| 1. |
- Hosseini, Sara, et al.
(författare)
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Comparative proteomic analysis of hyphae and germinating cysts of Phytophthora pisi and Phytophthora sojae.
- 2015
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Ingår i: Journal of Proteomics. - : Elsevier BV. - 1874-3919 .- 1876-7737. ; 117, s. 24-40
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Tidskriftsartikel (refereegranskat)abstract
- The recently described oomycete pathogen Phytophthora pisi causes root rot on pea and faba bean, while the closely related Phytophthora sojae is the causal agent of soybean root and stem rot. Differences in the pathogenicity factor repertoires that enable the two species to have distinct host specificity towards pea and soybean, were studied using tandem mass spectrometry in a global proteome study of hyphae and germinating cysts in P. pisi and P. sojae. In total 2775 proteins from P. pisi and 2891 proteins from P. sojae were identified. Fifty-eight orthologous proteins were more abundant in germinated cysts of both pathogens and thus identified as candidate proteins for the infective stage. Several of these proteins were associated with lipid transport and metabolism, and energy production. Twenty-three orthologous proteins were more abundant in hyphae of both pathogens and thus identified as candidate proteins for vegetative growth. Proteins uniquely present in germinating cysts of either P. pisi or P. sojae were considered as candidates for species-specific pathogenicity factors that may be involved in host specificity. Among these proteins were serine proteases, membrane transporters and a berberine-like protein. These results significantly expand the knowledge of the expressed proteome in P. pisi and P. sojae.
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| 2. |
- Abreha, Kibrom Berhe, et al.
(författare)
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Article leaf apoplast of field-grown potato analyzed by quantitative proteomics and activity-based protein profiling
- 2021
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Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 22:21
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Tidskriftsartikel (refereegranskat)abstract
- Multiple biotic and abiotic stresses challenge plants growing in agricultural fields. Most molecular studies have aimed to understand plant responses to challenges under controlled conditions. However, studies on field-grown plants are scarce, limiting application of the findings in agricultural conditions. In this study, we investigated the composition of apoplastic proteomes of potato cultivar Bintje grown under field conditions, i.e., two field sites in June–August across two years and fungicide treated and untreated, using quantitative proteomics, as well as its activity using activity-based protein profiling (ABPP). Samples were clustered and some proteins showed significant intensity and activity differences, based on their field site and sampling time (June–August), indicating differential regulation of certain proteins in response to environmental or developmental factors. Peroxidases, class II chitinases, pectinesterases, and osmotins were among the proteins more abundant later in the growing season (July–August) as compared to early in the season (June). We did not detect significant differences between fungicide Shirlan treated and untreated field samples in two growing seasons. Using ABPP, we showed differential activity of serine hydrolases and β-glycosidases under greenhouse and field conditions and across a growing season. Furthermore, the activity of serine hydrolases and β-glycosidases, including proteins related to biotic stress tolerance, decreased as the season progressed. The generated proteomics data would facilitate further studies aiming at understanding mechanisms of molecular plant physiology in agricultural fields and help applying effective strategies to mitigate biotic and abiotic stresses.
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| 3. |
- Ali, Ashfaq, et al.
(författare)
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Paranoid potato : phytophthora-resistant genotype shows constitutively activated defense
- 2012
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Ingår i: Plant Signaling and Behavior. - : Informa UK Limited. - 1559-2316 .- 1559-2324. ; 7:3, s. 400-408
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Tidskriftsartikel (refereegranskat)abstract
- Phytophthora is the most devastating pathogen of dicot plants. There is a need for resistance sources with different modes of action to counteract the fast evolution of this pathogen. In order to better understand mechanisms of defense against P. infestans, we analyzed several clones of potato. Two of the genotypes tested, Sarpo Mira and SW93-1015, exhibited strong resistance against P. infestans in field trials, whole plant assays and detached leaf assays. The resistant genotypes developed different sizes of hypersensitive response (HR)-related lesions. HR lesions in SW93-1015 were restricted to very small areas, whereas those in Sarpo Mira were similar to those in Solanum demissum, the main source of classical resistance genes. SW93-1015 can be characterized as a cpr (constitutive expressor of PR genes) genotype without spontaneous microscopic or macroscopic HR lesions. This is indicated by constitutive hydrogen peroxide (H₂O₂) production and PR1 (pathogenesis-related protein 1) secretion. SW93-1015 is one of the first plants identified as having classical protein-based induced defense expressed constitutively without any obvious metabolic costs or spontaneous cell death lesions.
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| 4. |
- Ali, Ashfaq, et al.
(författare)
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Quantitative proteomics and transcriptomics of potato in response to Phytophthora infestans in compatible and incompatible interactions
- 2014
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Ingår i: BMC Genomics. - : Springer Science and Business Media LLC. - 1471-2164. ; 15
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Tidskriftsartikel (refereegranskat)abstract
- Background: In order to get global molecular understanding of one of the most important crop diseases worldwide, we investigated compatible and incompatible interactions between Phytophthora infestans and potato (Solanum tuberosum). We used the two most field-resistant potato clones under Swedish growing conditions, which have the greatest known local diversity of P. infestans populations, and a reference compatible cultivar. Results: Quantitative label-free proteomics of 51 apoplastic secretome samples (PXD000435) in combination with genome-wide transcript analysis by 42 microarrays (E-MTAB-1515) were used to capture changes in protein abundance and gene expression at 6, 24 and 72 hours after inoculation with P. infestans. To aid mass spectrometry analysis we generated cultivar-specific RNA-seq data (E-MTAB-1712), which increased peptide identifications by 17%. Components induced only during incompatible interactions, which are candidates for hypersensitive response initiation, include a Kunitz-like protease inhibitor, transcription factors and an RCR3-like protein. More secreted proteins had lower abundance in the compatible interaction compared to the incompatible interactions. Based on this observation and because the well-characterized effector-target C14 protease follows this pattern, we suggest 40 putative effector targets. Conclusions: In summary, over 17000 transcripts and 1000 secreted proteins changed in abundance in at least one time point, illustrating the dynamics of plant responses to a hemibiotroph. Half of the differentially abundant proteins showed a corresponding change at the transcript level. Many putative hypersensitive and effector-target proteins were single representatives of large gene families.
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| 5. |
- Andersson, Karl-Magnus, et al.
(författare)
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Proteome of the nematode-trapping cells of the fungus Monacrosporium haptotylum.
- 2013
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Ingår i: Applied and Environmental Microbiology. - 0099-2240. ; 79:16, s. 4993-5004
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Tidskriftsartikel (refereegranskat)abstract
- Many nematophagous fungi use morphological structures called traps to capture nematodes by adhesion or mechanically. To better understand the cellular functions of adhesive traps, the trap cell proteome of the fungus Monacrosporium haptotylum was characterized. The trap of M. haptotylum consists of a unicellular structure called knob that develops at the apex of a hyphae. Proteins extracted from knobs and mycelia were analyzed using SDS-PAGE and LC-MS/MS. The peptide sequences were matched against predicted gene models from the recently sequenced M. haptotylum genome. In total, 336 proteins were identified, with 54 being expressed at significantly higher levels in the knobs than in the mycelia. The upregulated knob proteins included peptidases, small secreted proteins with unknown function and putative cell surface adhesins containing carbohydrate-binding domains including the WSC domain. Phylogenetic analysis showed that all upregulated WSC domain proteins belonged to a large, expanded cluster of paralogs in M. haptotylum. Several peptidases and homologs to experimentally verified proteins in other pathogenic fungi were also upregulated in the knob proteome. Complementary profiling of gene expression at the transcriptome level showed poor correlation between the upregulation of knob proteins and their corresponding transcripts. We propose that the traps of M. haptotylum contain many of the proteins needed in the early stages of infection, and that the trap cells can tightly control the translation and degradation of these proteins to minimize the cost of protein synthesis.
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| 6. |
- Antberg, Linn, et al.
(författare)
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Critical Comparison of Multidimensional Separation Methods for Increasing Protein Expression Coverage
- 2012
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Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 11:5, s. 2644-2652
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Tidskriftsartikel (refereegranskat)abstract
- We present a comparison of two-dimensional separation methods and how they affect the degree of coverage of protein expression in complex mixtures. We investigated the relative merits of various protein and peptide separations prior to acidic reversed-phase chromatography directly coupled to an ion trap mass spectrometer. The first dimensions investigated were density gradient organelle fractionation of cell extracts, 1D SDS-PAGE protein separation followed by digestion by trypsin or GluC proteases, strong cation exchange chromatography, and off-gel isoelectric focusing of tryptic peptides. The number of fractions from each first dimension and the total data accumulation RP-HPLC-MS/MS time was kept constant and the experiments were run in triplicate. We find that the most critical parameters are the data accumulation time, which defines the level of under-sampling and the avoidance of peptides from high expression level proteins eluting over the entire gradient.
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| 7. |
- Arrigoni, Giorgio, et al.
(författare)
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Chemical derivatization of phosphoserine and phosphothreonine containing peptides to increase sensitivity for MALDI-based analysis and for selectivity of MS/MS analysis
- 2006
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Ingår i: Proteomics. - : Wiley. - 1615-9861 .- 1615-9853. ; 6:3, s. 757-766
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Tidskriftsartikel (refereegranskat)abstract
- Protein phosphorylation is one of the most important and common ways of regulating protein function in cells. However, phosphopeptides are difficult to analyse, ionising poorly under standard MALDI conditions. Several methods have been developed to deal with the low sensitivity and specificity of phosphopeptide analysis. Here, we show an approach using a simple one-step beta-elimination/Michael addition reaction for the derivatization of phosphoserine and phosphothreonine. The substitution of the negatively charged phosphate group by a positively charged S-ethylpyridyl group greatly improves the ionisation of the modified peptides, especially in MALDI MS, increasing the sensitivity of the analysis. The modification allows the formation of a unique fragment ion at m/z 106 under mild collisional activation conditions, which can be used for parent (precursor) ion scanning in order to improve both the sensitivity and the selectivity of the analysis. The optimisation of the approach is described for a standard model peptide and protein and then applied to phosphorylation analysis in two biologically derived proteins purified from different experimental systems.
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| 8. |
- Ashraf, Muhammad Zubair, et al.
(författare)
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Comparative gut proteomics study revealing adaptive physiology of Eurasian spruce bark beetle, Ips typographus (Coleoptera: Scolytinae)
- 2023
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Ingår i: Frontiers in Plant Science. - 1664-462X. ; 14
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Tidskriftsartikel (refereegranskat)abstract
- The bark beetle, Ips typographus (L.), is a major pest of Norway spruce, Picea abies (L.), causing enormous economic losses globally. The adult stage of the I. typographus has a complex life cycle (callow and sclerotized); the callow beetles feed ferociously, whereas sclerotized male beetles are more aggressive and pioneers in establishing new colonies. We conducted a comparative proteomics study to understand male and female digestion and detoxification processes in callow and sclerotized beetles. Proteome profiling was performed using high-throughput liquid chromatography-mass spectrometry. A total of >3000 proteins were identified from the bark beetle gut, and among them, 539 were differentially abundant (fold change ±2, FDR <0.05) between callow and sclerotized beetles. The differentially abundant proteins (DAPs) mainly engage with binding, catalytic activity, anatomical activity, hydrolase activity, metabolic process, and carbohydrate metabolism, and hence may be crucial for growth, digestion, detoxification, and signalling. We validated selected DAPs with RT-qPCR. Gut enzymes such as NADPH-cytochrome P450 reductase (CYC), glutathione S-transferase (GST), and esterase (EST) play a crucial role in the I. typographus for detoxification and digesting of host allelochemicals. We conducted enzyme activity assays with them and observed a positive correlation of CYC and GST activities with the proteomic results, whereas EST activity was not fully correlated. Furthermore, our investigation revealed that callow beetles had an upregulation of proteins associated with juvenile hormone (JH) biosynthesis and chitin metabolism, whereas sclerotized beetles exhibited an upregulation of proteins linked to fatty acid metabolism and the TCA cycle. These distinctive patterns of protein regulation in metabolic and functional processes are specific to each developmental stage, underscoring the adaptive responses of I. typographicus in overcoming conifer defences and facilitating their survival. Taken together, it is the first gut proteomic study comparing males and females of callow and sclerotized I. typographus, shedding light on the adaptive ecology at the molecular level. Furthermore, the information about bark beetle handling of nutritionally limiting and defence-rich spruce phloem diet can be utilized to formulate RNAi-mediated beetle management.
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| 9. |
- Bendz, Maria, et al.
(författare)
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Quantification of Membrane Proteins Using Nonspecific Protease Digestions
- 2009
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Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 8:12, s. 5666-5673
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Tidskriftsartikel (refereegranskat)abstract
- We present a mass spectrometry-based method for the identification and quantification of membrane proteins using the low-specificity protease Proteinase K, at very high pH, to digest proteins isolated by a modified SDS-PAGE protocol. The resulting peptides are modified with a fragmentation-directing isotope labeled tag. We apply the method to quantify differences in membrane protein expression of Bacillus subtilis grown in the presence or absence of glucose.
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| 10. |
- Bengtsson, Therese, et al.
(författare)
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Proteomics and transcriptomics of the BABA-induced resistance response in potato using a novel functional annotation approach
- 2014
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Ingår i: BMC Genomics. - : Springer Science and Business Media LLC. - 1471-2164. ; 15
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Tidskriftsartikel (refereegranskat)abstract
- Background: Induced resistance (IR) can be part of a sustainable plant protection strategy against important plant diseases. beta-aminobutyric acid (BABA) can induce resistance in a wide range of plants against several types of pathogens, including potato infected with Phytophthora infestans. However, the molecular mechanisms behind this are unclear and seem to be dependent on the system studied. To elucidate the defence responses activated by BABA in potato, a genome-wide transcript microarray analysis in combination with label-free quantitative proteomics analysis of the apoplast secretome were performed two days after treatment of the leaf canopy with BABA at two concentrations, 1 and 10 mM. Results: Over 5000 transcripts were differentially expressed and over 90 secretome proteins changed in abundance indicating a massive activation of defence mechanisms with 10 mM BABA, the concentration effective against late blight disease. To aid analysis, we present a more comprehensive functional annotation of the microarray probes and gene models by retrieving information from orthologous gene families across 26 sequenced plant genomes. The new annotation provided GO terms to 8616 previously un-annotated probes. Conclusions: BABA at 10 mM affected several processes related to plant hormones and amino acid metabolism. A major accumulation of PR proteins was also evident, and in the mevalonate pathway, genes involved in sterol biosynthesis were down-regulated, whereas several enzymes involved in the sesquiterpene phytoalexin biosynthesis were up-regulated. Interestingly, abscisic acid (ABA) responsive genes were not as clearly regulated by BABA in potato as previously reported in Arabidopsis. Together these findings provide candidates and markers for improved resistance in potato, one of the most important crops in the world.
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