scholarly journals Identification of Novel Virulence-Associated Proteins Secreted to Xylem by Verticillium nonalfalfae During Colonization of Hop Plants

2016 ◽  
Vol 29 (5) ◽  
pp. 362-373 ◽  
Author(s):  
Marko Flajsman ◽  
Stanislav Mandelc ◽  
Sebastjan Radisek ◽  
Natasa Stajner ◽  
Jernej Jakse ◽  
...  
Keyword(s):  
Plant Pathogens ◽  
Xylem Sap ◽  
Hypothetical Protein ◽  
Secreted Proteins ◽  
In Planta ◽  
Fungal Virulence ◽  
Associated Proteins ◽  
Verticillium Nonalfalfae ◽  
Identification And Characterization

Plant pathogens employ various secreted proteins to suppress host immunity for their successful host colonization. Identification and characterization of pathogen-secreted proteins can contribute to an understanding of the pathogenicity mechanism and help in disease control. We used proteomics to search for proteins secreted to xylem by the vascular pathogen Verticillium nonalfalfae during colonization of hop plants. Three highly abundant fungal proteins were identified: two enzymes, α-N-arabinofuranosidase (VnaAbf4.216) and peroxidase (VnaPRX1.1277), and one small secreted hypothetical protein (VnaSSP4.2). These are the first secreted proteins so far identified in xylem sap following infection with Verticillium spp. VnaPRX1.1277, classified as a heme-containing peroxidase from Class II, similar to other Verticillium spp. lignin-degrading peroxidases, and VnaSSP4.2, a 14-kDa cysteine-containing protein with unknown function and with a close homolog in related V. alfalfae strains, were further examined. The in planta expression of VnaPRX1.1277 and VnaSSP4.2 genes increased with the progression of colonization, implicating their role in fungal virulence. Indeed, V. nonalfalfae deletion mutants of both genes exhibited attenuated virulence on hop plants, which returned to the level of the wild-type pathogenicity in the knockout complementation lines, supporting VnaPRX1.1277 and VnaSSP4.2 as virulence factors required to promote V. nonalfalfae colonization of hop plants.

scholarly journals Identification and Characterization of Two Novel Proteins Affecting Fission Yeast γ-tubulin Complex Function

2004 ◽  
Vol 15 (5) ◽  
pp. 2287-2301 ◽  
Author(s):  
Srinivas Venkatram ◽  
Joseph J. Tasto ◽  
Anna Feoktistova ◽  
Jennifer L. Jennings ◽  
Andrew J. Link ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Complex Function ◽  
Novel Proteins ◽  
Astral Microtubule ◽  
Different Types ◽  
Associated Proteins ◽  
Microtubule Formation ◽  
Identification And Characterization ◽  
Microtubule Function

The γ-tubulin complex, via its ability to organize microtubules, is critical for accurate chromosome segregation and cytokinesis in the fission yeast, Schizosaccharomyces pombe. To better understand its roles, we have purified the S. pombe γ-tubulin complex. Mass spectrometric analyses of the purified complex revealed known components and identified two novel proteins (i.e., Mbo1p and Gfh1p) with homology to γ-tubulin–associated proteins from other organisms. We show that both Mbo1p and Gfh1p localize to microtubule organizing centers. Although cells deleted for either mbo1+ or gfh1+ are viable, they exhibit a number of defects associated with altered microtubule function such as defects in cell polarity, nuclear positioning, spindle orientation, and cleavage site specification. In addition, mbo1Δ and gfh1Δ cells exhibit defects in astral microtubule formation and anchoring, suggesting that these proteins have specific roles in astral microtubule function. This study expands the known roles of γ-tubulin complex components in organizing different types of microtubule structures in S. pombe.

scholarly journals Comprehensive analysis of Verticillium nonalfalfae in silico secretome uncovers putative effector proteins expressed during hop invasion

2017 ◽  
Author(s):  
Kristina Marton ◽  
Marko Flajšman ◽  
Sebastjan Radišek ◽  
Katarina Košmelj ◽  
Jernej Jakše ◽  
...  
Keyword(s):  
Verticillium Wilt ◽  
Sequence Similarity ◽  
Vascular Plant ◽  
Xylem Sap ◽  
Fungal Growth ◽  
Effector Proteins ◽  
Specific Gene ◽  
In Planta ◽  
Verticillium Nonalfalfae ◽  
Spatio Temporal

AbstractBackgroundThe vascular plant pathogen Verticillium nonalfalfae causes Verticillium wilt in several important crops. VnaSSP4.2 was recently discovered as a V. nonalfalfae virulence effector protein in the xylem sap of infected hop. Here, we expanded our search for candidate secreted effector proteins (CSEPs) in the V. nonalfalfae predicted secretome using a bioinformatic pipeline built on V. nonalfalfae genome data, RNA-Seq and proteomic studies of the interaction with hop.ResultsThe secretome, rich in carbohydrate active enzymes, proteases, redox proteins and proteins involved in secondary metabolism, cellular processing and signaling, includes 263 CSEPs. Several homologs of known fungal effectors (LysM, NLPs, Hce2, Cerato-platanins, Cyanovirin-N lectins, hydrophobins and CFEM domain containing proteins) and avirulence determinants in the PHI database (Avr-Pita1 and MgSM1) were found. The majority of CSEPs were non-annotated and were narrowed down to 44 top priority candidates based on their likelihood of being effectors. These were examined by spatio-temporal gene expression profiling of infected hop. Among the highest in planta expressed CSEPs, five deletion mutants were tested in pathogenicity assays. A deletion mutant of VnaUn.279, a lethal pathotype specific gene with sequence similarity to SAM-dependent methyltransferase (LaeA), had lower infectivity and showed highly reduced virulence, but no changes in morphology, fungal growth or conidiation were observed.ConclusionsSeveral putative secreted effector proteins that probably contribute to V. nonalfalfae colonization of hop were identified in this study. Among them, LaeA gene homolog was found to act as a potential novel virulence effector of V. nonalfalfae. The combined results will serve for future characterization of V. nonalfalfae effectors, which will advance our understanding of Verticillium wilt disease.

scholarly journals Identification and Characterization of a Gene Cluster for Synthesis of the Polyketide Antibiotic 2,4-Diacetylphloroglucinol from Pseudomonas fluorescens Q2-87

1999 ◽  
Vol 181 (10) ◽  
pp. 3155-3163 ◽  
Author(s):  
M. Gita Bangera ◽  
Linda S. Thomashow
Keyword(s):  
Genomic Dna ◽  
Plant Pathogens ◽  
Polyketide Synthase ◽  
Open Reading Frames ◽  
Fungal Plant Pathogens ◽  
Repressor Proteins ◽  
Flanking Regions ◽  
Identification And Characterization ◽  
Reading Frames

The polyketide metabolite 2,4-diacetylphloroglucinol (2,4-DAPG) is produced by many strains of fluorescent Pseudomonas spp. with biocontrol activity against soilborne fungal plant pathogens. Genes required for 2,4-DAPG synthesis by P. fluorescensQ2-87 are encoded by a 6.5-kb fragment of genomic DNA that can transfer production of 2,4-DAPG to 2,4-DAPG-nonproducing recipientPseudomonas strains. In this study the nucleotide sequence was determined for the 6.5-kb fragment and flanking regions of genomic DNA from strain Q2-87. Six open reading frames were identified, four of which (phlACBD) comprise an operon that includes a set of three genes (phlACB) conserved between eubacteria and archaebacteria and a gene (phlD) encoding a polyketide synthase with homology to chalcone and stilbene synthases from plants. The biosynthetic operon is flanked on either side by phlEand phlF, which code respectively for putative efflux and regulatory (repressor) proteins. Expression in Escherichia coli of phlA, phlC, phlB, andphlD, individually or in combination, identified a novel polyketide biosynthetic pathway in which PhlD is responsible for the production of monoacetylphloroglucinol (MAPG). PhlA, PhlC, and PhlB are necessary to convert MAPG to 2,4-DAPG, and they also may function in the synthesis of MAPG.

scholarly journals Identification and Characterization of Phytoplasmal Genes, Employing a Novel Method of Isolating Phytoplasmal Genomic DNA

2003 ◽  
Vol 185 (22) ◽  
pp. 6513-6521 ◽  
Author(s):  
Sharon Melamed ◽  
Edna Tanne ◽  
Raz Ben-Haim ◽  
Orit Edelbaum ◽  
David Yogev ◽  
...  
Keyword(s):  
Genomic Dna ◽  
Plant Pathogens ◽  
Open Reading Frames ◽  
Rrna Genes ◽  
Unique Method ◽  
Novel Method ◽  
Identification And Characterization ◽  
Reading Frames ◽  
Large Background

ABSTRACT Phytoplasmas are unculturable, insect-transmissible plant pathogens belonging to the class Mollicutes. To be transmitted, the phytoplasmas replicate in the insect body and are delivered to the insect's salivary glands, from where they are injected into the recipient plant. Because phytoplasmas cannot be cultured, any attempt to recover phytoplasmal DNA from infected plants or insects has resulted in preparations with a large background of host DNA. Thus, studies of the phytoplasmal genome have been greatly hampered, and aside from the rRNA genes, only a few genes have hitherto been isolated and characterized. We developed a unique method to obtain host-free phytoplasmal genomic DNA from the insect vector's saliva, and we demonstrated the feasibility of this method by isolating and characterizing 78 new putative phytoplasmal open reading frames and their deduced proteins. Based on the newly accumulated information on phytoplasmal genes, preliminary characteristics of the phytoplasmal genome are discussed.

scholarly journals Identification and Characterization of a Candida albicans Mating Pheromone

2003 ◽  
Vol 23 (22) ◽  
pp. 8189-8201 ◽  
Author(s):  
Richard J. Bennett ◽  
M. Andrew Uhl ◽  
Mathew G. Miller ◽  
Alexander D. Johnson
Keyword(s):  
Candida Albicans ◽  
Fungal Pathogen ◽  
Cell Communication ◽  
Secreted Proteins ◽  
Pheromone Response ◽  
Mating Pheromone ◽  
A Cells ◽  
Bona Fide ◽  
Identification And Characterization

ABSTRACT Candida albicans, the most prevalent fungal pathogen of humans, has recently been shown to undergo mating. Here we describe a mating pheromone produced by C. albicans α cells and show that the gene which encodes it (MFα) is required for α cells, but not a cells, to mate. We also identify the receptor for this mating pheromone as the product of the STE2 gene and show that this gene is required for the mating of a cells, but not α cells. Cells of the a mating type respond to the α mating pheromone by producing long polarized projections, similar to those observed in bona fide mating mixtures of C. albicans a and α cells. During this process, transcription of approximately 62 genes is induced. Although some of these genes correspond to those induced in Saccharomyces cerevisiae by S. cerevisiae α-factor, most are specific to the C. albicans pheromone response. The most surprising class encode cell surface and secreted proteins previously implicated in virulence of C. albicans in a mouse model of disseminated candidiasis. This observation suggests that aspects of cell-cell communication in mating may have been evolutionarily adopted for host-pathogen interactions in C. albicans.

scholarly journals Characterization of aStaphylococcus aureusSurface Virulence Factor That Promotes Resistance to Oxidative Killing and Infectious Endocarditis

2010 ◽  
Vol 79 (1) ◽  
pp. 342-352 ◽  
Author(s):  
Natalia Malachowa ◽  
Petra L. Kohler ◽  
Patrick M. Schlievert ◽  
Olivia N. Chuang ◽  
Gary M. Dunny ◽  
...  
Keyword(s):  
Bacterial Infections ◽  
Hypothetical Protein ◽  
Cell Number ◽  
Deletion Strain ◽  
Human Neutrophils ◽  
Bacterial Cell Number ◽  
Hospital Acquired ◽  
Identification And Characterization

ABSTRACTStaphylococcus aureusis a prominent human pathogen and a leading cause of community- and hospital-acquired bacterial infections worldwide. Herein, we describe the identification and characterization of theS. aureus67.6-kDa hypothetical protein, named for thesurface factor promoting resistance tooxidativekilling (SOK) in this study. Sequence analysis showed that the SOK gene is conserved in all sequencedS. aureusstrains and homologous to the myosin cross-reactive antigen ofStreptococcus pyogenes.Immunoblotting and immunofluorescence analysis showed that SOK was copurified with membrane fractions and was exposed on the surface ofS. aureusNewman and RN4220. Comparative analysis of wild-typeS. aureusand an isogenic deletion strain indicated that SOK contributes to both resistance to killing by human neutrophils and to oxidative stress. In addition, theS. aureus sokdeletion strain showed dramatically reduced aortic valve vegetation and bacterial cell number in a rabbit endocarditis model. These results, plus the suspected role of the streptococcal homologue in certain diseases such as acute rheumatic fever, suggest that SOK plays an important role in cardiovascular and other staphylococcal infections.

scholarly journals Systematic Identification and Characterization of Novel Human Skin-Associated Genes Encoding Membrane and Secreted Proteins

PLoS ONE ◽  
2013 ◽  
Vol 8 (6) ◽  
pp. e63949 ◽  
Author(s):  
Peter Arne Gerber ◽  
Peter Hevezi ◽  
Bettina Alexandra Buhren ◽  
Cynthia Martinez ◽  
Holger Schrumpf ◽  
...  
Keyword(s):  
Human Skin ◽  
Secreted Proteins ◽  
Genes Encoding ◽  
Systematic Identification ◽  
Identification And Characterization

scholarly journals Characterization of the RND-Type Multidrug Efflux Pump MexAB-OprM of the Plant Pathogen Pseudomonas syringae

2008 ◽  
Vol 74 (11) ◽  
pp. 3387-3393 ◽  
Author(s):  
Savina O. Stoitsova ◽  
Yvonne Braun ◽  
Matthias S. Ullrich ◽  
Helge Weingart
Keyword(s):  
Pseudomonas Syringae ◽  
Plant Pathogens ◽  
Efflux Pump ◽  
Blot Hybridization ◽  
In Planta ◽  
Multidrug Efflux ◽  
Dot Blot ◽  
Multidrug Efflux Pump

ABSTRACT In gram-negative bacteria, transporters belonging to the RND family are the transporters most relevant for resistance to antimicrobial compounds. In Pseudomonas aeruginosa, a clinically important pathogen, the RND-type pump MexAB-OprM has been recognized as one of the major multidrug efflux systems. Here, homologues of MexAB-OprM in the plant pathogens Pseudomonas syringae pv. phaseolicola 1448A, P. syringae pv. syringae B728a, and P. syringae pv. tomato DC3000 were identified, and mexAB-oprM-deficient mutants were generated. Determination of MICs revealed that mutation of MexAB-OprM dramatically reduced the tolerance to a broad range of antimicrobials. Moreover, the ability of the mexAB-oprM-deficient mutants to multiply in planta was reduced. RNA dot blot hybridization revealed growth-dependent regulation of the mexAB-oprM operon in P. syringae; the expression of this operon was maximal in early exponential phase and decreased gradually during further growth.

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