| 1. |
- Bygdell, Joakim, et al.
(författare)
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Protein expression in tension wood formation monitored at high tissue resolution in Populus
- 2017
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Ingår i: Journal of Experimental Botany. - : Oxford University Press. - 0022-0957 .- 1460-2431. ; 68:13, s. 3405-3417
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Tidskriftsartikel (refereegranskat)abstract
- Tension wood (TW) is a specialized tissue with contractile properties that is formed by the vascular cambium in response to gravitational stimuli. We quantitatively analysed the proteomes of Populus tremula cambium and its xylem cell derivatives in stems forming normal wood (NW) and TW to reveal the mechanisms underlying TW formation. Phloem-, cambium-, and wood-forming tissues were sampled by tangential cryosectioning and pooled into nine independent samples. The proteomes of TW and NW samples were similar in the phloem and cambium samples, but diverged early during xylogenesis, demonstrating that reprogramming is an integral part of TW formation. For example, 14-3-3, reactive oxygen species, ribosomal and ATPase complex proteins were found to be up-regulated at early stages of xylem differentiation during TW formation. At later stages of xylem differentiation, proteins involved in the biosynthesis of cellulose and enzymes involved in the biosynthesis of rhamnogalacturonan-I, rhamnogalacturonan-II, arabinogalactan-II and fasciclin-like arabinogalactan proteins were up-regulated in TW. Surprisingly, two isoforms of exostosin family proteins with putative xylan xylosyl transferase function and several lignin biosynthesis proteins were also up-regulated, even though xylan and lignin are known to be less abundant in TW than in NW. These data provided new insight into the processes behind TW formation.
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| 2. |
- Kaur, S., et al.
(författare)
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Biochemical and proteomic analysis reveals oxidative stress tolerance strategies of Scenedesmus abundans against allelochemicals released by Microcystis aeruginosa
- 2019
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Ingår i: Algal Research. - : Elsevier B.V.. - 2211-9264. ; 41
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Tidskriftsartikel (refereegranskat)abstract
- We studied the possible survival strategies of a green alga, Scenedesmus abundans, against allelochemicals secreted by Microcystis aeruginosa. We exposed the monoculture of S. abundans to a cell free-filtrate (allelochemicals)of M. aeruginosa at the start of our experiment and measured the growth behaviour, morphological changes and oxidative stress markers. The results suggest that exposure to allelochemicals induced oxidative stress in S. abundans, which had significantly reduced the growth of green alga with certain morphological changes. However, after seven days, S. abundans found ways to reduce oxidative stress by recovering its morphology and growth close to that of control. To understand possible survival strategies of test alga, we measured biochemical as well as protein level changes in S. abundans. Biochemical response of the green alga clearly showed that as a response to allelochemicals, enzymatic and non-enzymatic antioxidants were induced. Proteomic analysis showed that exposure to allelochemicals induced accumulation of 13 proteins on the 2-DE gel of S. abundans, which falls in three functional categories, i.e., (i)energy metabolism (photosynthesis, carbon fixation and respiration), (ii)ROS scavenging enzymes and molecular chaperones, and (iii)amino acid and protein biosynthesis. After chronic oxidative stress, these proteins presumably retained glycolysis, pentose phosphate pathway and turnover rate of the Calvin-Benson cycle. Moreover, these proteins assisted in the adequate detoxification of ROS and played an important role in the damage removal and repair of oxidized proteins, lipids and nucleic acids. Therefore, our study anticipates that S. abundans embraces biochemical and proteomic reprogramming to thrives against allelochemicals released by M. aeruginosa.
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| 3. |
- Srivastava, Akanksha, et al.
(författare)
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Acute cadmium toxicity and post-stress recovery : Insights into coordinated and integrated response/recovery strategies of Anabaena sp. PCC 7120
- 2021
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Ingår i: Journal of Hazardous Materials. - : Elsevier BV. - 0304-3894 .- 1873-3336. ; 411
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Tidskriftsartikel (refereegranskat)abstract
- Cyanobacteria, the first photoautotrophs have remarkable adaptive capabilities against most abiotic stresses, including Cd. A model cyanobacterium, Anabaena sp. PCC 7120 has been commonly used to understand cyanobacterial plasticity under different environmental stresses. However, very few studies have focused on the acute Cd toxicity. In this context, Anabaena was subjected to 100 ?M Cd for 48 h (acute Cd stress, ACdS) and then transferred into the fresh medium for post-stress recovery (PSR). We further investigated the dynamics of morpho-ultrastructure, physiology, cytosolic proteome, thylakoidal complexes, chelators, and transporters after ACdS, as well as during early (ER), mid (MR), and late (LR) phases of PSR. The findings revealed that ACdS induced intracellular Cd accumulation and ROS production, altered morpho-ultrastructure, reduced photosynthetic pigments, and affected the structural organization of PSII, which subsequently hindered photosynthetic efficiency. Anabaena responded to ACdS and recovered during PSR by reprogramming the expression pattern of proteins/genes involved in cellular defense and repair; CO2 access, Calvin-Benson cycle, glycolysis, and pentose phosphate pathway; protein biosynthesis, folding, and degradation; regulatory functions; PSI-based cyclic electron flow; Cd chelation; and efflux. These modulations occurred in an integrated and coordinated manner that facilitated Anabaena to detoxify Cd and repair ACdS-induced cellular damage.
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| 4. |
- Srivastava, Akanksha, et al.
(författare)
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Cd-induced cytosolic proteome changes in the cyanobacterium Anabaena sp. PCC7120 are mediated by LexA as one of the regulatory proteins
- 2023
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Ingår i: Biochimica et Biophysica Acta - Proteins and Proteomics. - : Elsevier BV. - 1570-9639 .- 1878-1454. ; 1871:3, s. 140902-
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Tidskriftsartikel (refereegranskat)abstract
- LexA, a well-characterized transcriptional repressor of SOS genes in heterotrophic bacteria, has been shown to regulate diverse genes in cyanobacteria. An earlier study showed that LexA overexpression in a cyanobacterium, Anabaena sp. PCC7120 reduces its tolerance to Cd stress. This was later shown to be due to modulation of photosynthetic redox poising by LexA under Cd stress. However, due to the global regulatory nature of LexA and the prior prediction of AnLexA-box in a few heavy metal-responsive genes, we speculated that LexA has a broad role in Cd tolerance, with regulation over a variety of Cd stress-responsive genes in addition to photosynthetic genes. Thus, to further expand the knowledge on the regulatory role of LexA in Cd stress tolerance, a cytosolic proteome profiling of Anabaena constitutively overexpressing LexA upon Cd stress was performed. The proteomic study revealed 25 differentially accumulated proteins (DAPs) in response to the combined effect of LexA over -expression and Cd stress, and the other 11 DAPs exclusively in response to either LexA overexpression or Cd stress. The 36 identified proteins were related with a variety of functions, including photosynthesis, C -meta-bolism, antioxidants, protein turnover, post-transcriptional modifications, and a few unknown and hypothetical proteins. The regulation of LexA on corresponding genes, and six previously reported Cd efflux transporters, was further validated by the presence of AnLexA-boxes, transcript, and/or promoter analyses. In a nutshell, this study identifies the regulation of Anabaena LexA on several Cd stress-responsive genes of various functions, hence expanding the regulatory role of LexA under Cd stress.
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| 5. |
- Srivastava, A., et al.
(författare)
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Gamma (γ)-radiation stress response of the cyanobacterium Anabaena sp. PCC7120 : Regulatory role of LexA and photophysiological changes
- 2023
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Ingår i: Plant Science. - : Elsevier BV. - 0168-9452 .- 1873-2259. ; 326, s. 111529-
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Tidskriftsartikel (refereegranskat)abstract
- High radioresistance of the cyanobacterium, Anabaena sp. PCC7120 has been attributed to efficient DNA repair, protein recycling, and oxidative stress management. However, the regulatory network involved in these batteries of responses remains unexplored. In the present study, the role of a global regulator, LexA in modulating gamma (γ)-radiation stress response of Anabaena was investigated. Comparison of the cytosolic proteome profiles upon γ-radiation in recombinant Anabaena strains, AnpAM (vector-control) and AnlexA+ (LexA-overexpressing), revealed 41 differentially accumulated proteins, corresponding to 29 distinct proteins. LexA was found to be involved in the regulation of 27 of the corresponding genes based on the presence of AnLexA-Box, EMSA, and/or qRT-PCR studies. The majority of the regulated genes were found to be involved in C-assimilation either through photosynthesis or C-catabolism and oxidative stress alleviation. Photosynthesis, measured in terms of PSII photophysiological parameters and thylakoid membrane proteome was found to be affected by γ-radiation in both AnpAM and AnlexA+ cells, with LexA affecting them even under control growth conditions. Thus, LexA functioned as one of the transcriptional regulators involved in modulating γ-radiation stress response in Anabaena. This study could pave the way for a deeper understanding of the regulation of γ-radiation-responsive genes in cyanobacteria at large.
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| 6. |
- Srivastava, Akanksha, et al.
(författare)
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Physiological and thylakoid proteome analyses of Anabaena sp. PCC 7120 for monitoring the photosynthetic responses under cadmium stress
- 2021
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Ingår i: Algal Research. - : Elsevier BV. - 2211-9264. ; 54
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Tidskriftsartikel (refereegranskat)abstract
- Photosynthetic organisms are highly susceptible to cadmium (Cd) as it interferes with the structural and functional aspects of the photosynthesis. As a major group of photosynthetic prokaryotes, cyanobacteria are also affected by Cd. The inhibitory effects of Cd on cyanobacterial photosynthesis have been studied from an early stage, but the mechanism of Cd toxicity is still unclear. Therefore, we investigated the photosynthetic responses of Anabaena sp. PCC 7120 under Cd stress (LC50) at physiological and thylakoid proteome levels via chlorophyll a fluorescence measurements and blue native (BN)-SDS PAGE, respectively. The findings revealed that Cd exposure triggered the intracellular ROS production and negatively affected the photosynthetic performance. With Cd exposure, effective photochemical quantum yield of PSII (Y(II)) and photochemical quenching efficiency (qP and qL) were significantly reduced, whereas the non-regulated energy dissipation (Y(NO)) was increased. Fast fluorescence kinetic measurements showed that Cd exposure resulted in the inactivation of PSII reaction centres, which interfered with the transfer of energy from antenna complexes to reaction centre, decreased the ability to convert excitation energy into electron transport, increased thermal dissipation, and thus decreased linear electron flow efficiency. Conversely, the efficiency of electron transfer from intermediate carriers to final PSI acceptors was increased, which might be related to the induction of PSI-based cyclic electron flow. This assumption was confirmed by an enhanced post-illumination fluorescence transient. Consistent with these observations, BN-SDS PAGE showed that Cd exposure significantly decreased PSII subunit proteins accumulation, but slightly increased PSI proteins accumulation. Additionally, only minor reductions in the accumulation of ATP synthase/NDH-1 and Cytochrome b6f subunit proteins were observed. The transcript levels of most of the selected subunit proteins were in accordance with BN-SDS PAGE. Collectively, our experiments show that a shift to PSI-based cyclic electron flow was a protective strategy of Anabaena sp. PCC 7120 against Cd-induced PSII photo-damage.
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| 7. |
- Srivastava, Akanksha, et al.
(författare)
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Regulatory role of LexA in modulating photosynthetic redox poise and cadmium stress tolerance in the cyanobacterium, Anabaena sp. PCC7120
- 2022
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Ingår i: Environmental and Experimental Botany. - : Elsevier BV. - 0098-8472 .- 1873-7307. ; 195
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Tidskriftsartikel (refereegranskat)abstract
- Strategies developed by organisms to overcome disruption in redox poise of photosynthetic electron transport chain (pETC) are important for its survival under abiotic stress. The process needs to be tightly regulated for optimal functioning. While the redox poising processes are well known in cyanobacteria, understanding of their regulatory network is lacking. Since LexA is one of the known global regulators of stress response in the cyanobacterium Anabaena sp. PCC7120, its role in pETC redox poising was investigated using cadmium (Cd) as an abiotic stressor to disrupt photosynthesis. Assessment of the photosynthetic responses of recombinant Anabaena strains, AnlexA+ (LexA-overexpressing) and AnpAM (vector control), under unstressed and Cd-stressed conditions using transmission electron microscopy (TEM) and chlorophyll a fluorescence, indicated that some pETC redox poising responses, including PSII photodamage, energy dissipation, PSI photoprotection, and NDHmediated cyclic electron flow were decreased in AnlexA+ under unstressed conditions. Disturbance in pETC redox poise during Cd stress observed in Anabaena was accentuated upon overexpression of LexA. The decreased photodamage of PSII and increased photoinhibition of PSI in AnlexA+ in the presence or absence of Cd stress, correlated well with the changes in pETC complexes observed in blue native (BN)-PAGE and the regulation of over 70 of the 90 pETC component genes by LexA demonstrated through transcript, electromobility shift assay (EMSA), and bioinformatics studies. In a nutshell, LexA has been identified as one of the regulators involved in the streamlining of pETC redox poising responses under normal growth and during abiotic stress through transcriptional regulation of some of the redox-controlled pETC component genes.
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| 8. |
- Srivastava, Vaibhav, et al.
(författare)
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Alternative splicing studies of the reactive oxygen species gene network in Populus reveal two isoforms of high-isoelectric-point superoxide dismutase
- 2009
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 0032-0889 .- 1532-2548. ; 149:4, s. 1848-1859
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Tidskriftsartikel (refereegranskat)abstract
- Recent evidence has shown that alternative splicing (AS) is widely involved in the regulation of gene expression, substantially extending the diversity of numerous proteins. In this study, a subset of expressed sequence tags representing members of the reactive oxygen species gene network was selected from the PopulusDB database to investigate AS mechanisms in Populus. Examples of all known types of AS were detected, but intron retention was the most common. Interestingly, the closest Arabidopsis (Arabidopsis thaliana) homologs of half of the AS genes identified in Populus are not reportedly alternatively spliced. Two genes encoding the protein of most interest in our study (high-isoelectric-point superoxide dismutase [hipI-SOD]) have been found in black cottonwood (Populus trichocarpa), designated PthipI-SODC1 and PthipI-SODC2. Analysis of the expressed sequence tag libraries has indicated the presence of two transcripts of PthipI-SODC1 (hipI-SODC1b and hipI-SODC1s). Alignment of these sequences with the PthipI-SODC1 gene showed that hipI-SODC1b was 69 bp longer than hipI-SODC1s due to an AS event involving the use of an alternative donor splice site in the sixth intron. Transcript analysis showed that the splice variant hipI-SODC1b was differentially expressed, being clearly expressed in cambial and xylem, but not phloem, regions. In addition, immunolocalization and mass spectrometric data confirmed the presence of hipI-SOD proteins in vascular tissue. The functionalities of the spliced gene products were assessed by expressing recombinant hipI-SOD proteins and in vitro SOD activity assays.
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| 9. |
- Srivastava, Vaibhav, et al.
(författare)
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Downregulation of high-isoelectric-point extracellular superoxide dismutase mediates alterations in the metabolism of reactive oxygen species and developmental disturbances in hybrid aspen
- 2007
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Ingår i: The Plant Journal. - : Blackwell Publishing. - 0960-7412 .- 1365-313X. ; 49:1, s. 135-148
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Tidskriftsartikel (refereegranskat)abstract
- Transgenic hybrid aspen (Populus tremula L. x P. tremuloides Michx.) plants expressing a high-isoelectric-point superoxide dismutase (hipI-SOD) gene in antisense orientation were generated to investigate its function. Immunolocalization studies showed the enzyme to be localized extracellularly, in the secondary cell wall of xylem vessels and phloem fibers. The antisense lines of hipI-SOD exhibited a distinct phenotype; growth rate was reduced, stems were thinner and leaves smaller than in wild-type (WT) plants. The abundance of hipI-SOD was reduced in the bark and xylem of plants from these antisense lines. The vascular tissue of transgenic lines became lignified earlier than in WT plants and also showed an increased accumulation of reactive oxygen species (ROS). Xylem fibers and vessels were shorter and thinner in the transgenic lines than in WT plants. The total phenolic content was enhanced in the antisense lines. Furthermore, microarray analysis indicated that several enzymes involved in cell signaling, lignin biosynthesis and stress responses were upregulated in apical vascular tissues of transgenic plants. The upregulation of selected genes involved in lignin biosynthesis was also verified by real-time PCR. The results suggest that, in the transgenic plants, a premature transition into maturation occurs and the process is discussed in terms of the effects of increased accumulation of ROS due to reduced expression of hipI-SOD during development and differentiation.
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| 10. |
- Yadav, Sandhya, et al.
(författare)
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Comparison and optimization of protein extraction and two-dimensional gel electrophoresis protocols for liverworts
- 2020
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Ingår i: BMC Research Notes. - : Springer Nature. - 1756-0500. ; 13:1
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Tidskriftsartikel (refereegranskat)abstract
- Objective Liverworts possess historical adaptive strategies for abiotic stresses because they were the first plants that shifted from water to land. Proteomics is a state-of-the-art technique that can capture snapshots of events occurring at the protein level in many organisms. Herein, we highlight the comparison and optimization of an effective protein extraction and precipitation protocol for two-dimensional gel electrophoresis (2-DE) of liverworts. Results We compared three different protein extraction methods, i.e.,1.5 M Tris-HCl (pH 8.8), 50 mM Tris-HCl (pH 7.5), and polyvinylpolypyrrolidone (PVPP) extraction, followed by three precipitation methods, i.e., 80% ethanol, 80% acetone, and 20% tricholoroacetic acid (TCA)-acetone, in a liverwort Dumortiera hirsuta. Among these methods, 50 mM Tris-HCl (pH 7.5) extraction, followed by 20% TCA-acetone precipitation, appeared to be more suitable for 2-DE. Furthermore, we performed modifications during protein washing, re-solubilization in rehydration buffer and isoelectric focusing (IEF). The modifications provided us better results in terms of protein yield, resolution, spot numbers, and intensities for 2-DE gels of D. hirsuta and other two liverworts, i.e., Marchantia paleacea and Plagiochasma appendiculatum. Furthermore, we randomly selected spots from the 2-DE gel of D. hirsuta and identified using mass spectrometry, which confirms the applicability of this protocol for liverworts proteomics.
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