scholarly journals Proteome Analysis of Differentially Displayed Proteins As a Tool for the Investigation of Symbiosis

2000 ◽  
Vol 13 (9) ◽  
pp. 995-1009 ◽  
Author(s):  
Siria H. A. Natera ◽  
Nelson Guerreiro ◽  
Michael A. Djordjevic
Keyword(s):  
Sinorhizobium Meliloti ◽  
Cultured Cells ◽  
Proteome Analysis ◽  
Sequence Information ◽  
Down Regulation ◽  
Symbiotic Interaction ◽  
Two Dimensional Gel Electrophoresis ◽  
Nitrogen Acquisition ◽  
Flight Mass Spectrometry ◽  
Peptide Mass

Two-dimensional gel electrophoresis was used to identify differentially displayed proteins expressed during the symbiotic interaction between the bacterium Sinorhizobium meliloti strain 1021 and the legume Melilotus alba (white sweetclover). Our aim was to characterize novel symbiosis proteins and to determine how the two symbiotic partners alter their respective metabolisms as part of the interaction, by identifying gene products that are differentially present between the symbiotic and non-symbiotic states. Proteome maps from control M. alba roots, wild-type nodules, cultured S. meliloti, and S. meliloti bacteroids were generated and compared. Over 250 proteins were induced or up-regulated in the nodule, compared with the root, and over 350 proteins were down-regulated in the bacteroid form of the rhizobia, compared with cultured cells. N-terminal amino acid sequencing and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry peptide mass fingerprint analysis, in conjunction with data base searching, were used to assign putative identity to nearly 100 nodule, bacterial, and bacteroid proteins. These included the previously identified nodule proteins leghemoglobin and NifH as well as proteins involved in carbon and nitrogen metabolism in S. meliloti. Bacteroid cells showed down-regulation of several proteins involved in nitrogen acquisition, including glutamine synthetase, urease, a urea-amide binding protein, and a PII isoform, indicating that the bacteroids were nitrogen proficient. The down-regulation of several enzymes involved in polyhydroxybutyrate synthesis and a cell division protein was also observed. This work shows that proteome analysis will be a useful strategy to link sequence information and functional genomics.

Comparative proteomic analysis on human L-02 liver cells treated with varying concentrations of trichloroethylene

2007 ◽  
Vol 23 (2) ◽  
pp. 91-101 ◽  
Author(s):  
Jianjun Liu ◽  
Haiyan Huang ◽  
Xiumei Xing ◽  
Renrong Xi ◽  
Zhixiong Zhuang ◽  
...  
Keyword(s):  
Proteomic Analysis ◽  
Underlying Mechanism ◽  
Liver Cells ◽  
Control Group ◽  
Two Dimensional Gel Electrophoresis ◽  
Comparative Proteomic Analysis ◽  
Maldi Tof ◽  
Flight Mass Spectrometry ◽  
Peptide Mass ◽  
Tof Ms

To determine the differential proteomic expressions in human L-02 liver cells induced by varying concentrations of trichloroethylene (TCE), comparative proteomic analysis was performed on human L-02 liver cells which were treated with varying concentrations of TCE. According to the result of MTT test, we designed four different groups, in which the cells were treated with 0 μM (control group), 3, 10 or 40 μM TCE for 24 h, respectively. Comparative analysis of approximately 800 spots resolved by two-dimensional gel electrophoresis (2DE) in the soluble proteomes of L-02 cells from the four different groups resulted in 10 differential proteins. To identify the differential spots, matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) was carried out; if the results from the tool were insufficient, tandem MS (MALDI-TOF-TOF-MS) was then performed. The raw data of peptide mass fingerprints (PMFs) and MS/MS spectra were searched against the IPI human data base for exact matches. Then western blot was employed to verify the result of proteomic analysis, the following result confirmed that the results of proteomic analysis were reliable. These results might provide an insight into the underlying mechanism of TCE intoxication and find biological markers for diagnosis and therapy of TCE-induced diseases.

scholarly journals Biotin Limitation in Sinorhizobium meliloti Strain 1021 Alters Transcription and Translation

2003 ◽  
Vol 69 (2) ◽  
pp. 1206-1213 ◽  
Author(s):  
Elke B. Heinz ◽  
Wolfgang R. Streit
Keyword(s):  
Sinorhizobium Meliloti ◽  
Proteome Analysis ◽  
Copper Resistance ◽  
Genome Database ◽  
Reading Frame ◽  
Flight Mass Spectrometry ◽  
Special Adaptation ◽  
Gene Coding ◽  
Meliloti Strain ◽  
Metabolic Activities

ABSTRACT Most Sinorhizobium meliloti strains lack several key genes involved in microbial biotin biosynthesis, and it is assumed that this may be a special adaptation which allows the microbe to down-regulate metabolic activities in the absence of a host plant. To further explore this hypothesis, we employed two different strategies. (i) Searches of the S. meliloti genome database in combination with the construction of nine different gusA reporter fusions identified three genes involved in a biotin starvation response in this microbe. A gene coding for a protein-methyl carboxyl transferase (pcm) exhibited 13.6-fold-higher transcription under biotin-limiting conditions than cells grown in the presence of 40 nM biotin. Consistent with this observation, biotin-limiting conditions resulted in a significantly decreased survival of pcm mutant cells compared to parental cells or cells grown in the presence of 40 nM biotin. Further studies indicated that the autoinducer synthase gene, sinI, was transcribed at a 4.5-fold-higher level in early stationary phase in biotin-starved cells than in biotin-supplemented cells. Lastly, we observed that open reading frame smc02283, which codes for a putative copper resistance protein (CopC), was 21-fold down-regulated in response to biotin starvation. (ii) In a second approach, proteome analysis identified 10 proteins which were significantly down-regulated under the biotin-limiting conditions. Among the proteins identified by using matrix-assisted laser desorption ionization-time of flight mass spectrometry were the ϖ subunit of the RNA polymerase and the 50S ribosomal protein L7/L12 (L8) subunit, indicating that biotin-limiting conditions generally affect transcription and translation in S. meliloti.

scholarly journals Functional Genomic Analysis of Global Regulator NolR in Sinorhizobium meliloti

2005 ◽  
Vol 18 (12) ◽  
pp. 1340-1352 ◽  
Author(s):  
Hancai Chen ◽  
Ke Gao ◽  
Eva Kondorosi ◽  
Adam Kondorosi ◽  
Barry G. Rolfe
Keyword(s):  
Sinorhizobium Meliloti ◽  
Regulatory Protein ◽  
Proteome Analysis ◽  
Peptide Mass Fingerprinting ◽  
Nucleotide Metabolism ◽  
Conjugative Transfer ◽  
Environmental Stimuli ◽  
Peptide Mass ◽  
Functional Genomic Analysis ◽  
Associated Proteins

NolR is a regulator of nodulation genes present in species belonging to the genera Rhizobium and Sinorhizobium. The expression of the nolR gene in Sinorhizobium meliloti AK631 was investigated in relation to stage of growth, availability of nutrients, and different environmental stimuli using the nolR::lacZ fusion report system. It has been shown that the nolR gene is regulated in a population-density-dependent fashion and influenced by a number of environmental stimuli, including nutrients, pH, and oxygen. Exploration of the physiological functions of NolR under various laboratory conditions has shown that NolR is required for the optimal growth of the bacteria on solid media, optimal survival of the bacteria in carbon-starved minimal medium, and after heat shock challenge. NolR also is involved in recipient-induced conjugative transfer of a plasmid. Proteome analysis of strain AK631 and its Tn5-induced nolR-deficient mutant EK698 revealed that a functional NolR induced significant differences in the accumulation of 20 polypeptides in peptide mass fingerprinting early-log-phase cultures and 48 polypeptides in stationary-phase cultures. NolR acted mainly as a repressor in the early-log-phase cultures, whereas it acted as both repressor and activator in the stationaryphase cultures. The NolR protein and 59 NolR-associated proteins have been identified by peptide mass fingerprinting. The NolR protein was differentially expressed only in the NolR+ wild-type strain AK631 but not in its NolR- derivative EK698, confirming that no functional NolR was produced in the mutant. The NolR-associated proteins have diverse functions in amino acid metabolism, carbohydrate metabolism, lipid metabolism, nucleotide metabolism, energy metabolism, metabolism of Co-factors, and cellular adaptation and transportation. These results further support our previous proposal that the NolR is a global regulatory protein which is required for the optimization of nodulation, bacterial growth and survival, and conjugative transfer of a plasmid.

Comparative proteome analysis of drought-sensitive and drought-tolerant maize leaves under osmotic stress

2019 ◽  
Vol 99 (4) ◽  
pp. 467-479
Author(s):  
Yuhe Pei ◽  
Jianfen Bai ◽  
Xinmei Guo ◽  
Meiai Zhao ◽  
Qingmei Ma ◽  
...  
Keyword(s):  
Osmotic Stress ◽  
Proteome Analysis ◽  
Peptide Mass Fingerprinting ◽  
Inbred Lines ◽  
Limiting Factor ◽  
Two Dimensional Gel Electrophoresis ◽  
Maize Production ◽  
Drought Tolerant ◽  
Maize Inbred Lines ◽  
Peptide Mass

Drought is a major yield-limiting factor in maize production. Osmotic stress was applied to two maize inbred lines with polyethylene glycol 6000 treatments. Proteins from the leaves were analyzed by two-dimensional gel electrophoresis and peptide mass fingerprinting at two time points, 24 and 48 h after osmotic stress. Thirty-five proteins were differentially expressed between control and treatment groups in the two maize inbred lines. In ‘Qi319’, a drought-tolerant inbred line, there were five up-regulated proteins at 24 h and 13 up-regulated and one down-regulated protein at 48 h. In drought-sensitive line ‘Zheng58’, 10 proteins were up-regulated at 24 h, while six proteins were up-regulated at 48 h. The 35 proteins were subjected to mass spectrometry and 17 proteins were successfully identified. These proteins were classified into six categories: photosynthesis-related, energy and metabolism, signaling pathways, protein synthesis, defense-related, and unclassified.

scholarly journals Comparative Analysis of Extracellular and Intracellular Proteomes of Listeria monocytogenes Strains Reveals a Correlation between Protein Expression and Serovar

2008 ◽  
Vol 74 (23) ◽  
pp. 7399-7409 ◽  
Author(s):  
Emilie Dumas ◽  
Bruno Meunier ◽  
Jean-Louis Berdagué ◽  
Christophe Chambon ◽  
Mickaël Desvaux ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Peptide Mass Fingerprinting ◽  
Etiologic Agent ◽  
Extracellular Proteins ◽  
Health Concern ◽  
Two Dimensional Gel Electrophoresis ◽  
Extracellular Proteome ◽  
Flight Mass Spectrometry ◽  
Peptide Mass ◽  
Phylogenetic Lineages

ABSTRACT Listeria monocytogenes, the etiologic agent of listeriosis, remains a serious public health concern, with its frequent occurrence in food environments coupled with a high mortality rate. Among the 13 serovars, human listeriosis is mostly associated with the serovar 4b, 1/2b, and 1/2a strains. To investigate the diversity of L. monocytogenes, the intracellular and extracellular proteins of 12 strains were analyzed by two-dimensional gel electrophoresis. These strains had different origins, belonged to different serovars (4b, 1/2a, and 1/2b), and presented with different levels of virulence in chicken embryos. The clustering of the strains in two groups based on proteomic patterns is in agreement with the L. monocytogenes phylogenetic lineages. Statistical analysis did not allow for identification of proteins specific to the isolate origin or the virulence level of the strains, but 26 and 21 protein spots were shown to be significantly overexpressed and underexpressed, respectively, in the six strains of serovar 1/2a (lineage II) compared to strains of serovar 1/2b or 4b. Moreover, a penicillin-binding protein was specific for serovar 1/2b and two protein spots identified as a serine protease were specific to serovar 4b. These protein spots, identified through peptide mass fingerprinting using matrix-assisted laser desorption ionization-time of flight mass spectrometry, were essentially found in the extracellular proteome and may have uses as potential markers for serotyping and risk analysis.

scholarly journals Proteomic Analysis of Wild-Type Sinorhizobium meliloti Responses to N-Acyl Homoserine Lactone Quorum-Sensing Signals and the Transition to Stationary Phase

2003 ◽  
Vol 185 (17) ◽  
pp. 5029-5036 ◽  
Author(s):  
Hancai Chen ◽  
Max Teplitski ◽  
Jayne B. Robinson ◽  
Barry G. Rolfe ◽  
Wolfgang D. Bauer
Keyword(s):  
Quorum Sensing ◽  
Stationary Phase ◽  
Proteomic Analysis ◽  
Sinorhizobium Meliloti ◽  
Proteome Analysis ◽  
Peptide Mass Fingerprinting ◽  
Homoserine Lactone ◽  
Wild Type ◽  
Carbon And Nitrogen ◽  
Peptide Mass

ABSTRACT Proteome analysis revealed that two long-chain N-acyl homoserine lactones (AHLs) produced by Sinorhizobium meliloti 1021 induced significant differences in the accumulation of more than 100 polypeptides in early-log-phase cultures of the wild type. Fifty-six of the corresponding proteins have been identified by peptide mass fingerprinting. The proteins affected by addition of these two AHLs had diverse functions in carbon and nitrogen metabolism, energy cycles, metabolite transport, DNA synthesis, and protein turnover. Two hours of exposure to 3-oxo-C16:1-homoserine lactone (3-oxo-C16:1-HL) affected the accumulation of 40 of the 56 identified proteins, whereas comparable exposure to C14-HL affected 13 of the 56 proteins. Levels of four proteins were affected by both AHLs. Exposure to 3-oxo-C16:1-HL for 8 h affected the accumulation of 17 proteins, 12 of which had reduced accumulation. Of the 80 proteins identified as differing in accumulation between early-log- and early-stationary-phase cultures, only 13 were affected by exposure to 3-oxo-C16:1-HL or C14-HL. These results provide a foundation for future studies of the functions regulated by AHL quorum sensing in S. meliloti and help to establish proteomic analysis as a powerful global approach to the identification of quorum-sensing regulatory patterns in wild-type bacteria.

scholarly journals Comparative Proteome Analysis of Brucella melitensis Vaccine Strain Rev 1 and a Virulent Strain, 16M

2002 ◽  
Vol 184 (18) ◽  
pp. 4962-4970 ◽  
Author(s):  
Michel Eschenbrenner ◽  
Mary Ann Wagner ◽  
Troy A. Horn ◽  
Jo Ann Kraycer ◽  
Cesar V. Mujer ◽  
...  
Keyword(s):  
Vaccine Strain ◽  
Brucella Melitensis ◽  
Virulent Strain ◽  
Iron Acquisition ◽  
Proteome Analysis ◽  
Peptide Mass Fingerprinting ◽  
Computer Assisted ◽  
Two Dimensional Gel Electrophoresis ◽  
Peptide Mass ◽  
Amino Acid Binding

ABSTRACT The genus Brucella consists of bacterial pathogens that cause brucellosis, a major zoonotic disease characterized by undulant fever and neurological disorders in humans. Among the different Brucella species, Brucella melitensis is considered the most virulent. Despite successful use in animals, the vaccine strains remain infectious for humans. To understand the mechanism of virulence in B. melitensis, the proteome of vaccine strain Rev 1 was analyzed by two-dimensional gel electrophoresis and compared to that of virulent strain 16M. The two strains were grown under identical laboratory conditions. Computer-assisted analysis of the two B. melitensis proteomes revealed proteins expressed in either 16M or Rev 1, as well as up- or down-regulation of proteins specific for each of these strains. These proteins were identified by peptide mass fingerprinting. It was found that certain metabolic pathways may be deregulated in Rev 1. Expression of an immunogenic 31-kDa outer membrane protein, proteins utilized for iron acquisition, and those that play a role in sugar binding, lipid degradation, and amino acid binding was altered in Rev 1.

Proteome Analysis of CD4+ T Cells Reveals Differentially Expressed Proteins in Infertile Polycystic Ovary Syndrome Patients

2020 ◽  
Vol 20 ◽  
Author(s):  
Fatemeh Nasri ◽  
Maryam Zare ◽  
Mehrnoosh Doroudchi ◽  
Behrouz Gharesi-Fard
Keyword(s):  
Mass Spectrometry ◽  
T Cells ◽  
T Lymphocytes ◽  
Gel Electrophoresis ◽  
Cd4 T Cells ◽  
Polycystic Ovary ◽  
Proteome Analysis ◽  
Two Dimensional ◽  
Ovary Syndrome ◽  
Two Dimensional Gel Electrophoresis

Background: Polycystic ovary syndrome (PCOS) is the most frequent endocrine disorder affecting 6–7% of premenopausal women. Recent studies revealed that the immune system especially CD4+ T helper cells are important in the context PCOS. Proteome analysis of CD4+ T lymphocytes can provide valuable information regarding the biology of these cells in the context of PCOS. Objective: To investigate immune dysregulation in CD4+ T lymphocytes at the protein level in the context of PCOS using two-dimensional gel electrophoresis (2DE) and mass spectrometry (MS). Methods: In the present study, we applied two-dimensional gel electrophoresis / mass spectrometry to identify proteins differentially expressed by peripheral blood CD4+ T cells in ten PCOS women compared with ten healthy women. Western blot technique was used to confirm the identified proteins. Results: Despite the overall proteome similarities, there were significant differences in the expression of seven spots between two groups (P <0.05). Three proteins, namely phosphatidylethanolamine-binding protein 1, proteasome activator complex subunit 1 and triosephosphate isomerase 1 were successfully identified by Mass technique and confirmed by western blot. All characterized proteins were over-expressed in CD4+ T cells from patients compared to CD4+ T cells from controls (P <0.05). In-silico analysis suggested that the over-expressed proteins interact with other proteins involved in cellular metabolism especially glycolysis and ferroptosis pathway. Conclusion: These findings suggest that metabolic adjustments in CD4+ T lymphocytes, which is in favor of increased glycolysis and Th2 differentiation are important in the context of PCOS.

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