Identification of lily pollen 14-3-3 isoforms and their subcellular and time-dependent expression profile

2011 ◽  
Vol 392 (3) ◽  
Author(s):  
Heidi Pertl ◽  
Simon Rittmann ◽  
Waltraud X. Schulze ◽  
Gerhard Obermeyer
Keyword(s):  
Expression Profile ◽  
Pollen Germination ◽  
Temporal Dynamics ◽  
Pollen Grains ◽  
Lilium Longiflorum ◽  
Mass Spectrometry Analysis ◽  
Primary Metabolism ◽  
Two Dimensional Gel Electrophoresis ◽  
Spectrometry Analysis ◽  
Lily Pollen

Abstract 14-3-3 proteins are major regulators in plant development and physiology including primary metabolism and signal transduction pathways, typically via a phosphorylation-dependent interaction with a target protein. Four full-length 14-3-3 isoforms were identified in pollen grains of Lilium longiflorum by screening of a cDNA library and RACE (rapid amplification of cDNA ends)-PCR. Mass spectrometry analysis of partially purified 14-3-3s confirmed the presence of the four isoforms but also indicated the presence of additional, less abundant 14-3-3 isoforms in lily pollen. Separation of partially purified 14-3-3 proteins by two-dimensional gel electrophoresis resulted in nine spots that mainly contained the four major 14-3-3 isoforms. In a first step to examine putative physiological roles of specific 14-3-3 isoforms, their subcellular expression profile during pollen germination and tube growth was monitored using a characterized set of antibodies against 14-3-3 proteins with distinct crossreactivity. The abundance profile of 14-3-3 proteins associated with the cytosol, endomembranes (tonoplast, endoplasmic reticulum, Golgi, mitochondria) and plasma membrane showed high spatial-temporal dynamics. This indicates different targets of 14-3-3 proteins at different organelles and time points during pollen germination and growth.

scholarly journals Impact of Manganese on Pollen Germination and Tube Growth in Lily

2021 ◽  
Vol 74 ◽  
Author(s):  
Thomas Sawidis ◽  
Gülriz Baycu ◽  
Elżbieta Weryszko-Chmielewska ◽  
Aneta Sulborska
Keyword(s):  
Pollen Tube ◽  
Pollen Germination ◽  
Surrounding Medium ◽  
Pollen Grains ◽  
Lilium Longiflorum ◽  
Tube Growth ◽  
Low Concentrations ◽  
Main Effect

Abstract In vitro culture of Lilium longiflorum pollen grains was carried out to determine the role of manganese in pollen germination and pollen tube growth. Pollen germination was adversely affected by the presence of manganese (>10 −8 M), whereas low concentrations (10 −12 –10 −10 M) stimulated the process. Manganese caused morphological anomalies during tube growth, characterized by irregular pollen tube thickening and swollen tips. The main effect was the anomalous cell wall formation at the tip, in which the presence of several organelles reduced the number of secretory vesicles. A loose network of fibrillar material and spherical aggregates, mostly in the tip region, was detected, and this material was progressively loosened into the surrounding medium. As a response to potential toxicity, the excess manganese was isolated in vacuoles, which formed an internal barrier against penetration of manganese to the tip area. Elevated manganese concentrations might affect plant reproduction, resulting in anomalies in gamete development. Consequently, the loss in genetic diversity and decreased fruit set ultimately lower yield.

scholarly journals Understanding Mechanisms of Salinity Tolerance in Barley by Proteomic and Biochemical Analysis of Near-Isogenic Lines

2020 ◽  
Vol 21 (4) ◽  
pp. 1516 ◽  
Author(s):  
Juan Zhu ◽  
Yun Fan ◽  
Sergey Shabala ◽  
Chengdao Li ◽  
Chao Lv ◽  
...  
Keyword(s):  
Salinity Tolerance ◽  
Crop Production ◽  
Biochemical Analysis ◽  
Mass Spectrometry Analysis ◽  
Near Isogenic Lines ◽  
Ros Scavenging ◽  
Hordeum Vulgare L ◽  
Two Dimensional Gel Electrophoresis ◽  
Spectrometry Analysis ◽  
Isogenic Lines

Salt stress is one of the major environmental factors impairing crop production. In our previous study, we identified a major QTL for salinity tolerance on chromosome 2H on barley (Hordeum vulgare L.). For further investigation of the mechanisms responsible for this QTL, two pairs of near-isogenic lines (NILs) differing in this QTL were developed. Sensitive NILs (N33 and N53) showed more severe damage after exposure to 300 mM NaCl than tolerant ones (T46 and T66). Both tolerant NILs maintained significantly lower Na+ content in leaves and much higher K+ content in the roots than sensitive lines under salt conditions, thus indicating the presence of a more optimal Na+/K+ ratio in plant tissues. Salinity stress caused significant accumulation of H2O2, MDA, and proline in salinity-sensitive NILs, and a greater enhancement in antioxidant enzymatic activities at one specific time or tissues in tolerant lines. One pair of NILs (N33 and T46) were used for proteomic studies using two-dimensional gel electrophoresis. A total of 53 and 51 differentially expressed proteins were identified through tandem mass spectrometry analysis in the leaves and roots, respectively. Proteins which are associated with photosynthesis, reactive oxygen species (ROS) scavenging, and ATP synthase were found to be specifically upregulated in the tolerant NIL. Proteins identified in this study can serve as a useful resource with which to explore novel candidate genes for salinity tolerance in barley.

scholarly journals Proteomic Analysis of Liver from Human Lipoprotein(a) Transgenic Mice Shows an Oxidative Stress and Lipid Export Response

2018 ◽  
Vol 2018 ◽  
pp. 1-11
Author(s):  
Euan J. Rodger ◽  
Carolyn M. Porteous ◽  
Gregory T. Jones ◽  
Michael Legge ◽  
Torsten Kleffmann ◽  
...  
Keyword(s):  
Human Liver ◽  
Density Lipoprotein ◽  
Bioinformatic Analysis ◽  
Liver Cells ◽  
Mass Spectrometry Analysis ◽  
Two Dimensional Gel Electrophoresis ◽  
Lipoprotein A ◽  
Spectrometry Analysis ◽  
Human Liver Cells ◽  
Proteomic Approach

Background. Mouse models of hypercholesterolaemia have been used to identify arterial proteins involved in atherosclerosis. As the liver is extremely sensitive to dyslipidemia, one might expect major changes in the abundance of liver proteins in these models even before atherosclerosis develops. Methods. Lipid levels were measured and a proteomic approach was used to quantify proteins in the livers of mice with an elevated low-density lipoprotein (LDL) and the presence of lipoprotein(a) [Lp(a)] but no atherosclerosis. Results. The livers of Lp(a) mice showed an increased triglyceride but reduced phospholipid and oxidised lipid content. Two-dimensional gel electrophoresis and mass spectrometry analysis identified 24 liver proteins with significantly increased abundance in Lp(a) mice (P<0.05). A bioinformatic analysis of the 24 proteins showed the major effect was that of an enhanced antioxidant and lipid efflux response with significant increases in antioxidant (Park7, Gpx1, Prdx6, and Sod1) and lipid metabolism proteins (Fabp4, Acaa2, apoA4, and ApoA1). Interestingly, human liver cells treated with Lp(a) showed significant increases in Gpx1 and Prdx6 but not Sod1 or Park7. Conclusions. The presence of human LDL and Lp(a) in mice promotes an enhanced flux of lipids into the liver which elicits an antioxidant and lipid export response before the onset of atherosclerosis. The antioxidant response can be reproduced in human liver cells treated with Lp(a).

scholarly journals Proteomic-Based Approach for the Identification of Tumor Markers Associated with Hepatocellular Carcinoma

2001 ◽  
Vol 17 (4) ◽  
pp. 217-223 ◽  
Author(s):  
Philip Shalhoub ◽  
Sarah Kern ◽  
Sophie Girard ◽  
Laura Beretta
Keyword(s):  
Hepatocellular Carcinoma ◽  
Tumor Antigens ◽  
Humoral Response ◽  
Mass Spectrometry Analysis ◽  
Major Type ◽  
Two Dimensional Gel Electrophoresis ◽  
Spectrometry Analysis ◽  
Immune Response To Cancer ◽  
Specific Proteins ◽  
The Common

There is increasing evidence for an immune response to cancer in humans, demonstrated in part by the identification of autoantibodies to tumor antigens. The identification of panels of tumor antigens that elicit a humoral response may have utility in cancer screening, diagnosis or in establishing prognosis. Several approaches are currently available for the identification of tumor antigens. We have used a proteomic-based approach for the identification of tumor antigens that induce an antibody response which we have applied to hepatocellular carcinoma, a major type of cancer worldwide. Two-dimensional gel electrophoresis allows simultaneous separation of several thousand individual proteins from tumor tissue or tumor cell lines. Proteins eliciting a humoral response in HCC were identified by 2-D Western blotting using sera from patients with hepatocellular carcinoma, followed by mass spectrometry analysis and database search. The common occurrence of autoantibodies to specific proteins may have utility for HCC screening and diagnosis.

Elevated Plasma Levels of Crosslinked Fibrinogen Gamma-chain Dimer Indicate Cancer-related Fibrin Deposition and Fibrinolysis

2001 ◽  
Vol 85 (03) ◽  
pp. 494-501 ◽  
Author(s):  
Werner Steinkellner ◽  
Klaus Holzmann ◽  
Andrea Gsur ◽  
Rudolf Grimm ◽  
Christian Ensinger ◽  
...  
Keyword(s):  
Solid Tumor ◽  
Acute Phase Proteins ◽  
Mass Spectrometry Analysis ◽  
Plasma Samples ◽  
Fibrin Deposition ◽  
Gamma Chain ◽  
Two Dimensional Gel Electrophoresis ◽  
Spectrometry Analysis ◽  
Chain Dimer ◽  
Potential Marker

SummaryCancer-related fibrin deposition and fibrinolysis were investigated by two-dimensional gel electrophoresis of human solid tumor and effusion specimen in addition to plasma samples. Fibrinogen gamma-chain dimer indicating fibrin deposition and plasmin-generated fibrinogen beta-chain fragments were identified in various solid tumor types by amino acid sequencing, mass spectrometry analysis and Western blotting. In tumor-associated effusions, these techniques allowed to observe plasmin-generated fragments of fibrinogen alpha, beta and gamma-chains in addition to elevated levels of acute-phase proteins. Similar observations were made in case of inflammation-associated effusions. No fibrin degradation product was observed in plasma samples, however, high amounts of fibrinogen gamma-chain dimer crosslinked by transglutaminase were detected in plasma from tumor patients, but not in plasma from controls and patients suffering acute infections and/or inflammations. This finding demonstrated that high transglutaminase activity may be associated with cancer. The presented data indicate that the amount of crosslinked fibrinogen gamma-chain dimer in plasma may correlate with tumor-associated fibrin deposition. The tumor-biological relevance of this potential marker protein is discussed.

scholarly journals Multiple plasma proteins control atrial natriuretic peptide (ANP) aggregation

2004 ◽  
Vol 33 (2) ◽  
pp. 335-341 ◽  
Author(s):  
C Torricelli ◽  
E Capurro ◽  
A Santucci ◽  
A Paffetti ◽  
C D’Ambrosio ◽  
...  
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Mass Spectrometry Analysis ◽  
High Density Lipoproteins ◽  
Protective Mechanism ◽  
Two Dimensional Gel Electrophoresis ◽  
Spectrometry Analysis ◽  
Dimeric Form ◽  
Atrial Natriuretic

We have recently demonstrated that human α-atrial natriuretic peptide (α-hANP), an amyloidogenic peptide responsible for isolated atrial amyloidosis, binds to a dimeric form of apo A-I belonging to small high-density lipoproteins (HDL). This binding phenomenon is considered a protective mechanism since it inhibits or strongly reduces the ANP aggregation process. The observation that plasma exhibits at least four times greater amyloid inhibitory activity than HDL prompted us to determine whether small HDL are the only ANP plasma-binding factors. After incubation of whole plasma with labelled ANP, the macromolecular complexes were subjected to two-dimensional gel electrophoresis followed by autoradiography. The results presented here provide novel evidence of additional binding proteins, in addition to apo A-I dimer, able to bind ANP in vitro and to prevent its aggregation. The mass spectrometry analysis of the radioactive spots identified them as albumin, α-1 antitrypsin, orosomucoid and apo A-IV-TTR complex. The putative impact of these findings in the amyloidogenic/antiamyloidogenic peptides network is discussed.

scholarly journals Modulation of Glucose Transport Causes Preferential Utilization of Aromatic Compounds in Pseudomonas putida CSV86

2007 ◽  
Vol 189 (21) ◽  
pp. 7556-7562 ◽  
Author(s):  
Aditya Basu ◽  
Rahul Shrivastava ◽  
Bhakti Basu ◽  
Shree K. Apte ◽  
Prashant S. Phale
Keyword(s):  
Organic Acids ◽  
Pseudomonas Putida ◽  
Glucose Transport ◽  
Aromatic Compounds ◽  
Carbon Sources ◽  
Mass Spectrometry Analysis ◽  
Periplasmic Space ◽  
Two Dimensional Gel Electrophoresis ◽  
Spectrometry Analysis ◽  
Glucose Binding

ABSTRACT Pseudomonas putida CSV86 utilizes aromatic compounds in preference to glucose and coutilizes aromatics and organic acids. Protein analysis of cells grown on different carbon sources, either alone or in combination, revealed that a 43-kDa periplasmic-space protein was induced by glucose and repressed by aromatics and succinate. Two-dimensional gel electrophoresis and liquid chromatography-tandem mass spectrometry analysis identified this protein as closely resembling the sugar ABC transporter of Pseudomonas putida KT2440. A partially purified 43-kDa protein showed glucose binding activity and was specific for glucose. The results demonstrate that the aromatic- and organic acid-mediated repression of a periplasmic-space glucose binding protein and consequent inhibition of glucose transport are responsible for this strain's ability to utilize aromatics and organic acids in preference to glucose.

scholarly journals Two-dimensional gel electrophoresis to identify arcelins from Phaseolus vulgaris with inmuno-proteomic analysis

2018 ◽  
Vol 36 (2) ◽  
pp. 114-119
Author(s):  
Carolina Bernal ◽  
Daynet Sosa ◽  
Iván Galindo-Castro ◽  
Nardy Diez
Keyword(s):  
Mass Spectrometry ◽  
Proteomic Analysis ◽  
Gel Electrophoresis ◽  
Mass Spectrometry Analysis ◽  
Two Dimensional ◽  
Storage Pests ◽  
Molecular Weight Range ◽  
Two Dimensional Gel Electrophoresis ◽  
Breeding Programs ◽  
Spectrometry Analysis

The present study used proteomics to analyze the expression of lectin-like proteins, specifically arcelins, in P. vulgaris cultivar varieties from Venezuela. A PAGE-SDS analysis of 30 commercial accessions of P. vulgaris showed significant differences in the molecular weight range of lectin-like proteins (arcelins). Eight different accessions were selected based on their electroforetic mobility for the proteomic analysis. Arcelin immuno-detection of two dimentional electrophoresed proteins was used to easily display the different arcelin proteomic profiles of the studied accessions. Mass spectrometry analysis confirmed the arcelin nature of these proteins. This is the first report on arcelin evaluation of the Venezuelan germoplasm of P. vulgaris with the aim of enhancing breeding programs by identifying accession materials with resistance to bean storage pests.

Comparative Proteomic Analysis of Hydrogen Peroxide-induced Protein Expression in Streptococcus pneumoniae D39

2021 ◽  
Vol 18 ◽  
Author(s):  
Sungkyoung Lee ◽  
Myoung-Ro Lee ◽  
Songmee Bae ◽  
Min-Kyu Kwak
Keyword(s):  
Streptococcus Pneumoniae ◽  
Protein Identification ◽  
Peptide Mapping ◽  
Virulent Strain ◽  
Anaerobic Growth ◽  
Differentially Expressed ◽  
Mass Spectrometry Analysis ◽  
Primary Metabolism ◽  
Human Respiratory Tract ◽  
Two Dimensional Gel Electrophoresis

Background: Streptococcus pneumoniae is a leading cause of human respiratory tract infection. Despite the lack of activities of antioxidative enzymes, including cytochromes, hemoproteins, and peroxidases/catalases, traits conferring the aerotolerant-anaerobic growth of this bacterium are conserved, with high efficacy of antioxidative actions, in an oxygen-rich environment. Objective: Through proteome analysis, this study’s intention was to evaluate differentially expressed proteins and/or gene products modeled in a highly virulent strain, S. pneumoniae D39, exogenously-treated with millimolar concentrations of H2O2. Method: For two-dimensional gel electrophoresis (2-DE) analysis, following one dimensional isoelectric focusing with an immobilized pH gradient of pH 4-7, the most significantly mobilized proteins expressed were separated by SDS-PAGE in the second dimension. With a total of 431 protein spots detected, certain proteins were excised, in-gel trypsin digested, and analyzed by combination with MALDI-TOF and LC-ESI-MS/MS for mass spectrometric peptide mapping and protein identification. Utilizing mass spectrometry analysis of spots excised from 2-DE, the selected protein spots were identified with a variety of databases and MASCOT. Results: With the aid of comparisons to proteome reference maps, the most differentially expressed 38 proteins, those with approximately 1.4-fold or more increase and/or decrease or with multiple isoforms exhibiting variable pI values, were induced by treatment of exogenous 2 mM H2O2. The identified proteins were seen to be involved in pneumococcal pathogenesis and primary metabolism, amongst others. Conclusion: This is the first study to convincingly document proteomic information associated with pathophysiological adaptation under the given oxidative conditions, and corresponding potential antioxidative mechanisms, in S. pneumoniae.

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