Functional analysis of the theobroma cacao NPR1 gene in arabidopsis

<p>Abstract</p> <p>Background</p> <p>The <it>Arabidopsis thaliana NPR1 </it>gene encodes a transcription coactivator (NPR1) that plays a major role in the mechanisms regulating plant defense response. After pathogen infection and in response to salicylic acid (SA) accumulation, NPR1 translocates from the cytoplasm into the nucleus where it interacts with other transcription factors resulting in increased expression of over 2000 plant defense genes contributing to a pathogen resistance response.</p> <p>Results</p> <p>A putative <it>Theobroma cacao NPR1 </it>cDNA was isolated by RT-PCR using degenerate primers based on homologous sequences from <it>Brassica</it>, <it>Arabidopsis </it>and <it>Carica papaya</it>. The cDNA was used to isolate a genomic clone from <it>Theobroma cacao </it>containing a putative <it>TcNPR1 </it>gene. DNA sequencing revealed the presence of a 4.5 kb coding region containing three introns and encoding a polypeptide of 591 amino acids. The predicted TcNPR1 protein shares 55% identity and 78% similarity to <it>Arabidopsis </it>NPR1, and contains each of the highly conserved functional domains indicative of this class of transcription factors (BTB/POZ and ankyrin repeat protein-protein interaction domains and a nuclear localization sequence (NLS)). To functionally define the <it>TcNPR1 </it>gene, we transferred <it>TcNPR1 </it>into an <it>Arabidopsis npr1 </it>mutant that is highly susceptible to infection by the plant pathogen <it>Pseudomonas syringae </it>pv. tomato DC3000. Driven by the constitutive CaMV35S promoter, the cacao <it>TcNPR1 </it>gene partially complemented the <it>npr1 </it>mutation in transgenic <it>Arabidopsis </it>plants, resulting in 100 fold less bacterial growth in a leaf infection assay. Upon induction with SA, <it>TcNPR1 </it>was shown to translocate into the nucleus of leaf and root cells in a manner identical to <it>Arabidopsis </it>NPR1. Cacao NPR1 was also capable of participating in SA-JA signaling crosstalk, as evidenced by the suppression of JA responsive gene expression in <it>TcNPR1 </it>overexpressing transgenic plants.</p> <p>Conclusion</p> <p>Our data indicate that the <it>TcNPR1 </it>is a functional ortholog of <it>Arabidopsis NPR1</it>, and is likely to play a major role in defense response in cacao. This fundamental knowledge can contribute to breeding of disease resistant cacao varieties through the application of molecular markers or the use of transgenic strategies.</p

Erratum to: Theobroma cacao L. pathogenesis-related gene tandem array members show diverse expression dynamics in response to pathogen colonization

The original version of the manuscript [1] contained an incorrectly named Criollo gene ID on chromosome 1 in the first sentence, under the subheading “Organization of PR gene families into tandem arrays”. The second gene on chromosome 1, Tc##_g######, should therefore be Tc01_g000020.The original version of the manuscript [1] contained an incorrectly named Criollo gene ID on chromosome 1 in the first sentence, under the subheading “Organization of PR gene families into tandem arrays”. The second gene on chromosome 1, Tc##_g######, should therefore be Tc01_g000020

Population Structure and Molecular Characterization of Nigerian Field Genebank Collections of Cacao, Theobroma cacao L.

AbstractInadequate knowledge of the population structure and diversity present often hamper the efficient use of germplasm collections. Using a high through-put system, twelve microsatellite loci were used to analyze genetic diversity and population structure in a national field genebank repository of 243 cacao accessions grouped into 11 populations based on their known sources. Based on multi-locus profiles, the Bayesian method was used for individual assignment to verify membership in each population, determine mislabeling and ancestry of some important accessions used in breeding program. A total of 218 alleles was revealed with a mean number of 18.2 alleles per locus. Gene diversity (He= 0.70) and allelic richness (4.34 alleles per locus) were highest in the F1 hybrid population. Differential mating system was suggested as responsible for the observed deficit and excess of heterozygotes observed among the populations. Analysis of molecular variance showed that within-population variance accounted for 63.0% of the total variance while the rest 37% was accounted for by the among-population variance. Cluster dendrogram based on UPGMA revealed two main subsets. The first group was made up of the Amelonado/Trinitario ancestry and the other of Nanay/Parinari ancestry. We found that Nanay and Parinari populations were the major source of Upper Amazon genes utilized while a large proportion of genetic diversity in the field genebank remained under-utilized in development of improved cultivars released to farmers in Nigeria. This study showed that the presence of alleles of the Upper Amazon Forasteros (Nanay, Parinari and Iquitos Mixed Calabacillo) genetic materials in the locally available accessions predated the formal large scale introduction of Upper Amazon materials in 1944. This is the first report of population structure of field genebank collections of cacao in Nigeria since more than seven decades of formal cacao breeding research

Towards the understanding of the cocoa transcriptome: Production and analysis of an exhaustive dataset of ESTs of Theobroma cacao L. generated from various tissues and under various conditions

Theobroma cacao L., is a tree originated from the tropical rainforest of South America. It is one of the major cash crops for many tropical countries. T. cacao is mainly produced on smallholdings, providing resources for 14 million farmers. Disease resistance and T. cacao quality improvement are two important challenges for all actors of cocoa and chocolate production. T. cacao is seriously affected by pests and fungal diseases, responsible for more than 40% yield losses and quality improvement, nutritional and organoleptic, is also important for consumers. An international collaboration was formed to develop an EST genomic resource database for cacao. Fifty-six cDNA libraries were constructed from different organs, different genotypes and different environmental conditions. A total of 149,650 valid EST sequences were generated corresponding to 48,594 unigenes, 12,692 contigs and 35,902 singletons. A total of 29,849 unigenes shared significant homology with public sequences from other species. Gene Ontology (GO) annotation was applied to distribute the ESTs among the main GO categories. A specific information system (ESTtik) was constructed to process, store and manage this EST collection allowing the user to query a database. To check the representativeness of our EST collection, we looked for the genes known to be involved in two different metabolic pathways extensively studied in other plant species and important for T. cacao qualities: the flavonoid and the terpene pathways. Most of the enzymes described in other crops for these two metabolic pathways were found in our EST collection. A large collection of new genetic markers was provided by this ESTs collection. This EST collection displays a good representation of the T. cacao transcriptome, suitable for analysis of biochemical pathways based on oligonucleotide microarrays derived from these ESTs. It will provide numerous genetic markers that will allow the construction of a high density gene map of T. cacao. This EST collection represents a unique and important molecular resource for T. cacao study and improvement, facilitating the discovery of candidate genes for important T. cacao trait variation. (Résumé d'auteur

Pervasive effects of a dominant foliar endophytic fungus on host genetic and phenotypic expression in a tropical tree

It is increasingly recognized that macro-organisms (corals, insects, plants, vertebrates) consist of both host tissues and multiple microbial symbionts that play essential roles in their host’s ecological and evolutionary success. Consequently, identifying benefits and costs of symbioses, as well as mechanisms underlying them are research priorities. All plants surveyed under natural conditions harbor foliar endophytic fungi (FEF) in their leaf tissues, often at high densities. Despite producing no visible effects on their hosts, experiments have nonetheless shown that FEF reduce pathogen and herbivore damage. Here, combining results from three genomic, and two physiological experiments, we demonstrate pervasive genetic and phenotypic effects of the apparently asymptomatic endophytes on their hosts. Specifically, inoculation of endophyte-free (E−) Theobroma cacao leaves with Colletotrichum tropicale (E+), the dominant FEF species in healthy T. cacao, induces consistent changes in the expression of hundreds of host genes, including many with known defensive functions. Further, E+ plants exhibited increased lignin and cellulose content, reduced maximum rates of photosynthesis (Amax), and enrichment of nitrogen-15 and carbon-13 isotopes. These phenotypic changes observed in E+ plants correspond to changes in expression of specific functional genes in related pathways. Moreover, a cacao gene (Tc00g04254) highly up-regulated by C. tropicale also confers resistance to pathogen damage in the absence of endophytes or their products in host tissues. Thus, the benefits of increased pathogen resistance in E+ plants are derived in part from up-regulation of intrinsic host defense responses, and appear to be offset by potential costs including reduced photosynthesis, altered host nitrogen metabolism, and endophyte heterotrophy of host tissues. Similar effects are likely in most plant-endophyte interactions, and should be recognized in the design and interpretation of genetic and phenotypic studies of plantsIt is increasingly recognized that macro-organisms (corals, insects, plants, vertebrates) consist of both host tissues and multiple microbial symbionts that play essential roles in their host’s ecological and evolutionary success. Consequently, identifying benefits and costs of symbioses, as well as mechanisms underlying them are research priorities. All plants surveyed under natural conditions harbor foliar endophytic fungi (FEF) in their leaf tissues, often at high densities. Despite producing no visible effects on their hosts, experiments have nonetheless shown that FEF reduce pathogen and herbivore damage. Here, combining results from three genomic, and two physiological experiments, we demonstrate pervasive genetic and phenotypic effects of the apparently asymptomatic endophytes on their hosts. Specifically, inoculation of endophyte-free (E−) Theobroma cacao leaves with Colletotrichum tropicale (E+), the dominant FEF species in healthy T. cacao, induces consistent changes in the expression of hundreds of host genes, including many with known defensive functions. Further, E+ plants exhibited increased lignin and cellulose content, reduced maximum rates of photosynthesis (Amax), and enrichment of nitrogen-15 and carbon-13 isotopes. These phenotypic changes observed in E+ plants correspond to changes in expression of specific functional genes in related pathways. Moreover, a cacao gene (Tc00g04254) highly up-regulated by C. tropicale also confers resistance to pathogen damage in the absence of endophytes or their products in host tissues. Thus, the benefits of increased pathogen resistance in E+ plants are derived in part from up-regulation of intrinsic host defense responses, and appear to be offset by potential costs including reduced photosynthesis, altered host nitrogen metabolism, and endophyte heterotrophy of host tissues. Similar effects are likely in most plant-endophyte interactions, and should be recognized in the design and interpretation of genetic and phenotypic studies of plant

A genome survey of Moniliophthora perniciosa gives new insights into Witches' Broom Disease of cacao

<p>Abstract</p> <p>Background</p> <p>The basidiomycete fungus <it>Moniliophthora perniciosa </it>is the causal agent of Witches' Broom Disease (WBD) in cacao (<it>Theobroma cacao</it>). It is a hemibiotrophic pathogen that colonizes the apoplast of cacao's meristematic tissues as a biotrophic pathogen, switching to a saprotrophic lifestyle during later stages of infection. <it>M. perniciosa</it>, together with the related species <it>M. roreri</it>, are pathogens of aerial parts of the plant, an uncommon characteristic in the order Agaricales. A genome survey (1.9× coverage) of <it>M. perniciosa </it>was analyzed to evaluate the overall gene content of this phytopathogen.</p> <p>Results</p> <p>Genes encoding proteins involved in retrotransposition, reactive oxygen species (ROS) resistance, drug efflux transport and cell wall degradation were identified. The great number of genes encoding cytochrome P450 monooxygenases (1.15% of gene models) indicates that <it>M. perniciosa </it>has a great potential for detoxification, production of toxins and hormones; which may confer a high adaptive ability to the fungus. We have also discovered new genes encoding putative secreted polypeptides rich in cysteine, as well as genes related to methylotrophy and plant hormone biosynthesis (gibberellin and auxin). Analysis of gene families indicated that <it>M. perniciosa </it>have similar amounts of carboxylesterases and repertoires of plant cell wall degrading enzymes as other hemibiotrophic fungi. In addition, an approach for normalization of gene family data using incomplete genome data was developed and applied in <it>M. perniciosa </it>genome survey.</p> <p>Conclusion</p> <p>This genome survey gives an overview of the <it>M. perniciosa </it>genome, and reveals that a significant portion is involved in stress adaptation and plant necrosis, two necessary characteristics for a hemibiotrophic fungus to fulfill its infection cycle. Our analysis provides new evidence revealing potential adaptive traits that may play major roles in the mechanisms of pathogenicity in the <it>M. perniciosa</it>/cacao pathosystem.</p