  
Suncana Moslavac ◽  
Kerstin Nicolaisen ◽  
Oliver Mirus ◽  
Fadi Al Dehni ◽  
Rafael Pernil ◽  
...  
Keyword(s):  
Morphological Changes ◽  
Protein Export ◽  
Filamentous Cyanobacterium ◽  
Sole Nitrogen Source ◽  
Wild Type ◽  
Anabaena Sp ◽  
Pcc 7120 ◽  
Export System ◽  
Diazotrophic Growth ◽  
Mutant Cells

ABSTRACT The filamentous cyanobacterium Anabaena sp. strain PCC 7120 forms heterocysts in a semiregular pattern when it is grown on N2 as the sole nitrogen source. The transition from vegetative cells to heterocysts requires marked metabolic and morphological changes. We show that a trimeric pore-forming outer membrane β-barrel protein belonging to the TolC family, Alr2887, is up-regulated in developing heterocysts and is essential for diazotrophic growth. Mutants defective in Alr2887 did not form the specific glycolipid layer of the heterocyst cell wall, which is necessary to protect nitrogenase from external oxygen. Comparison of the glycolipid contents of wild-type and mutant cells indicated that the protein is not involved in the synthesis of glycolipids but might instead serve as an exporter for the glycolipid moieties or enzymes involved in glycolipid attachment. We propose that Alr2887, together with an ABC transporter like DevBCA, is part of a protein export system essential for assembly of the heterocyst glycolipid layer. We designate the alr2887 gene hgdD (heterocyst glycolipid deposition protein).

scholarly journals An amidase is required for proper intercellular communication in the filamentous cyanobacteriumAnabaenasp. PCC 7120

2017 ◽  
Vol 114 (8) ◽  
pp. E1405-E1412 ◽  
Author(s):  
Zhenggao Zheng ◽  
Amin Omairi-Nasser ◽  
Xiying Li ◽  
Chunxia Dong ◽  
Yan Lin ◽  
...  
Keyword(s):  
Intercellular Communication ◽  
Electron Tomography ◽  
Filamentous Cyanobacterium ◽  
Channel Formation ◽  
Wild Type ◽  
In The Wild ◽  
Z Ring ◽  
Green Fluorescent ◽  
20 Nm ◽  
Pcc 7120

Channels that cross cell walls and connect the cytoplasm of neighboring cells in multicellular cyanobacteria are pivotal for intercellular communication. We find that the product of the geneall1140of the filamentous cyanobacteriumAnabaenasp. PCC 7120 is required for proper channel formation.All1140encodes an amidase that hydrolyses purified peptidoglycans. An All1140-GFP fusion protein is located at the Z-ring in the periplasmic space during most of the cell cycle. Anall1140-null mutant (M40) was unable to grow diazotrophically, and no mature heterocysts were observed in the absence of combined nitrogen. Expression of two key genes,hetRandpatS, was studied in M40 using GFP as a reporter. Upon nitrogen step-down, the patterned distribution of green fluorescent cells in filaments seen in the wild type were not observed in mutant M40. Intercellular communication in M40 was studied by measuring fluorescence recovery after photobleaching (FRAP). Movement of calcein (622 Da) was aborted in M40, suggesting that the channels connecting the cytoplasm of neighboring cells are impaired in the mutant. The channels were examined with electron tomography; their diameters were nearly identical, 12.7 nm for the wild type and 12.4 nm for M40, suggesting that AmiC3 is not required for channel formation. However, when the cell wall sacculi isolated by boiling were examined by EM, the average sizes of the channels of the wild type and M40 were 20 nm and 12 nm, respectively, suggesting that the channel walls of the wild type are expandable and that this expandability requires AmiC3.

scholarly journals Contribution of the Shigella flexneri Sit, Iuc, and Feo Iron Acquisition Systems to Iron Acquisition In Vitro and in Cultured Cells

2003 ◽  
Vol 71 (4) ◽  
pp. 1919-1928 ◽  
Author(s):  
L. J. Runyen-Janecky ◽  
S. A. Reeves ◽  
E. G. Gonzales ◽  
S. M. Payne
Keyword(s):  
Iron Transport ◽  
Cultured Cells ◽  
Iron Acquisition ◽  
Shigella Flexneri ◽  
Luria Broth ◽  
Iron Chelator ◽  
Wild Type ◽  
Cell Monolayers ◽  
Serovar Typhimurium

ABSTRACT Shigella flexneri possesses multiple iron acquisition systems, including proteins involved in the synthesis and uptake of siderophores and the Feo system for ferrous iron utilization. We identified an additional S. flexneri putative iron transport gene, sitA, in a screen for S. flexneri genes that are induced in the eukaryotic intracellular environment. sitA was present in all Shigella species and in most enteroinvasive Escherichia coli strains but not in any other E. coli isolates tested. The sit locus consists of four genes encoding a potential ABC transport system. The deduced amino acid sequence of the S. flexneri sit locus was homologous to the Salmonella enterica serovar Typhimurium Sit and Yersinia pestis Yfe systems, which mediate both manganese and iron transport. The S. flexneri sit promoter was repressed by either iron or manganese, and the iron repression was partially dependent upon Fur. A sitA::cam mutation was constructed in S. flexneri. The sitA mutant showed reduced growth, relative to the wild type, in Luria broth containing an iron chelator but formed wild-type plaques on Henle cell monolayers, indicating that the sitA mutant was able to acquire iron and/or manganese in the host cell. However, mutants defective in two of these iron acquisition systems (sitA iucD, sitA feoB, and feoB iucD) formed slightly smaller plaques on Henle cell monolayers. A strain carrying mutations in sitA, feoB, and iucD did not form plaques on Henle cell monolayers.

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