Biomedical subjects

Pierre R Fobert

Publications and source records attributed to Pierre R Fobert.

9 recordsLinked to original sources

PubMed2006

The coactivator function of Arabidopsis NPR1 requires the core of its BTB/POZ domain and the oxidation of C-terminal cysteines.

NONEXPRESSOR OF PATHOGENESIS-RELATED GENES1 (NPR1) regulates systemic acquired resistance (SAR) in Arabidopsis thaliana, and current models propose that after treatment with salicylic acid (SA), Cys-82 and Cys-216 of NPR1 are reduced, leading to nuclear import. The interaction of nucleus-localized NPR1 with TGA transcription factors results in the activation of defense genes, including the SAR marker PATHOGENESIS-RELATED-1 (PR-1), and the deployment of SAR. Little is known about how TGA factors or NPR1 regulate transcription or whether a TGA-NPR1 complex forms on DNA. We show that TGA2 and NPR1 are recruited to PR-1 independently of each other and of SA treatment. Consistent with the result that a triple knockout in TGA2/5/6 derepresses PR-1, in vivo plant transcription assays revealed that TGA2 is not an autonomous transcription activator but is a transcriptional repressor in both untreated and SA-treated cells. However, after stimulation with SA, TGA2 is incorporated into a transactivating complex with NPR1, forming an enhanceosome that requires the core of the NPR1 BTB/POZ domain (residues 80 to 91) and the oxidation of NPR1 Cys-521 and Cys-529. These Cys residues are found in a new type of transactivation domain that we term Cys-oxidized. These data further our understanding of the mechanism by which TGA2 and NPR1 activate Arabidopsis PR-1.

Amino Acid Sequence↗

PubMed2005

Comparison of transcript profiling on Arabidopsis microarray platform technologies.

To date there have been few systematic studies comparing the results of transcript profiling from different microarray platform technologies. We evaluated in detail two different Arabidopsis thaliana microarray platforms: our own Genomic Amplicon arrays and the Qiagen long oligonucleotide arrays designed by Operon; furthermore, we cross-validated these arrays against the Affymetrix AG and ATH1 GeneChips. Data were obtained from all three platforms in each of two separate experiments; (1) at 2 h and (2) 8 h following a salicylic acid treatment applied to both wild-type and npr1-3 mutant plants. A total of 20 hybridizations were performed, analyzing the expression of 26,814 unique locus IDs. We demonstrate that intensity rank is a key variable that affects both inter-platform and cross-platform reproducibility. Although general agreement between platform technologies is low, data derived from high signal intensities (90th percentile) can correlate as well between differing platforms as replicates within the same platform (r=0.4-0.7). We also show that the identification of differentially expressed genes by significance analysis of microarrays is influenced by signal intensity and that overlap between significant gene lists from different platform technologies was as high as 67% when low intensity values were removed. Validation of 41 genes by Northern blot hybridization showed that all platform technologies performed well, qualitatively confirming 83-100% of differential gene expression. Our results suggest that the potential for the broad integration of microarray data from different platforms and laboratories is promising.

Arabidopsis↗

PubMed2005

Redox control of systemic acquired resistance.

Changes in gene expression during systemic acquired resistance (SAR) require the phenolic signaling molecule salicylic acid (SA) and are modulated by the interaction between the NON EXPRESSOR OF PATHOGENESIS-RELATED GENES1 (NPR1) protein and members of the TGA family of transcription factors. In the past two years, the activities of NPR1 and of the TGA factors TGA1 and TGA4 have been shown to be modulated by SA-induced oxidoreduction (redox) modifications of key cysteine residues. Reduction of two conserved cysteines in NPR1 leads to its monomerization and nuclear localization, which is required for the activation of PATHOGENESIS-RELATED(PR) genes. Reduction of conserved cysteines in TGA1 and TGA4 enables their interaction with NPR1, which acts as a redox-sensitive cofactor in stimulating TGA1 DNA-binding activity. The identity of the redox mediators that are involved in regulating NPR1 and TGA factors is unknown. However, a novel thioredoxin interacts with the disease resistance protein Cf-9 and modulates Cf-9-dependent signaling. These results begin to provide a mechanistic understanding of how SAR is regulated by redox conditions.

Gene Expression Regulation, Plant↗

PubMed2005

An Arabidopsis NPR1-like gene, NPR4, is required for disease resistance.

The Arabidopsis genome contains six NPR1-related genes. Given the pivotal role played by NPR1 in controlling salicylic acid (SA)-mediated gene expression and disease resistance, functional characterization of other family members appears to be justified. Reverse genetics was used to analyze the role of one NPR1-like gene, which we called NPR4. The NPR4 protein shares 36% identity with NPR1 and interacts with the same spectrum of TGA transcription factors in yeast two-hybrid assays. Plants with T-DNA insertions in NPR4 are more susceptible to the virulent bacterial pathogen Pseudomonas syringe pv. tomato DC3000. This phenotype is complemented by expression of the wild type NPR4 coding region. As determined by the parasite reproduction, the npr4-1 mutant is more susceptible to the fungal pathogen Erysiphe cichoracearum, but does not differ markedly from wild type in its interaction with virulent and avirulent strains of the oomycete Peronospora parasitica. In leaves of wild-type plants, NPR4 mRNA levels increase following pathogen challenge or SA treatment, and decrease rapidly following methyl jasmonic acid (MeJA) treatment. Transcripts of the pathogenesis-related (PR) genes PR-1, PR-2, and PR-5 are only marginally reduced in the npr4-1 mutant following pathogen challenge or SA treatment. This reduction of PR gene expression is more pronounced when leaves are challenged with the bacterial pathogen following SA treatment. Expression of the jasmonic acid-dependent pathway marker gene PDF1.2 is compromised in npr4-1 leaves following application of MeJA or a combination of SA and MeJA. These results indicate that NPR4 is required for basal defense against pathogens, and that it may be implicated in the cross-talk between the SA- and JA-dependent signaling pathways.

Amino Acid Sequence↗

PubMed2004

A "Whirly" transcription factor is required for salicylic acid-dependent disease resistance in Arabidopsis.

Transcriptional reprogramming is critical for plant disease resistance responses; its global control is not well understood. Salicylic acid (SA) can induce plant defense gene expression and a long-lasting disease resistance state called systemic acquired resistance (SAR). Plant-specific "Whirly" DNA binding proteins were previously implicated in defense gene regulation. We demonstrate that the potato StWhy1 protein is a transcriptional activator of genes containing the PBF2 binding PB promoter element. DNA binding activity of AtWhy1, the Arabidopsis StWhy1 ortholog, is induced by SA and is required for both SA-dependent disease resistance and SA-induced expression of an SAR response gene. AtWhy1 is required for both full basal and specific disease resistance responses. The transcription factor-associated protein NPR1 is also required for SAR. Surprisingly, AtWhy1 activation by SA is NPR1 independent, suggesting that AtWhy1 works in conjunction with NPR1 to transduce the SA signal. Our analysis of AtWhy1 adds a critical component to the SA-dependent plant disease resistance response.

Aldehyde Oxidoreductases↗

PubMed2004

Discovery of functional genes for systemic acquired resistance in Arabidopsis thaliana through integrated data mining.

Various data mining techniques combined with sequence motif information in the promoter region of genes were applied to discover functional genes that are involved in the defense mechanism of systemic acquired resistance (SAR) in Arabidopsis thaliana. A series of K-Means clustering with difference-in-shape as distance measure was initially applied. A stability measure was used to validate this clustering process. A decision tree algorithm with the discover-and-mask technique was used to identify a group of most informative genes. Appearance and abundance of various transcription factor binding sites in the promoter region of the genes were studied. Through the combination of these techniques, we were able to identify 24 candidate genes involved in the SAR defense mechanism. The candidate genes fell into 2 highly resolved categories, each category showing significantly unique profiles of regulatory elements in their promoter regions. This study demonstrates the strength of such integration methods and suggests a broader application of this approach.

Algorithms↗

PubMed2003

The tomato transcription factor Pti4 regulates defense-related gene expression via GCC box and non-GCC box cis elements.

The tomato transcription factor Pti4, an ethylene-responsive factor (ERF), interacts physically with the disease resistance protein Pto and binds the GCC box cis element that is present in the promoters of many pathogenesis-related (PR) genes. We reported previously that Arabidopsis plants expressing Pti4 constitutively express several GCC box-containing PR genes and show reduced disease symptoms compared with wild-type plants after inoculation with Pseudomonas syringae pv tomato or Erysiphe orontii. To gain insight into how genome-wide gene expression is affected by Pti4, we used serial analysis of gene expression (SAGE) to compare transcripts in wild-type and Pti4-expressing Arabidopsis plants. SAGE provided quantitative measurements of >20,000 transcripts and identified the 50 most highly expressed genes in Arabidopsis vegetative tissues. Comparison of the profiles from wild-type and Pti4-expressing Arabidopsis plants revealed 78 differentially abundant transcripts encoding defense-related proteins, protein kinases, ribosomal proteins, transporters, and two transcription factors (TFs). Many of the genes identified were expressed differentially in wild-type Arabidopsis during infection by Pseudomonas syringae pv tomato, supporting a role for them in defense-related processes. Unexpectedly, the promoters of most Pti4-regulated genes did not have a GCC box. Chromatin immunoprecipitation experiments confirmed that Pti4 binds in vivo to promoters lacking this cis element. Potential binding sites for ERF, MYB, and GBF TFs were present in statistically significantly increased numbers in promoters regulated by Pti4. Thus, Pti4 appears to regulate gene expression directly by binding the GCC box and possibly a non-GCC box element and indirectly by either activating the expression of TF genes or interacting physically with other TFs.

Arabidopsis↗

PubMed2003

Proliferating Floral Organs (Pfo), a Lotus japonicus gene required for specifying floral meristem determinacy and organ identity, encodes an F-box protein.

To study flower development in the model legume Lotus japonicus, a population of transgenic plants containing a maize transposable element (Ac) in their genome was screened for floral mutants. One mutation named proliferating floral organs (pfo) causes plants to produce a large number of sepal-like organs instead of normal flowers. It segregates as a single recessive Mendelian locus, and causes sterility. Scanning electron microscopy revealed that pfo affects the identity, number and arrangement of floral organs. Sepal-like organs form in the first whorl, and secondary floral meristems are produced in the next whorl. These in turn produce sepal-like organs in the first whorl and floral meristems in the second whorl, and the process is reiterated. Petals and stamens are absent while carpels are either absent or reduced. The pfo phenotype was correlated with the presence of an Ac insertion yielding a 1.6-kb HindIII restriction fragment on Southern blots. Both the mutant phenotype and this Ac element are unstable. Using the transposon as a tag, the Pfo gene was isolated. Conceptual translation of Pfo predicts a protein containing an F-box, with high overall similarity to the Antirrhinum FIMBRIATA, Arabidopsis UNUSUAL FLORAL ORGANS and Pisum sativum Stamina pistilloida proteins. This suggests that Pfo may regulate floral organ identity and meristem determinacy by targeting proteins for ubiquitination.

Amino Acid Sequence↗

PubMed2003

The Arabidopsis NPR1 disease resistance protein is a novel cofactor that confers redox regulation of DNA binding activity to the basic domain/leucine zipper transcription factor TGA1.

The Arabidopsis NPR1 protein is essential for regulating salicylic acid-dependent gene expression during systemic acquired resistance. NPR1 interacts differentially with members of the TGA class of basic domain/Leu zipper transcription factors and regulates their DNA binding activity. Here, we report that although TGA1 does not interact with NPR1 in yeast two-hybrid assays, treatment with salicylic acid induces the interaction between these proteins in Arabidopsis leaves. This phenomenon is correlated with a reduction of TGA1 Cys residues. Furthermore, site-directed mutagenesis of TGA1 Cys-260 and Cys-266 enables the interaction with NPR1 in yeast and Arabidopsis. Together, these results indicate that TGA1 relies on the oxidation state of Cys residues to mediate the interaction with NPR1. An intramolecular disulfide bridge in TGA1 precludes interaction with NPR1, and NPR1 can only stimulate the DNA binding activity of the reduced form of TGA1. Unlike its animal and yeast counterparts, the DNA binding activity of TGA1 is not redox regulated; however, this property is conferred by interaction with the NPR1 cofactor.

Amino Acid Sequence↗