Comparative proteomic analysis of the plant-virus interaction in resistant and susceptible ecotypes of maize infected with sugarcane mosaic virus.

Sugarcane mosaic virus (SCMV) is an important viral pathogen and has caused serious losses in grain and forage yield. To identify candidate SCMV resistance proteins and to explore the molecular mechanisms involved in the plant-SCMV interaction, we conducted proteomic analyses of leaf samples from resistant and susceptible ecotypes of maize infected with SCMV. Proteins were analyzed by quantitative two-dimensional differential gel electrophoresis (2D-DIGE), and 93 protein spots showed statistically significant differences after virus inoculation. Functional categorization showed that SCMV-responsive proteins were mainly involved in energy and metabolism, stress and defense responses, photosynthesis, and carbon fixation. The majority of the identified proteins were located in chloroplast and cytoplasm based on bioinformatic analysis. Among these identified proteins, 17 have not been identified previously as virus-responsive proteins, and 7 were new and did not have assigned functions. Western blotting analyses confirmed the expression patterns of proteins of specific interest, and the genes encoding these proteins were further analyzed by real-time PCR. The results of this study showed overlapping and specific proteomic responses to SCMV infection between resistant and susceptible maize ecotypes. This study provides further insight into the molecular events during compatible and incompatible interactions between viruses and host plants. BIOLOGICAL SIGNIFICANCE: Sugarcane mosaic virus (SCMV) is an important viral pathogen and has caused serious losses in grain and forage yield. However, little is known about host-SCMV interactions from the proteome perspective. This study analyzed proteomic changes in resistant and susceptible plants that are infected with SCMV using DIGE based proteomics. We identified 17 proteins that have not been identified previously as virus-responsive proteins, and 7 new proteins without assigned functions. These proteins are interesting candidates for future research, as they may be associated with new biological functions and play important roles in plant-virus interactions. Real-time RT-PCR analysis of genes encoding several proteins of interest provided indication on whether the changes in protein abundance were regulated at the mRNA level. The results of this study showed overlapping and specific proteomic responses to SCMV infection between resistant and susceptible ecotypes. After inoculation, the proteins involved in energy and metabolism, stress and defense responses, photosynthesis and other four functional groups showed significant changes in both ecotypes, which suggested that SCMV infection influenced these physiological processes in both the resistant Siyi and the susceptible Mo17. However, the oxidative burst was more pronounced during incompatible plant-SCMV interactions, as compared to those defined as compatible. We also observed an increase of enzymes involved in glycolysis and gluconeogenesis pathways in the resistant maize ecotype Siyi, while decrease in the susceptible maize ecotype Mo17. In addition, there is a marked increase of guanine nucleotide-binding protein beta submit in the resistant Siyi, which suggests a possible involvement of G-protein associated pathways in the resistant responses of maize to SCMV. These observations may possibly reveal protein targets/markers that are useful in the design of future diagnosis or plant protection strategies and provide new insights into the molecular mechanism of plant-virus interactions.

Shotgun proteomic analysis of plasma from dairy cattle suffering from footrot: characterization of potential disease-associated factors.

The plasma proteome of healthy dairy cattle and those with footrot was investigated using a shotgun LC-MS/MS approach. In total, 648 proteins were identified in healthy plasma samples, of which 234 were non-redundant proteins and 123 were high-confidence proteins; 712 proteins were identified from footrot plasma samples, of which 272 were non-redundant proteins and 138 were high-confidence proteins. The high-confidence proteins showed significant differences between healthy and footrot plasma samples in molecular weight, isoelectric points and the Gene Ontology categories. 22 proteins were found that may differentiate between the two sets of plasma proteins, of which 16 potential differential expression (PDE) proteins from footrot plasma involved in immunoglobulins, innate immune recognition molecules, acute phase proteins, regulatory proteins, and cell adhesion and cytoskeletal proteins; 6 PDE proteins from healthy plasma involved in regulatory proteins, cytoskeletal proteins and coagulation factors. Of these PDE proteins, haptoglobin, SERPINA10 protein, afamin precursor, haptoglobin precursor, apolipoprotein D, predicted peptidoglycan recognition protein L (PGRP-L) and keratan sulfate proteoglycan (KS-PG) were suggested to be potential footrot-associated factors. The PDE proteins PGRP-L and KS-PG were highlighted as potential biomarkers of footrot in cattle. The resulting protein lists and potential differentially expressed proteins may provide valuable information to increase understanding of plasma protein profiles in cattle and to assist studies of footrot-associated factors.

Proteomics analysis of differentially expressed proteins in schistosomula and adult worms of Schistosoma japonicum.

Schistosoma japonicum has a complex lifecycle and exhibits dramatic changes in its biology and morphology at different developmental stages. The schistosomulum and adult worm are two stages of this complex lifecycle and differentially expressed proteins in these two stages should be important for survival, development, and reproduction of the parasites. In this study, soluble and hydrophobic proteins were extracted from eggs, cercariae, schistosomula (8d and 19d), and male and female adult worms (42d) of Schistosoma japonicum, and separated by two-dimensional (2D) gel electrophoresis. A total of 1376±52, 928±61, 1465±41, 1230±30, 904±34, and 1080±26 soluble proteins and 1437±44, 845±53, 986±22, 1145±35, 1066±39, and 1123±45 hydrophobic proteins were separated from eggs, cercariae, schistosomula (8d and 19d), and male and female adult worms (42d), respectively. There were 65±14, 27±7, 37±17 and 48±9 soluble protein spots only present in schistosomula (8d and/or 19d) and adult schistosomes (male and/or female). We successfully identified 22 spots from schistosomula and 11 spots from adult schistosomes by mass spectrometry. Quantitative real-time RT-PCR was used to examine six differentially expressed proteins at the transcription level. These proteins only found in schistosomula or adults stage by the proteomics analysis were highly expressed in the corresponding stage at mRNA level. Bioinformatics analysis showed that the differentially expressed proteins from schistosomula were mainly involved in cellular metabolic processes, stress response and developmental process. Differentially expressed proteins from adult schistosomes were involved with gene expression and protein metabolism processes. The results of this study might provide new insights to stimulate further exploration of the mechanism of growth and development in schistosomes and help identify candidate molecules for developing new vaccines or drugs.

Proteomic changes in articular cartilage of human endemic osteoarthritis in China.

Kashin-Beck disease (KBD) is a chronic endemic osteochondropathy with unclear pathogenesis. It is a degenerative disease similar to osteoarthritis, but with different manifestations of cartilage damage. The aim of this investigation was to show the protein changes in KBD cartilage and to identify the candidate proteins in order to understand the pathogenesis of the disease. Proteins were extracted from the media of primary cell cultures of KBD and normal chondrocytes, and separated by two-dimensional fluorescence difference gel electrophoresis (2-D DIGE). MALDI-TOF/TOF analysis revealed statistically significant differences in 27 proteins from KBD chondrocyte cultures, which consisted of 17 up-regulated and ten down-regulated proteins. The results were further validated by Western blot analysis. The proteins identified are mainly involved in cellular redox homeostasis and stress response (MnSOD, Hsp27, Peroxiredoxin-1, and Cofilin-1), glycolysis (PGK-1, PGM-1, α-enolase), and cell motility and cytoskeletal organization (Actin, Calponin-2, and Keratin). These KBD-associated proteins indicate that cytoskeletal remodeling, glycometabolism, and oxidative stress are abnormal in KBD articular cartilage.

[The tear proteomics analysis of conjunctivochalasis].

OBJECTIVE: To analysis tear proteins by using shotgun strategy, and to study pathogenesis mechanism of conjunctivochalasis by comparing tear proteins between the conjunctivochalasis and normal body. METHODS: Tears were obtained from 8 conjunctivochalasis cases and 8 normal controls. Fifteen microliters of tears was collected by microcapillary tubes from each eye. Shotgun strategy was used for tear protein analysis. Trypsin digestion in-solution, separation of peptide mixture by reverse-phase high-pressure liquid chromatograph (RP-HPLC) followed by electrospray ionization mass spectrometry (ESI-MS/MS) identification and bioinformation analysis were used in this study. RESULTS: Three hundred and fifty-six proteins were excised in conjunctivochalasis patients and 352 proteins in the normal controls, and among them 119 proteins were the same. The functions of tear proteins were classified with GOA, which found some apoptosis regulation proteins and apoptosis related proteins and inflammatory response related proteins in conjuntivochalasis but not in normal controls group. Defensin was also found in conjuntivochalasis. CONCLUSION: Shotgun strategy can separate and analysis tear proteins effectively, which provide a new method for tear protein component of conjunctivochalasis. The special component in conjunctivochalasis tear show conjunctivochalasis maybe related to cell apoptosis and inflammatory.