Protein Spots Research Articles

Proteomic Analysis Reveals Differential Protein Expression Induced by Inhibition of Prolyl Oligopeptidase in Filarial Parasites.

Prolyl oligopeptidase (POP) plays a crucial role in the processing and degradation of neuropeptides and regulates inositol trisphosphate (IP3) signaling in mammals. We have reported that POP inhibition leads to IP3-mediated calcium efflux leading to mitochondrial-mediated apoptosis in the filarial parasite Setaria cervi. This study further elucidates the effect of altered calcium homeostasis on the proteome of filarial parasites. Adult parasites were treated with POP's specific inhibitor, Z-Pro-prolinal (ZPP), for 7h. Cytosolic and mitochondrial proteome was analyzed using 2D gel electrophoresis coupled with MALDI-MS/MS. Phosphoproteins were also analyzed in the cytosolic fraction of the parasites. The phosphoprotein analysis revealed 7, and 9 spots in the cytosolic fraction of control and ZPP-treated parasites, respectively. The two identified protein spots in the treated set were found to be involved in G protein signaling. In cytosolic fraction, 109 and 112 protein spots were observed in control and treated parasites, respectively. Of these, 56 upregulated and 32 downregulated protein spots were observed in the treated set. On the other hand, 50 and 47 protein spots were detected in the mitochondrial fraction of control and treated parasites, respectively. Of these spots, 18 upregulated and 12 down-regulated protein spots were found in treated parasites. In silico analysis showed that the identified proteins were involved in energy metabolism, calcium signaling, stress response, and cytoskeleton organization. These findings correlate with our previous results suggesting the important regulatory role of POP in signaling and different metabolic pathways of filarial parasites.

Effect of hazelnut skin by-product supplementation in lambs' diets: Implications on plasma and muscle proteomes and first insights on the underlying mechanisms

This study aimed to evaluate the effect of hazelnut skin by-product supplementation on lamb meat quality characteristics and plasma and muscle proteomes. Twenty-two Valle del Belice male lambs were divided into two experimental groups: control (C), fed a maize-barley diet and hazelnut (H), fed hazelnut skin by-product as maize partial replacer in the concentrate diet. The meat of lambs fed hazelnut skin showed greater values of lightness, redness, yellowness, and chroma color parameters together with the highest myofibril fragmentation index. Two-dimensional electrophoresis and mass spectrometry applied on plasma proteome identified 20 protein spots corresponding to 18 unique gene names to be differently expressed due to hazelnut skin by-product substitution. For the early post-mortem muscle, 23 protein spots (42 unique gene names) were significantly up-regulated due to hazelnut skin by-product supplementation. Four proteins these being APOA1, PHB, ACTG1 and ALB, were found to be common to the two proteomes suggesting that these proteins could be candidate biomarkers to monitor in vita and post-mortem lamb meat quality traits. This study evidenced the main mechanisms involved in the supplementation of hazelnut skin by-product in lambs' diet and confirmed the possibility of using plasma proteome as a non-invasive way to predict lamb meat quality. SignificanceMaximizing the use of agro-industrial by-products as replacers of traditional feedstuff for improving animal products is one of the important challenges to preserving natural resources and guaranteeing environmental sustainability. Hazelnut (Corylus avellana L.) skin, obtained as a results of hazelnut roasting, represents a valuable by-products due to its high content in unsaturated fatty acids, tannins, and vitamins. Thus, including hazelnut skin by-product in small ruminant nutrition could reduce the costs of animal feedings for farmers as well as improve meat nutritional and sensorial characteristics. Additionally, monitoring the meat quality characteristics with fast, accurate, and non-invasive tools to find, before slaughter, animals with desired quality characteristics is of growing interest in the last years. In this regard, the objectives of this study were to assess i) the effect of hazelnut skin supplementation on lamb meat quality characteristics and plasma and muscle proteomes, and ii) whether analyzing plasma proteome by using a gel-based proteomic approach could effectively offer a more readily available option for determining lamb meat quality. Taken together, the proteomic approach applied to plasma and muscle proteomes, allowed us to reveal the pathways and the potential candidate plasma biomarkers to predict lamb meat production in the pre-slaughter phase.

Comparative Appraisal of Leaf Proteomic and Mass Spectrometry Analyses During FusariumWilt Infection in Resistance and Susceptible Genotypes of Castor (Ricinus communis L.).

The resistant and susceptible genotypes of castor were utilized for leaf proteomic study during Fusarium wilt infection. The histopathological study was observed under SEM and it confirmed that the infection of Fusarium oxysporum f. sp. ricini was higher in the root of susceptible JI-35, while incompatible interaction is observed in resistant SKI-215 genotype. The acidic and neutral proteins were maximally up-expressed with 2 to 171 kDa in treated resistant and 2 to 150 kDa in treated susceptible interactions. In resistant genotype, the leaf proteins were recognized with 3.0- and 5.8-fold higher at infection stage and post-infection stage, respectively, as compared to susceptible genotype. The highly up expressions of leaf acidic (4.76 pI) and basic (8.77 pI) proteins were found with 224.94- and 61.68-fold change, respectively during the post-infection stage in treated resistance compared to its control. The protein spots at 4.76 pI and 8.77 pI were characterized with nanoLC-MS Triple TOF and were recognized as signalling molecules small GTP binding protein (23 kDa) and actin (8 kDa), respectively, on the basis of mass spectrometry and peptide sequences. However, basic and neutral proteins were up regulated as 30.11- and 20.30-fold changes in treated susceptible compared to its control. These proteins were identified as HSP90 (10 kDa) and LEA (27 kDa) proteins. The 148 kDa protein is recognized as histidine kinase in incompatible resistant interaction compared to compatible susceptible (serine threonine protein kinase, 65 kDa) as common acidic protein at 3.80 pI during infection stage. Some acidic proteins were maximally up-regulated in the leaf of resistant castor genotype and played a significant role in defense response.

Analysis of virulence proteins in pathogenic Acinetobacter baumannii to provide early warning of zoonotic risk

Acinetobacter baumannii is a ubiquitous opportunistic pathogen usually with low virulence. In recent years, reports of increased pathogenicity of A. baumannii in livestock due to the migratory behaviour of wildlife have attracted public health attention. Our previous study reported that an A. baumannii strain isolated from dead chicks, CCGGD201101, showed enhanced pathogenicity, but the mechanism for increased virulence is not understood. Here, to screen potential virulence factors, the proteomes of the isolated strain CCGGD201101 and the standard strain ATCC19606 of A. baumannii were compared, and the possible virulence-enhancing mechanisms were further analysed. The 50 % lethal dose (LD50) values of CCGGD201101 and standard strain ATCC19606 in ICR mice were determined to verify their bacterial toxicity. 2D fluorescence difference gel electrophoresis (2D-DIGE) combined with matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF/TOF-MS) and quantitative real-time PCR (RTqPCR) were applied to screen and identify differentially expressed proteins or genes that may be related to virulence enhancement. Bioinformatics analyses based on proteinprotein interaction (PPI) networks were used to explore the function of potential virulence proteins. The pathogenicity of potential virulence factors was assessed by phylogenetic analyses and an animal infection model. The results showed that the LD50 of CCGGD201101 for mice was 1.186 × 106 CFU/mL, and the virulence was increased by 180.5-fold compared to ATCC19606. Forty-seven protein spots were significantly upregulated for the A. baumannii CCGGD201101 strain (fold change ≥1.5, p < 0.05). In total, 14 upregulated proteins were identified using proteomic analysis, and the mRNA expression levels of these proteins were nearly identical, with few exceptions. According to the PPI network and phylogenetic analyses, the I78 family peptidase inhibitor, 3-oxoacyl-ACP reductase FabG, and glycine zipper were screened as being closely related to the pathogenicity of bacteria. Furthermore, the I78 overexpression strains exhibited higher lethality in mouse infection models, which indicated that the I78 family peptidase inhibitor was a potential new virulence factor to enhance the pathogenicity of the A. baumannii CCGGD201101 strain. The present study helped us to better understand the mechanisms of virulence enhancement and provided a scientific basis for establishing an early warning system for enhanced virulence of A. baumannii from animals.

The HSP90 inhibitor 17-AAG induced calcium-mediated apoptosis in filarial parasites.

The available antifilarial medications are effective only against the larval stage of the filarial parasite. As a result, there is a pressing need for an adulticidal drug. The development of drugs requires the identification of molecular targets that are critical for parasite life. In this study, we observed the effect of 17-N-allyl-17-demethoxygeldanamycinon the survival of adult filarial parasites. The 17-N-allyl-17-demethoxygeldanamycin (17-AAG) is a derivative of geldanamycin (GA), which is an inhibitor of heat shock protein(HSP)90. It is less toxic as compared to geldanamycin. The motility and viability of the adult filarial parasite Setaria cervi were decreased on exposure to 17-AAG at 2.5 and 5.0 μM/ml concentrations. The 17-AAG treated parasites showed induction of oxidative stress as evidenced by decreased activity of various antioxidant enzymes like glutathione s-transferase, glutathione reductase, thioredoxin reductase, and an increase in ROS production in comparison to control. Oxidative stress may lead to altered calcium homeostasis. Indeed, in 17-AAG treated worms, there was a rise in calcium in the cytosol and mitochondria, as well as a decrease in the ER. We also observed enhanced activity of phospholipase C in the treated parasite, suggesting the opening of calcium channels located on the ER membrane. ER stress is marked by a reduced level of protein disulfide isomerase. Further, 17-AAG treated worms showed an increase in apoptotic marker enzyme activities like calpain, cyt-c, and caspase-3. The 2D-gel electrophoresis technique showed 142 protein spots in the control and 112 spots in the 17-AAG treated parasite. Thus, 17-AAG induced oxidative stress, and altered calcium, and proteostasis of parasites, which led to apoptosis.

Analysis of Membrane Proteins of Streptomycin-Resistant Mycobacterium tuberculosis Isolates

Background: Drug-resistant tuberculosis remains a health security threat and resistance to second-line drugs limits the options for treatment. Consequently, there is an utmost need for identifying and characterizing new biomarkers/drug targets of prime importance. Membrane proteins have an anticipated role in biological processes and could qualify as biomarkers/drug targets. Streptomycin (SM) is recommended as a second-line treatment regimen only when amikacin resistance has been confirmed. As extensively drug-resistant (XDR) isolates are frequently cross-resistant to second-line injectable drugs, an untapped potential for the continued use of SM has been suggested. Objective: The study aimed to analyze the membrane proteins overexpressed in SM resistant isolates of Mycobacterium tuberculosis using proteomics approaches. Methods: Membrane proteins were extracted employing sonication and ultracentrifugation. Twodimensional gel electrophoresis (2DGE) of membrane proteins was performed and identification of proteins was done by liquid chromatography-mass spectrometry (LCMS) and bioinformatics tools. Results: On analyzing the two-dimensional (2D) gels, five protein spots were found overexpressed in the membrane of SM resistant isolates. Docking analysis revealed that SM might bind to the conserved domain of overexpressed proteins and Group-based prediction system-prokaryotic ubiquitinlike protein (GPS-PUP) predicted potential pupylation sites within them. Conclusion: These proteins might be of diagnostic importance for detecting the cases early and for exploring effective control strategies against drug-resistant tuberculosis, particularly SM.

Analysis of Expression of Diabetic Nephropathy-Related Protein and Stem Cell Tissue Repair Under Nano Membrane Concentration Technology

This study aimed to study protein expression in diabetic nephropathy (DN) rats and stem cell tissue repair based on nanomembrane concentration technology. Based on the polymer porous self-assembled nanomembrane technology, the content of total protein and albumin in the serum of rats in the control and experimental groups were measured. The obtained images were adopted to analyze the expression of cytoplasmic proteins and membrane proteins, and then the mechanism of stem cell tissue repair function was studied. The results showed that at 56 weeks of age, in contrast to control group, the total protein content in the serum of the experimental group evidently decreased. At 36 weeks of age, the cytoplasmic protein samples of diabetic rats in the experimental group were subjected to three two-dimensional protein electrophoresis. It was found that there were about 701 spots in each gel, and the matching rate was about 87.5%. In contrast to control group, 16 cytoplasmic proteins and 23 membrane proteins of the experimental group changed. The results of the differentially expressed protein analysis indicated that the change trends of protein spots. Based on polymer porous self-assembled nano-film technology, the expression of differential proteins in DN rats was analyzed, which would supplement new research assistance to the mechanism of DN.

Preparing mare milk sample for proteomic analysis using two-dimensional electrophoresis

There is no universal method to prepare physiological fluids for 2-DE proteomic analysis. Furthermore, interspecies differences in milk composition require the formulation of a species-specific sample preparation procedure. The study was carried out on mare's milk which was prepared for 2-DE in four different methods: the first sample (M1) was defatted, sample M2 was defatted and after casein precipitation, sample M3 was sample M2 after reduction of high molecular proteins and sample M4 was desalted sample M3. The milk samples prepared in different methods were separated by 1-DE and 2-DE. The obtained gels were analysed qualitatively and quantitatively. Furthermore, selected protein spots were identified by MALDI-TOF MS. Analysis of 1-DE and 2-DE gel images indicated that the optimal procedure for preparing mare milk samples for 2-DE and identification of proteins by MS is a method based on defatting and precipitation of caseins. The preparation of mare's milk samples by defatting and then precipitation of caseins is enough to obtain an optimal 2-DE separation for identification of proteins of this body fluid. The method of caseins precipitation should be improved in order to reduce the proportion of these high-abundance protein in milk samples which could increase in the identification of low-abundance proteins in mare's milk.

Proteomic Analysis of the Inflorescence Stem Mechanical Strength Difference in Herbaceous Peonies (Paeonia lactiflora Pall.).

The herbaceous peony (Paeonia lactiflora Pall.) is a traditional rare flower in China, and production of its cut flowers has developed gradually in many places of the world. However, the inflorescence stems of some P. lactiflora cultivars have such low mechanical strength that the cut flower production was severely restricted. To better understand the causes of this problem from a protein expression level, two P. lactiflora cultivars with different inflorescence stem mechanical strengths were analyzed by two-dimensional electrophoresis and MALDI-TOF/MS. More than 1700 clear protein spots were detected, 53 of which varied significantly. Moreover, 23 of the differentially expressed proteins were identified and confirmed and are involved in various biological processes such as metabolism, protein biosynthesis and transport, signal transduction, and defensive response. Especially, cinnamyl alcohol dehydrogenase (CAD) and xyloglucan endotransglucosylase/hydrolase (XTH) were strongly connected to the inflorescence stem mechanical strength in P. lactiflora.

Identification of Allergenic Proteins in Velvet Mesquite (Prosopis velutina) Pollen: An Immunoproteomics Approach.

Velvet mesquite (Prosopis velutina) is a native legume of the southwestern United States and northwestern Mexico, contributing significantly to the desert ecosystem and playing key ecological roles. It is also an important cause of allergic respiratory disease widely distributed in the Sonoran, Chihuahuan, and Mojave Deserts. However, no allergens from velvet mesquite pollen have been identified to date. Pollen proteins were extracted and analyzed by one- and two-dimensional electrophoresis and immunoblotting using a pool of 11 sera from mesquite-sensitive patients as the primary antibody. IgE-recognized protein spots were identified by mass spectrometry and bioinformatics analysis. Twenty-four unique proteins, including proteins well known as pollen, food, airway, or contact allergens and four proteins not previously reported as pollen allergens, were identified. This is the first report on allergenic proteins in velvet mesquite pollen. These findings will contribute to the development of specific diagnosis and treatment of mesquite pollen allergy.

Genetic structure and proteomic analysis associated in potato to Rhizoctonia solani AG-3PT-stem canker and black scurf

Rhizoctonia solani AG-3PT is a necrotrophic plant pathogenic fungus affecting potato production globally. A total of 20 isolates of R. solani AG-3PT were subjected to pathogenicity tests, genetic diversity and proteomic analyses. The genetic diversity of R. solani AG-3PT isolates with different disease virulence was analyzed using 11 simple sequence repeats (SSR) primer pairs. Cluster analysis grouped the isolates into five main categories, indicating a high genetic diversity and recombination among the isolates, with no correlation between the groups and geographical origin. The protein pattern of 20 isolates of R. solani AG-3PT with different disease severity were analyzed by SDS-PAGE. Protein profiles were different that maybe associated with different levels of their aggressiveness. Based on the pathogenicity test results, isolates Rs-13 and Rs-4 were classified as virulent and least virulent, respectively. Here, we used proteomics strategy to compare mycelial proteins of virulent and least virulent isolates in order to further characterize them at the molecular level. The 2-DE results showed a differential expression of 27 protein spots, of which 11 well-focused protein spots were subjected to LC-MS-MS. The functions of the identified proteins indicated that they play roles in the fungal development, sporulation, carbohydrate metabolic process, lipid metabolic process, protein modification, and nucleotide metabolic process. The differences in pathogenicity between Rs-13 and Rs-4 isolate can be explained by differences in the expression levels of proteins involved in these pathways. The results of this research can enrich our knowledge about the virulence strategies of R. solani AG-3PT and provide useful information for the development of future fungicides. A combination of genetic diversity assessments of pathogen and understanding the biology and virulence mechanisms of pathogen, can help us in development of innovative and sustainable approaches to slow down the emergence of pathogen.

Potato tuber skin proteome is enriched with defensive components to prevent the further infection of common scab into tuber flesh

Potato common scab is a soil-borne disease caused mainly by Streptomyces scabies that seriously affects tuber quality and production. To understand the potential molecular mechanisms of potato tuber responded to common scab, a comparative proteomics approach was applied to analyze the proteome alteration of tuber skin and flesh after infected with common scab by two-dimensional gel electrophoresis (2-DE). Quantitative image analysis showed that the abundance of 36 and 26 protein spots in tuber skin and flesh of scab-infected potato were significantly altered (p < 0.05) more than 1.5-fold, respectively. All these differentially abundant proteins were successfully identified by MALDI-TOF/TOF MS, which were mainly involved in defense, bioenergy and metabolism, storage, chaperones, redox homeostasis, miscellaneous and other unknown functions. Compared with tuber flesh, more defense-related proteins were up-regulated in tuber skin, and several cell wall reorganization and lignification-related enzymes were also up-regulated in tuber skin. It appeared that the tuber skin as a barrier might be enriched with components of defense response to prevent the further infection of pathogen into tuber flesh. Several chaperones were up-regulated in tuber skin and flesh, suggesting a key role in reconstructing normal protein conformation and cell homeostasis in tubers. Conversely, common scab causes the down-regulation of many patatin proteins, which might inhibit tuber swelling. This study would provide some new insight into the potential response mechanisms of tuber infected with common scab, and help in developing strategies to improve resistance to common scab in potato breeding.

Hydrogen peroxide initiates oxidative stress and proteomic alterations in meningothelial cells

Meningothelial cells (MECs) are fundamental cells of the sheaths covering the brain and optic nerve, where they build a brain/optic nerve-cerebral spinal fluid (CSF) barrier that prevents the free flow of CSF from the subarachnoid space, but their exact roles and underlying mechanisms remain unclear. Our attempt here was to investigate the influence elicited by hydrogen peroxide (H2O2) on functional changes of MECs. Our study showed that cell viability of MECs was inhibited after cells were exposed to oxidative agents. Cells subjected to H2O2 at the concentration of 150 µM for 24 h and 48 h exhibited an elevation of reactive oxygen species (ROS) activity, decrease of total antioxidant capacity (T-AOC) level and reduced mitochondrial membrane potential (ΔΨm) compared with control cells. 95 protein spots with more than twofold difference were detected in two dimensional electrophoresis (2DE) gels through proteomics assay following H2O2 exposure for 48 h, 10 proteins were identified through TOF/MS analysis. Among the proteomic changes explored, 8 proteins related to energy metabolism, mitochondrial function, structural regulation, and cell cycle control were downregulated. Our study provides key insights that enhance our understanding of the role of MECs in the pathology of brain and optic nerve disorders.

Cofilin-1 and profilin-1 expression in lung microvascular endothelial cells exposed to titanium dioxide nanoparticles

Air pollutants exacerbate chronic airway diseases, such as asthma and chronic obstructive pulmonary disease (COPD). However, the underlying mechanisms are yet to be determined. While a number of studies have reported adverse effects of nanoparticles on humans, little is known about their effects on the respiratory system. To examine the protein expression in human lung microvascular endothelial cells (HMVEC-L) exposed to titanium dioxide (TiO2) nanoparticles, a common air pollutant. A proteomics approach using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF MS) was used to determine the differences in protein expression at 8 h and 24 h, following the treatment of HMVEC-L with 20-μM or 40-μM TiO2 nanoparticles. Human lung microvascular endothelial cells treated with 20-μM TiO2 nanoparticles showed alterations of 7 protein spots, including molecules related to calcium regulation, transport, cytoskeleton, and muscle contraction. The treatment of HMVEC-L with 40-μM TiO2 nanoparticles resulted in alterations of 4 protein spots, with molecular functions related to the cytoskeleton, myosin regulation, actin modulation, as well as guanosine diphosphate (GDP) and guanosine triphosphate (GTP) regulation. To validate these results, immunohistochemical staining and western blotting analyses were performed on lung tissues collected from mice exposed to TiO2 nanoparticles. Cofilin-1 and profilin-1 were expressed in the endothelium, epithelium and inflammatory cells, and decreased in lung tissues of TiO2 nanoparticle-exposed mice compared to sham-treated controls. These results suggest that some of the differentially expressed proteins may play important roles in airway diseases caused by TiO2 nanoparticle exposure.

Mercury metalloproteomic profile in muscle tissue of Arapaima gigas from the Brazilian Amazon.

Metalloproteomics is an innovative methodology for identifying of protein-associated mercury. Thus, we analyzed the muscle proteome of Arapaima gigas (pirarucu), collected in the Madeira River of the Brazilian Amazon, to identify protein-associated mercury, with the aim of identifying possible mercury biomarkers in fish muscle tissue. After obtaining the protein pellet, we conducted two-dimensional electrophoresis (2D PAGE) to fractionate the muscle proteome. Total mercury in muscle tissue and protein pellets and mapping of mercury content in protein spots of the 2D PAGE gels was determined using graphite furnace atomic absorption spectrometry (GFAAS). The protein-associated mercury identification was performed using liquid chromatography coupled with sequence mass spectrometry (LC‒MS/MS). Total mercury determinations by GFAAS indicated concentrations on the order of 153 ± 1.90mgkg-1 and 142 ± 1.50mgkg-1 (total precipitation of protein fraction) and 139 ± 1.45mgkg-1 (fractional precipitation of protein fraction) in muscle tissue and protein pellets, respectively. Mercury concentrations in the range of 48 ± 0.90 to 165 ± 3.00mgkg-1 were found in twelve protein spots. Among the 2D PAGE protein spots, eleven Hg-binding proteins were identified using LC‒MS/MS, which showed characteristics of mercury exposure biomarkers for important metabolic functions, such as five parvalbumin isoforms, triosephosphate isomerase, cofilin 2 (muscle), and fructose-bisphosphate aldolases.

Shrimp pmo-miR-750 regulates the expression of sarcoplasmic calcium-binding protein facilitating virus infection in Penaeus monodon

MicroRNAs (miRNAs) regulate gene expression post-transcriptionally and play crucial roles in antiviral responses. Penaeus monodon miR-750 (pmo-miR-750) was found to be strongly up-regulated in the late phase of white spot syndrome virus (WSSV) infection, but its function remains uncharacterized. Herein, the targets that were translationally down-regulated in the shrimp stomach following a pmo-miR-750 mimic injection were identified using two-dimensional gel electrophoresis. Sarcoplasmic calcium-binding protein (Scp) and actin1 (Act1) were revealed to be down-regulated protein spots. The genuine binding of pmo-miR-750 mimic to Scp but not Act1 mRNA was validated in vitro. In addition, a negative correlation between the Scp transcript and pmo-miR-750 expression level in WSSV-infected P. monodon stomach implies that pmo-miR-750 regulates Scp expression in vivo. When injected into WSSV-infected shrimp, the pmo-miR-750 mimic suppressed Scp expression but significantly increased the WSSV copy number. Consistent with the miRNA mimic-mediated Scp suppression, the loss of function assay of Scp in WSSV-challenged shrimp by RNA interference revealed a decreased survival rate with a dramatic increase in viral copy number. Besides that, apoptosis was activated in the hemocytes of the Scp knockdown shrimp upon WSSV infection. Collectively, our findings reveal that up-regulated pmo-miR-750 suppresses Scp expression at both the transcript and protein levels in the late stage of WSSV infection, which contributes to modulating apoptosis and eventually enabling viral propagation.

Identification and characterization of salt-responsive proteins in mango (Mangifera indica L.)

Increasing salinity is a cause of concern for meeting UN sustainable development goals and needs urgent mitigation strategies. The widespread use of salinity tolerant rootstocks in salt-sensitive fruits provides long-term solutions to the harsh effects of soil salinization. Polyembryonic mango cultivar 13-1, a salinity-tolerant variety from Israel, was used to unravel initial salt adaptive mechanism by imposing a salinity screen at 200mM NaCl. Differentially accumulated proteins were separated through 2-D electrophoresis (pH gradient 4.0-7.0), and identified through properties of pI and molecular weight by annotation against Citrus isoelectric focusing database. Protein spots (309) were detected on Coomassie-stained gels and about 22 spots were found differentially expressed in control and stress. Overproduction of stress-related proteins like polygalacturonase (97 kDa/4.9PI) and alcohol dehydrogenase (38kDa/5.6PI) is linked to enhanced cell wall integrity, transpiration rate regulation and ionic maintenance in adaptability mechanism. The upregulated phenylpropanoid pathway proteins p-coumaroyl ester, Flavanone3-hydroxylase-2 and UDP-glycosyl transferase are also involved in stress alleviation through flavonoid accumulation. Glutathione S-transferase was also identified with 2.21-fold accumulation in plants exposed to salinity stress, thereby elucidating its role in oxidative stress mitigation. Cell wall and cytoskeleton metabolism-related proteins were also found to be associated with salinity adaptation in mango cv 13-1. Differential accumulation of proteins implicated in signal transduction pathway, transcription regulation and hormone signaling were also identified. Thus, role of differentially expressed proteins under initial salinity stress conditions provide new insights molecular adjustment mechanisms orchestrated by mango rootstock variety by hormone signaling, osmotic arrangements, cytoskeleton modifications, phenol accumulation and transcription regulation.

Leaf proteome reveals the alterations in photosynthesis and defense-related proteins between potato tetraploid cultivars and diploid wild species

Potato (Solanum tuberosum L.) as the important food crop worldwide has abundant morphological and genetic diversity. To understand the underlying molecular mechanisms determining phenotypic differences in wild species and cultivated potato, a comparative proteomics approach was applied to analyze leaf proteome alteration among three tetraploid cultivars and three diploid wild species using two-dimensional gel electrophoresis (2-DE). Quantitative image analysis showed a total of 47 protein spots with significantly altered abundance (>3-fold, P < 0.05), and 45 differentially abundant proteins were identified by MALDI-TOF/TOF MS. These proteins exhibited both the qualitative and quantitative changes. Most of them were involved in photosynthesis, cell defense and rescue, protein biosynthesis, which might exhibit the main differences between tetraploid cultivars and diploid wild species. The photosynthesis and protein biosynthesis-related proteins were up-regulated or only present in tetraploid cultivars, suggesting the higher photosynthetic efficiency and more newly synthesized peptides. It might contribute to some superior traits of tetraploid cultivars, such as larger leaf size, greater growth vigor, better tuber yield and quality. However, some cell defense and rescue-related proteins, especially the pathogenesis-related proteins and antioxidant enzymes, were up-regulated or only present in diploid wild species. It might be responsible for stronger resistance to diseases and pests or tolerance to environmental stresses in diploid wild species. This study would provide valuable information for the underlying molecular mechanisms of potato genetic diversity, and help in developing strategies for the utilization of wild species for potato improvement.

First insights about the underlying mechanisms of Martina Franca donkey meat tenderization during aging: A proteomic approach

This study aimed to evaluate the effect of different aging times (1, 6 and 14 days) on the tenderization rate and protein changes in ten Martina Franca donkey striploins using a proteomic approach. During aging, a progressive decrease in share force, hardness, gumminess, and chewiness together with an increase in myofibril fragmentation index were observed. Proteolysis monitored by immunoblotting revealed a progressive degradation of desmin and fast troponin-T over time and an increase of their degradation products up to 14 days aging. Proteomics revealed by means of two-dimensional electrophoresis 37 protein spots corresponding to 15 proteins to change significantly by increasing aging time. These proteins belong to three pathways, these being “muscle contraction, structure, and associated proteins” (9 proteins); ii) “energy metabolism” (5 proteins); and iii) “chaperone” (1 protein). This study is the first to highlight the possible role of interconnected pathways in driving the final quality of donkey meat and predicting its texture.

Identification of Candidate Genes Associated with Trichothecene Biosynthesis in Fusarium graminearum Species Complex Combined with Transcriptomic and Proteomic Analysis.

The Fusarium graminearum species complex is the main causal agent of wheat head blight worldwide. Trichothecenes produced by the pathogen in infected grains have important food safety implications. Previously reported studies on trichothecene production have all focused on the conditions conducive to mycotoxin production, while the molecular mechanisms of trichothecene biosynthesis in Fusarium strains under normal or non-inducing conditions are still unclear. Here, a global analysis of the fungal gene expression of three strains using the Affymetrix Fusarium GeneChip under non-inducing conditions is reported. Differentially expressed genes were identified among strains with different trichothecene-production ability, and some novel genes associated with trichothecene biosynthesis were found by bioinformatics analysis. To verify the transcriptome results, proteomic analyses of the three strains were conducted under the same culture conditions. In total, 69 unique fungal proteins were identified in 77 protein spots. Combined with transcriptome and proteome analysis, 27 novel genes were predicted to be associated with trichothecene mycotoxin production. A protein, encoded by FGSG_01403, was found to be associated with trichothecene production via proteome analysis. Gene knock-out mutations of FGSG_01403 resulted in mutants with increased production of trichothecenes. Future functional analysis of the candidate genes identified in this study may reveal new insights into the negative regulation of trichothecene production in the Fusarium graminearum species complex.