Shivam Yadav

UV-B stress induced metabolic rearrangements explored with comparative proteomics in three Anabaena species.

Comparative proteomics together with physiological variables revealed different responses among three species of diazotrophic cyanobacterium Anabaena exposed to UV-B stress at the same time points. Perceptible decline in PSII activity, ATP pool, nitrogenase activity and respiration rate was observed for all the three species; this being maximum in Anabaena doliolum, followed by Anabaena sp. PCC 7120 and minimum in Anabaena L31. Statistical analysis of the protein abundance divided majority of them as early accumulated in A. L31, late accumulated in A. sp. PCC 7120 and downregulated in A. doliolum. Tolerance of A. L31 may be ascribed to post-translational modification reflected through the highest number of protein isoforms in its proteome followed by A. PCC 7120 and A. doliolum. Furthermore, increase in abundance of cyanophycinase, glutamine synthetase and succinate semialdehyde dehydrogenase in A. L31 suggests operation of an alternate pathway for assimilation of nitrogen and carbon under UV-B stress. An early accumulation of four proteins viz., glutamate ammonia ligase (Alr2328), transketolase (Alr3344), inorganic pyrophosphatase (All3570), and trigger protein (Alr3681) involved respectively in amino acid metabolism, energy metabolism, biosynthesis of cofactor and trigger protein and chaperone like activity across three species, suggests them to be marker of UV-B stress in Anabaena spp. This article is part of a Special Issue entitled: Proteomics in India.

A Novel Aldo-Keto Reductase (AKR17A1) of Anabaena sp. PCC 7120 Degrades the Rice Field Herbicide Butachlor and Confers Tolerance to Abiotic Stresses in E. coli

Present study deals with the identification of a novel aldo/keto reductase, AKR17A1 from Anabaena sp. PCC7120 and adds on as 17th family of AKR superfamily drawn from a wide variety of organisms. AKR17A1 shares many characteristics of a typical AKR such as— (i) conferring tolerance to multiple stresses like heat, UV-B, and cadmium, (ii) excellent activity towards known AKR substrates (isatin and 2-nitrobenzaldehyde), and (iii) obligate dependence on NADPH as a cofactor for enzyme activity. The most novel attribute of AKR17A1, first reported in this study, is its capability to metabolize butachlor, a persistent rice field herbicide that adversely affects agro-ecosystem and non-target organisms. The AKR17A1 catalyzed- degradation of butachlor resulted into formation of 1,2-benzene dicarboxylic acid and 2,6 bis (1,1, dimethylethyl) 4,-methyl phenol as the major products confirmed by GC-MS analysis.

Overexpression of phytochelatin synthase ( pcs ) enhances abiotic stress tolerance by altering the proteome of transformed Anabaena sp. PCC 7120

The present study provides data on the insertion of an extra copy of phytochelatin synthase (alr0975) in Anabaena sp. PCC 7120. The recombinant strain (AnFPN-pcs) compared to wild type showed approximately 22.3% increase in growth rate under UV-B, NaCl, heat, CuCl2, carbofuran, and CdCl2. It also registered 2.25-fold enhanced nitrogenase activity and 5-fold higher phytochelatin production. A comparison of the protein profile of wild type with the recombinant strain revealed that recombinant strain accumulated proteins belonging to the following categories: (i) detoxification (nutrient stress induced DNA binding protein, Mn-SOD, Alr0946 (CalA)), (ii) protein folding and modification (molecular chaperone DnaK, FKBP-type peptidyl-prolyl cis-trans isomerase), (iii) nucleotide and amino acid biosynthesis (dihydroorotase and Ketol-acid reductoisomerase), (iv) photosynthesis and respiration (coproporphyrinogen III oxidase, phycocyanin alpha chain, ferredoxin-NADP+ reductase), and (v) transport (sugar transport ATP-binding protein). Thus, it can be concluded that, above category proteins with their respective role in scavenging reactive oxygen species, proper folding of unfolded proteins, and protection of protein from degradation, sustained carbon fixation and energy pool and active transport of sugar together conceivably help the recombinant cyanobacterium (AnFPN-pcs) to cope with abiotic stress employed in the present study. Such recombinant strains have potential for future use as biofertilizer.

Comparative proteomics of wild type, An+ahpC and An∆ahpC strains of Anabaena sp. PCC7120 demonstrates AhpC mediated augmentation of photosynthesis, N2-fixation and modulation of regulatory network of antioxidative proteins.

UNLABELLED Alkylhydroperoxide reductase (AhpC), a 1-Cys peroxiredoxin is well known for maintaining the cellular homeostasis. Present study employs proteome approach to analyze and compare alterations in proteome of Anabaena PCC7120 in overexpressing (An+ahpC), deletion (An∆ahpC) and its wild type. 2-DE based analysis revealed that the major portion of identified protein belongs to energy metabolism, protein folding, modification and stress related proteins and carbohydrate metabolism. The two major traits discernible from An+ahpC were (i) augmentation of photosynthesis and nitrogen fixation (ii) modulation of regulatory network of antioxidative proteins. Increased accumulation of proteins of light reaction, dark reaction, pentose phosphate pathway and electron transfer agent FDX for nitrogenase in An+ahpC and their simultaneous downregulation in AnΔahpC demonstrates its role in augmenting photosynthesis and nitrogen fixation. Proteomic data was nicely corroborated with physiological, biochemical parameters displaying upregulation of nitrogenase (1.6 fold) PSI (1.08) and PSII (2.137) in An+ahpC. Furthermore, in silico analysis not only attested association of AhpC with peroxiredoxins but also with other players of antioxidative defense system viz. thioredoxin and thioredoxin reductase. Above mentioned findings are in agreement with 33-40% and 40-60% better growth performance of An+ahpC over wild type and An∆ahpC respectively under abiotic stresses, suggesting its role in maintenance of metabolic machinery under stress. SIGNIFICANCE Present work explores key role of AhpC in mitigating stress in Anabaena PCC7120 through combined proteomic, biochemical and in silico investigations. This study is the first attempt to analyze and compare alterations in proteome of Anabaena PCC7120 following addition (overexpressing strain An+ahpC) and deletion (mutant An∆ahpC) of AhpC against its wild type. The effort resulted in two major traits in An+ahpC as (i) augmentation of photosynthesis and nitrogen fixation (ii) modulation of regulatory network of antioxidative proteins.

Effects of UV-B and heavy metals on nitrogen and phosphorus metabolism in three cyanobacteria.

Cyanobacteria sp. (diazotrophic and planktonic) hold a major position in ecosystem, former one due to their intrinsic capability of N2‐fixation and later because of mineralization of organic matter. Unfortunately, their exposure to variety of abiotic stresses is unavoidable. Comparative analysis of interactive effect of UV‐B and heavy metals (Cd/Zn) on nitrogen and phosphorus metabolism of three cyanobacteria (Anabaena, Microcystis, Nostoc) revealed additive inhibition (χ2 significant p < 0.05) of NH4+ and PO43− uptake whereas increase in nitrate uptake upon UV‐B + heavy metal exposure. Glutamine synthetase and Alkaline phosphatase activity was reduced after all treatments whereas Nitrate reductase activity showed slight stimulation in UV‐B and UV‐B + heavy metals treatment. Combination of UV‐B and metals seems more detrimental to the NH4+, PO43− uptake, GS and APA activity. A significant stimulation in NO3− uptake and NR activity following exposure to UV‐B in all the three cyanobacteria suggests UV‐B‐induced structural change(s) in the enzyme/carriers. Metals seem to compete for the binding sites of the enzymes and carriers; as noticed for Anabaena and Microcystis showing change in Km while no change in the Km value of Nostoc suggests non‐competitive nutrient uptake. Higher accumulation and more adverse effect on Na+ and K+ efflux proposes Cd as more toxic compared to Zn.

Exploring the membrane proteome of the diazotropic cyanobacterium Anabaena PCC7120 through gel-based proteomics and in silico approaches ☆

UNLABELLED This paper focuses on the gel-based membrane proteomics from diazotrophic cyanobacterium Anabaena PCC7120 by modifying the protocol of Hall et al. [1]. The bioinformatic analysis revealed that 59 (29 integral, 30 peripheral) of the 67 proteins identified were membrane proteins. Of the 29 integral proteins, except Alr0834, the remaining 28 contained 1-12 transmembrane helices. Sixteen integral proteins harboring signal peptides (Sec/TAT/LipoP) suggest that protein targeting in Anabaena involves both sec-dependent and sec-independent pathways. While majority of photosynthesis and respiration proteins (21 of 24) were confined to broad pH gradient the hypothetical and unknown (12 of 13), and cell envelope proteins (3 of 3) preferred the narrow pH range. Of the 5 transporters and binding proteins, Na(+)/H(+)-exchanging protein and Alr2372 were present in broad, pstS1 and cmpD in narrow and cmpA was common to both pH ranges. The distribution of proteins across pH gradient, thus clearly indicates the functional and structural diversity in membrane proteome of Anabaena. It requires mention that protochlorophyllide oxido-reductase, Na(+)/H(+)-exchanging protein, All1355, Alr2055, Alr3514, Alr2903 and Alr2751 were new entries to the 2DE membrane protein profile of Anabaena. This study demonstrates suitability of the modified protocol for the study of membrane protein from filamentous cyanobacteria. SIGNIFICANCE Anabaena sp. PCC7120 is used as a model organism due to its agriculture significance as biofertilizer, close resemblance with higher plant chloroplast and availability of full genome sequence. Although cytosolic proteome has been explored a lot membrane proteins are still understudied as they are notoriously difficult to display using 2-D technology. Identification and characterization of these proteins is therefore required to elucidate and understand cellular mechanisms. The purpose of this study was to develop a protocol suitable for membrane protein extraction from Anabaena. Additionally, by homology comparison or domain assignment a possible function could be ascribed to novel uncharacterized proteins which will serve as a useful reference for further detailed studies of membrane system in filamentous cyanobacteria. Resolution of membrane proteins ranging from least (single transmembrane helix) to highly hydrophobic (several transmembrane helices) one on 2D gels recommends the gel based approach for identification of membrane proteomics from filamentous cyanobacteria. This article is part of a Special Issue entitled: Proteomics in India.

Role of Algae as a Biofertilizer

Sustainable agriculture is advantageous over conventional agriculture for its capacity to accomplish food demand by utilizing environmental resources without negatively affecting it. The beneficiary role of blue-green algae (BGA) by way of supporting the nitrogen economy of paddy fields and enhancing rice productivity is well documented. The simple presence of BGA in soil results in formation of soil aggregates, which reduces soil loss during rainy season and regulates aeration, soil temperature, hence, improving physical and chemical properties of soil vis-a-vis physical environment of the crop. BGA are helpful in restoring soil nutrients by secreting exopolysaccharides and bioactive substances. They have the potential to mobilize insoluble forms of inorganic phosphates. Algalization has been employed for reducing the amount of exchangeable sodium, which results in altered soil pH and leads to reclamation of sodic soils. Some red algae used as biofertilizers have been found to augment growth nutritional value and yield of crop plants. This chapter provides an overview of the role of algae as biofertilizers.

Signal Perception and Mechanism of Salt Toxicity/Tolerance in Photosynthetic Organisms: Cyanobacteria to Plants

High salt concentration represents one of the most significant abiotic constraints, affecting all life forms including plants and cyanobacteria. Soil salinity curtails plant growth by way of osmotic, ionic and oxidative stresses resulting in multiple inhibitory effects on various physiological processes such as growth, photosynthesis, respiration and cellular metabolism. In order to combat high salinity, various adaptive strategies employed include ion homeostasis achieved by ion transport and compartmentalization of injurious ions, osmotic homeostasis by accumulation of compatible solutes/osmolytes and upregulation of antioxidant defence mechanism. The aforesaid processes are executed through SOS and MAPK signalling pathways leading to modulation of gene expression. Salt stress signal transduction pathways initiate through sensing extracellular Na+ ions causing modification of constitutively expressed transcription factors. This modification is responsible for expression of early transcriptional activators such as CBF/DREB gene family which eventually activate stress tolerance effector genes such as osmolyte biosynthesis genes, detoxification enzymes, and chaperones. Various genes/cDNAs encoding proteins involved in these adaptive mechanisms have been isolated and identified. Bioinformatic predictions through docking revealed interaction of salt across the species at conserved domains and motifs as a possible mechanism for response of a particular protein under salt stress. In this chapter, major aspects of salt stress are reviewed with emphasis on its detrimental consequences and biochemical and molecular mechanisms of signal transduction in plants and cyanobacteria under high salinity.

Cyanobacterial Biodiversity and Biotechnology: A Promising Approach for Crop Improvement

Abstract Cyanobacteria due to their remarkable evolutionary advances such as the presence of oxygenic photosynthesis have been considered as an ideal system for plant-based studies in order to assess fundamental biochemical processes like carbon and nitrogen assimilation and photosynthesis processes. Moreover, the exclusive ability of both photosynthesis and nitrogen fixation together with adaptability to various environmental fluctuations of few genera makes them ubiquitous. Cyanobacterial genes involved in carbon metabolism, fatty acid biosynthesis, and pigment biosynthesis have been exploited as substitute for homologous gene sources, targeting enhanced plant productivity and nutritional values. Present chapter sheds light on key contributions of cyanobacterial biodiversity and biotechnology along with the future prospects for developing transgenic crops of high yield and nutritive value utilizing cyanobacterial genes.

Nickel and arsenite-induced differential oxidative stress and antioxidant responses in two Anabaena species

In recent years, release of chemical pollutants has increased due to anthropogenic activities. Heterocystous filamentous cyanobacteria constitute dominant paddy microflora and are excellent biofertilizers augmenting rice productivity. Cyanobacteria are frequently exposed to toxic metals, nickel and arsenic are one of the major toxicants present. We exposed two species of diazotrophic cyanobacteria Anabaena sp. PCC 7120 and Anabaena doliolum, to sub‐lethal concentrations (15.0 and 9.0 μM) of Ni2+ and (17.0 and 11 mM) of arsenite (AsIII) and analyzed at different days of treatments (0, 1, 7, and 15 days) for oxidative damage and antioxidative biomarkers. Lipid peroxidation was enhanced (1.5‐ to 2.5‐fold increase in MDA content), indicating damaging effects of Ni2+ and As(III) on membrane. Although Ni2+ and As(III), both induced oxidative stress in both species, Anabaena PCC 7120 experienced less stress than A. doliolum. This could be explained by a higher activity of antioxidant enzymes catalase (CAT), ascorbate peroxidase (APX), glutathione reductase (GR) in Anabaena PCC 7120 (4.6‐, 2.0‐ and 1.4‐fold [Ni2+] 3.2‐, 2.5‐, and 2.08‐fold [As]) compared to A. doliolum (4.2‐, 2.5‐, and 1.3‐fold [Ni2+] and 3.2‐, 3.33‐, and 1.8‐fold [As]). Moreover, superoxide dismutase registered less inhibition in Anabaena sp. PCC 7120 (1.5 and 1.8) compared to A. doliolum (1.8 and 2.3) under Ni2+ and As(III) stress. In addition to, IBR revealed that As(III) imposes severe impact on both strain, however, A. doliolum suffers most. Therefore, the study demonstrates interspecies variation in survival strategy of two Anabaena species and difference in potential of two different toxicants to produce oxidative stress.