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Title: A genetic screen to isolate type III effectors translocated into pepper cells during Xanthomonas infection

Abstract

The bacterial pathogen Xanthomonas campestris pv. vesicatoria (Xcv) uses a type III secretion system (TTSS) to translocate effector proteins into host plant cells. The TTSS is required for Xcv colonization, yet the identity of many proteins translocated through this apparatus is not known. We used a genetic screen to functionally identify Xcv TTSS effectors. A transposon 5 (Tn5)-based transposon construct including the coding sequence for the Xcv AvrBs2 effector devoid of its TTSS signal was randomly inserted into the Xcv genome. Insertion of the avrBs2 reporter gene into Xcv genes coding for proteins containing a functional TTSS signal peptide resulted in the creation of chimeric TTSS effector::AvrBs2 fusion proteins. Xcv strains containing these fusions translocated the AvrBs2 reporter in a TTSS-dependent manner into resistant BS2 pepper cells during infection, activating the avrBs2-dependent hypersensitive response (HR). We isolated seven chimeric fusion proteins and designated the identified TTSS effectors as Xanthomonas outer proteins (Xops). Translocation of each Xop was confirmed by using the calmodulin-dependent adenylate cydase reporter assay. Three xop genes are Xanthomonas spp.-specific, whereas homologs for the rest are found in other phytopathogenic bacteria. XopF1 and XopF2 define an effector gene family in Xcv. XopN contains a eukaryotic protein fold repeatmore » and is required for full Xcv pathogenicity in pepper and tomato. The translocated effectors identified in this work expand our knowledge of the diversity of proteins that Xcv uses to manipulate its hosts.« less

Publication Date:
Research Org.:
Stanford University, Stanford, CA
Sponsoring Org.:
USDOE Office of Science (SC)
OSTI Identifier:
947124
Report Number(s):
DOE-ER15443 Final Report
DOE Contract Number:  
FG02-03ER15443
Resource Type:
Journal Article
Journal Name:
PNAS
Additional Journal Information:
Journal Volume: 101; Journal Issue: 47
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; bacterial plant pathogenesis, virulence proteins

Citation Formats

. A genetic screen to isolate type III effectors translocated into pepper cells during Xanthomonas infection. United States: N. p., 2004. Web. doi:10.1073/pnas.0407383101.
. A genetic screen to isolate type III effectors translocated into pepper cells during Xanthomonas infection. United States. https://doi.org/10.1073/pnas.0407383101
. 2004. "A genetic screen to isolate type III effectors translocated into pepper cells during Xanthomonas infection". United States. https://doi.org/10.1073/pnas.0407383101.
@article{osti_947124,
title = {A genetic screen to isolate type III effectors translocated into pepper cells during Xanthomonas infection},
author = {},
abstractNote = {The bacterial pathogen Xanthomonas campestris pv. vesicatoria (Xcv) uses a type III secretion system (TTSS) to translocate effector proteins into host plant cells. The TTSS is required for Xcv colonization, yet the identity of many proteins translocated through this apparatus is not known. We used a genetic screen to functionally identify Xcv TTSS effectors. A transposon 5 (Tn5)-based transposon construct including the coding sequence for the Xcv AvrBs2 effector devoid of its TTSS signal was randomly inserted into the Xcv genome. Insertion of the avrBs2 reporter gene into Xcv genes coding for proteins containing a functional TTSS signal peptide resulted in the creation of chimeric TTSS effector::AvrBs2 fusion proteins. Xcv strains containing these fusions translocated the AvrBs2 reporter in a TTSS-dependent manner into resistant BS2 pepper cells during infection, activating the avrBs2-dependent hypersensitive response (HR). We isolated seven chimeric fusion proteins and designated the identified TTSS effectors as Xanthomonas outer proteins (Xops). Translocation of each Xop was confirmed by using the calmodulin-dependent adenylate cydase reporter assay. Three xop genes are Xanthomonas spp.-specific, whereas homologs for the rest are found in other phytopathogenic bacteria. XopF1 and XopF2 define an effector gene family in Xcv. XopN contains a eukaryotic protein fold repeat and is required for full Xcv pathogenicity in pepper and tomato. The translocated effectors identified in this work expand our knowledge of the diversity of proteins that Xcv uses to manipulate its hosts.},
doi = {10.1073/pnas.0407383101},
url = {https://www.osti.gov/biblio/947124}, journal = {PNAS},
number = 47,
volume = 101,
place = {United States},
year = {Tue Nov 23 00:00:00 EST 2004},
month = {Tue Nov 23 00:00:00 EST 2004}
}