Glycolipid composition of the heterocyst envelope of Anabaena sp. PCC 7120 is crucial for diazotrophic growth and relies on the UDP-galactose 4-epimerase HgdA

The nitrogenase complex in the heterocysts of the filamentous freshwater cyanobacterium Anabaena sp. PCC 7120 fixes atmospheric nitrogen to allow diazotrophic growth. The heterocyst cell envelope protects the nitrogenase from oxygen and consists of a polysaccharide and a glycolipid layer that are formed by a complex process involving the recruitment of different proteins. Here we studied the function of the putative nucleoside-diphosphate-sugar epimerase HgdA, which along with HgdB and HgdC is essential for deposition of the glycolipid layer and growth without a combined nitrogen source. Using site-directed mutagenesis and single homologous recombination approach, we performed a thoroughly functional characterization of HgdA and confirmed that the glycolipid layer of the hgdA mutant heterocyst is aberrant as shown by transmission electron microscopy and chemical analysis. The hgdA gene was expressed during late stages of the heterocyst differentiation. GFP-tagged HgdA protein localized inside the heterocysts. The purified HgdA protein had UDP-galactose 4-epimerase activity in vitro. This enzyme could be responsible for synthesis of heterocyst-specific glycolipid precursors, which could be transported over the cell wall by the ABC transporter components HgdB/HgdC.

Amino acid transport systems required for diazotrophic growth in the cyanobacterium Anabaena sp. strain PCC 7120.

Journal of Bacteriology ◽

10.1128/jb.177.11.3150-3157.1995 ◽

1995 ◽

Vol 177 (11) ◽

pp. 3150-3157 ◽

Cited By ~ 54

Author(s):

M L Montesinos ◽

A Herrero ◽

E Flores

Keyword(s):

Amino Acid ◽

Amino Acid Transport ◽

Transport Systems ◽

Acid Transport ◽

Cyanobacterium Anabaena ◽

Anabaena Sp ◽

Pcc 7120 ◽

Heterocyst-specific flavodiiron protein Flv3B enables oxic diazotrophic growth of the filamentous cyanobacterium Anabaena sp. PCC 7120

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1407327111 ◽

2014 ◽

Vol 111 (30) ◽

pp. 11205-11210 ◽

Cited By ~ 37

Author(s):

M. Ermakova ◽

N. Battchikova ◽

P. Richaud ◽

H. Leino ◽

S. Kosourov ◽

...

Keyword(s):

Filamentous Cyanobacterium ◽

Cyanobacterium Anabaena ◽

Anabaena Sp ◽

Pcc 7120 ◽

NrrA Directly Regulates Expression of hetR during Heterocyst Differentiation in the Cyanobacterium Anabaena sp. Strain PCC 7120

Journal of Bacteriology ◽

10.1128/jb.01314-06 ◽

2006 ◽

Vol 188 (24) ◽

pp. 8520-8525 ◽

Cited By ~ 62

Author(s):

Shigeki Ehira ◽

Masayuki Ohmori

Keyword(s):

Transcription Initiation ◽

Response Regulator ◽

Nitrogen Deprivation ◽

Heterocyst Differentiation ◽

Combined Nitrogen ◽

Regulatory Cascade ◽

Cyanobacterium Anabaena ◽

Anabaena Sp ◽

ABSTRACT Heterocyst differentiation in the cyanobacterium Anabaena sp. strain PCC 7120 requires NtcA, the global nitrogen regulator in cyanobacteria, and HetR, the master regulator of heterocyst differentiation. Expression of hetR is upregulated by nitrogen deprivation, and its upregulation depends on NtcA. However, it has not yet been revealed how NtcA regulates the expression of hetR. In the experiments presented here, it was confirmed that NrrA (All4312), a nitrogen-responsive response regulator, was required for the upregulation of hetR. The use of the nitrogen-responsive transcription initiation sites (TISs) for the hetR gene depended upon NrrA. NrrA bound specifically to the region upstream of TISs located at positions −728 and −696 in vitro. Overexpression of nrrA resulted in enhanced hetR expression and heterocyst formation. A molecular regulatory cascade is proposed whereby NtcA upregulates the expression of nrrA upon limitation of combined nitrogen in the medium and then NrrA upregulates the expression of hetR, leading to heterocyst differentiation.

Cell Envelope Components Influencing Filament Length in the Heterocyst-Forming Cyanobacterium Anabaena sp. Strain PCC 7120

Journal of Bacteriology ◽

10.1128/jb.02128-14 ◽

2014 ◽

Vol 196 (23) ◽

pp. 4026-4035 ◽

Cited By ~ 14

Author(s):

M. Burnat ◽

E. Schleiff ◽

E. Flores

Keyword(s):

Cell Envelope ◽

Filament Length ◽

Cyanobacterium Anabaena ◽

Anabaena Sp ◽

The pknH gene restrictively expressed in heterocysts is required for diazotrophic growth in the cyanobacterium Anabaena sp. strain PCC 7120

Microbiology ◽

10.1099/mic.0.057729-0 ◽

2012 ◽

Vol 158 (6) ◽

pp. 1437-1443 ◽

Cited By ~ 12

Author(s):

Shigeki Ehira ◽

Masayuki Ohmori

Keyword(s):

Cyanobacterium Anabaena ◽

Anabaena Sp ◽

Pcc 7120 ◽

Cold-stress-altered phosphorylation of EF-Tu in the cyanobacterium Anabaena sp. strain PCC 7120

Canadian Journal of Microbiology ◽

10.1139/w07-030 ◽

2007 ◽

Vol 53 (5) ◽

pp. 551-558 ◽

Cited By ~ 4

Author(s):

Bassam El-Fahmawi ◽

George W. Owttrim

Keyword(s):

Cold Stress ◽

Prolonged Exposure ◽

De Novo ◽

Affinity Purification ◽

Translation Elongation ◽

Cyanobacterium Anabaena ◽

Anabaena Sp ◽

Growth of prokaryotes at reduced temperature results in the formation of a cold-adapted ribosome through association with de novo synthesized polypeptides. In vitro and in vivo phosphorylation studies combined with affinity purification and mass spectrometry identified that the phosphorylation status of translation elongation factor EF-Tu was altered in response to cold stress in the photosynthetic, Gram-negative cyanobacterium Anabaena sp. strain PCC 7120. In response to a temperature downshift from 30 to 20 °C, EF-Tu was rapidly and transiently hyperphosphorylated during the acclimation phase followed by a reduction in phosphorylation below background levels in response to prolonged exposure. EF-Tu was identified as a phosphothreonine protein. Unexpectedly, ribosomal protein S2 was also observed to be a phosphoprotein continuously phosphorylated during cold stress. The phosphorylation status of EF-Tu has previously been associated with translational regulation in other systems, with a reduction in translation elongation occurring in response to phosphorylation. These results provide evidence for a novel mechanism by which translation is initially downregulated in response to cold stress in Anabaena.

A novel glutaredoxin domain-containing peroxiredoxin ‘All1541’ protects the N2-fixing cyanobacterium Anabaena PCC 7120 from oxidative stress

Biochemical Journal ◽

10.1042/bj20111877 ◽

2012 ◽

Vol 442 (3) ◽

pp. 671-680 ◽

Cited By ~ 23

Author(s):

Manisha Banerjee ◽

Anand Ballal ◽

Shree K. Apte

Keyword(s):

Oxidative Stress ◽

Peroxidase Activity ◽

Bioinformatic Analysis ◽

Open Reading Frames ◽

Site Directed Mutagenesis ◽

Cysteine Residues ◽

Anabaena Pcc 7120 ◽

Cyanobacterium Anabaena ◽

Prxs (peroxiredoxins) are ubiquitous thiol-based peroxidases that detoxify toxic peroxides. The Anabaena PCC 7120 genome harbours seven genes/ORFs (open reading frames) which have homology with Prxs. One of these (all1541) was identified to encode a novel Grx (glutaredoxin) domain-containing Prx by bioinformatic analysis. A recombinant N-terminal histidine-tagged All1541 protein was overexpressed in Escherichia coli and purified. Analysis with the protein alkylating agent AMS (4-acetamido-4′-maleimidyl-stilbene-2,2′-disulfonate) showed All1541 to form an intra-molecular disulfide bond. The All1541 protein used glutathione (GSH) more efficiently than Trx (thioredoxin) to detoxify H2O2. Deletion of the Grx domain from All1541 resulted in loss of GSH-dependent peroxidase activity. Employing site-directed mutagenesis, the cysteine residues at positions 50 and 75 were identified as peroxidatic and resolving cysteine residues respectively, whereas both the cysteine residues within the Grx domain (positions 181 and 184) were shown to be essential for GSH-dependent peroxidase activity. On the basis of these data, a reaction mechanism has been proposed for All1541. In vitro All1541 protein protected plasmid DNA from oxidative damage. In Anabaena PCC 7120, all1541 was transcriptionally activated under oxidative stress. Recombinant Anabaena PCC 7120 strain overexpressing All1541 protein showed superior oxidative stress tolerance to H2O2 as compared with the wild-type strain. The results suggest that the glutathione-dependent peroxidase All1541 plays an important role in protecting Anabaena from oxidative stress.

NrrA directly regulates expression of the fraF gene and antisense RNAs for fraE in the heterocyst-forming cyanobacterium Anabaena sp. strain PCC 7120

Microbiology ◽

10.1099/mic.0.076703-0 ◽

2014 ◽

Vol 160 (5) ◽

pp. 844-850 ◽

Cited By ~ 5

Author(s):

Shigeki Ehira ◽

Masayuki Ohmori

Keyword(s):

Response Regulator ◽

Repeat Sequence ◽

Nitrogen Deprivation ◽

Antisense Rnas ◽

Regulated Expression ◽

Cyanobacterium Anabaena ◽

Anabaena Sp ◽

Dna Strand ◽

Pcc 7120 ◽

The heterocystous cyanobacterium Anabaena sp. strain PCC 7120 grows as linear multicellular filaments that can contain hundreds of cells. Heterocysts, which are specialized cells for nitrogen fixation, are regularly intercalated among photosynthetic vegetative cells, and these cells are metabolically dependent on each other. Thus, multicellularity is essential for diazotrophic growth of heterocystous cyanobacteria. In Anabaena sp. strain PCC 7120, the fraF gene, which is required to limit filament length, is induced by nitrogen deprivation. The fraF transcripts extend to the fraE gene, which lies on the opposite DNA strand and could possess dual functionality, mRNAs for fraF and antisense RNAs for fraE. In the present study, we found that NrrA, a nitrogen-regulated response regulator, directly regulated expression of fraF. Induction of fraF by nitrogen deprivation was abolished by the nrrA disruption. NrrA specifically bound to the promoter region of fraF, and recognized an inverted repeat sequence. Thus, it is concluded that NrrA controls expression of mRNAs for fraF and antisense RNAs for fraE in response to nitrogen deprivation.

Expanding the Natural Products Heterologous Expression Repertoire in the Model Cyanobacterium Anabaena sp. Strain PCC 7120: Production of Pendolmycin and Teleocidin B-4

10.26434/chemrxiv.11316098.v1 ◽

2019 ◽

Cited By ~ 1

Author(s):

Patrick Videau ◽

Kaitlyn Wells ◽

Arun Singh ◽

Jessie Eiting ◽

Philip Proteau ◽

...

Keyword(s):

Natural Products ◽

Genome Mining ◽

Gene Clusters ◽

Combinatorial Biosynthesis ◽

Test Case ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Cyanobacterium Anabaena ◽

Anabaena Sp ◽

Cyanobacteria are prolific producers of natural products and genome mining has shown that many orphan biosynthetic gene clusters can be found in sequenced cyanobacterial genomes. New tools and methodologies are required to investigate these biosynthetic gene clusters and here we present the use of <i>Anabaena </i>sp. strain PCC 7120 as a host for combinatorial biosynthesis of natural products using the indolactam natural products (lyngbyatoxin A, pendolmycin, and teleocidin B-4) as a test case. We were able to successfully produce all three compounds using codon optimized genes from Actinobacteria. We also introduce a new plasmid backbone based on the native <i>Anabaena</i>7120 plasmid pCC7120ζ and show that production of teleocidin B-4 can be accomplished using a two-plasmid system, which can be introduced by co-conjugation.