Project description:Stem rust of wheat is a deleterious fungal disease across the globe causing severe yield losses. Although, many stem rust resistance genes (Sr) are being used in wheat breeding programs, new emerging stem rust pathotypes are a challenge to important Sr genes. In recent years, multiple studies on leaf and yellow rust molecular mechanism have been done, however, for stem rust such studies are lacking. Current study investigated stem rust induced response in the susceptible wheat genotype C306 and its Near Isogenic Line (NIL) for Sr24 gene, HW2004, using microarray analysis to understand the transcriptomic differences at different stages of infection. Results showed that HW2004 has higher basal levels of several important genes involved in pathogen detection, defence, and display early activation of multiple defence mechanisms. Further Gene Ontology (GO) and pathway analysis identified important genes responsible for pathogen detection, downstream signalling cascades and transcription factors (TFs) involved in activation and mediation of defence responses. Results suggest that generation of Reactive Oxygen Species (ROS), cytoskeletal rearrangement, activation of multiple hydrolases, and lipid metabolism mediated biosynthesis of certain secondary metabolites are collectively involved in Sr24-mediated defence in HW2004, in response to stem rust infection. Novel and unannotated, but highly responsive genes were also identified, which may also contribute towards resistance phenotype. Furthermore, certain DEGs also mapped close to the Sr24-linked marker on Thinopyrum elongatum translocated fragment on wheat 3E chromosome, which advocate further investigations for better insights of the Sr24-mediated stem rust resistance.
Project description:Aim:To characterise a recently discovered stem rust resistance locus on wheat chromosome 7AL. Transcriptome analysis by RNA-sequencing, in association with microscopic observations, was used to compare responses to the Puccinia graminis f. sp. tritici pathogen of the susceptible line Columbus, and two independent backcrossed resistant lines containing the locus, Columbus-NS765 and Columbus-NS766. Results: Microscopic observations of infected leaves revealed that the resistance conferred by the 7AL resistance locus was initiated by two days post-inoculation, upon the entry of the stem rust fungus into the plant through the stoma. Death of guard and epidermal cells adjacent to the fungal points of entry was observed to be clearly more frequent in resistant lines than in the susceptible genotype, suggesting that the resistance response is similar in all genotypes, but enhanced in the resistant lines. Transcriptomic analysis, combined with assignment of genes to wheat chromosomes, revealed a disporportionately high number of differentially expressed genes were located on chromosomes 7AL and 6A. A number of genes annotated as cysteine-rich receptor-like kinases were located on chromosome 7AL. Closer investigation indicated that the encoded proteins were in fact putative receptor-like cytoplasmic kinases (RLCKs). One of the putative RLCK genes contained a SNP marker previously shown to co-segregate with the 7AL resistance locus. The large number of differentially expressed genes on chromosome 6A indicated the presence of a large introgression on this chromosome that co-segregated with stem rust resistance in the two independent resistant lines, but its role in the resistance response is currently unclear. Conclusions: This study represents the first transcriptome analysis of responses to stem rust in wheat, and the first investigation of the resistance conferred by the newly-discovered wheat 7AL stem rust resistance locus. Microscopy showed the resistance response was associated with pre-haustorial cell death. Results of the RNA-seq, which has the resolution to discriminate between homeologous wheat genes, along with assignment of differentially expressed genes to wheat chromosomes, suggested putative receptor-like cytoplasmic kinases linked to the 7AL locus as candidate resistance genes for further investigation.
Project description:The wheat gene Lr1 imparts resistance to certain races of the leaf rust pathogen Puccinia triticina. However, the biological basis underlying the complex resistance mechanisms remains largely unknown. Gene expression profiling analyses using the Affymetrix GeneChipM-BM-. Wheat Genome Array were performed with six independent Lr1 transgenic lines sampled before inoculation (0h) and at 6 and 24 h after inoculation with the avirulent P. triticina race CCDS. For the 6 and 24 h time points, 3M-BM- 384 probe sets corresponding to 2M-BM- 978 transcripts were differentially expressed as compared to the plants before inoculation. Genes involved in defence signalling were triggered as early as 6 h post inoculation (HPI) and included COI1, the primary component of jasmonic acid signalling. By 24 HPI, a shift in plant cellular metabolism, namely carbon conservation, was notable. The enzymes of glycolysis and gluconeogenesis were down-regulated, however, enzymes linked to alternative carbon sequestration processes such as the glyoxylate cycle were up-regulated. Also at 24 HPI, various secondary signalling molecules belonging to the various hormone signalling pathways were triggered. These microarray data suggest that the Lr1 mediated leaf rust resistance is achieved by early onset of defence signalling followed by co-ordinated interplay of metabolism alteration and elicitation of defence responses. We used microarrays to detail the early gene expression events underlying Lr1-mediated resistance to Puccinia triticina pathogen in wheat. Three biological replicates with six independent Lr1 transgenic lines were used in this study to eliminate the possibility of false discovery of genes due to copy number variation or the position of insertion of the transgene Lr1. Early time points of 6 and 24 HPI were compared to 0 h (prior to inoculation) to capture the early signalling events involved in Lr1-mediated response to rust infection.
Project description:Oligoarray expression profiling was carried out in poplar leaves upon infection with rust in order to identify genes expressed during tree defense response. For this purpose, we inoculated detached leaves of the interamerican hybrid poplar Populus trichocarpa x Populus deltoides 'Beaupré' grown in greenhouse either with spores of avirulent strain 93ID6 (incompatible interaction I48) or spores of virulent strain 98AG31 (compatible interaction C48) of the pathogenic rust fungus Melampsora larici-populina. Besides, we mock-inoculated 'Beaupré' leaves with water (control condition, T48). Detached leaves were maintained in vitro in controled conditions to allow fungal infection and colonization of plant tissue. Leaves were sampled 48 hours post-inoculation after that the fungus attempt to penetrate plant cells in mesophyll. Keywords: Plant tissue infection, Plant defense response, Oligonucleotide array
Project description:cDNA macroarray expression profiling was carried out in poplar leaves upon infection with rust in order to identify genes expressed during tree defense response. For this purpose, we inoculated detached leaves of the interamerican hybrid poplar Populus trichocarpa x Populus deltoides 'Beaupré' grown in greenhouse either with spores of avirulent strain 93ID6 (incompatible interaction, I) or spores of virulent strain 98AG31 (compatible interaction, C) of the pathogenic rust fungus Melampsora larici-populina. Besides, we mock-inoculated 'Beaupré' leaves with water (control condition, T). Detached leaves were maintained in vitro in controled conditions to allow fungal infection and colonization of plant tissue. Leaves were sampled at 12, 24 and 48 hours post-inoculation (hpi) in a time-course experiment before (12 hpi) and after (24 and 48 hpi) that the fungus attempt to penetrate plant cells in mesophyll. Keywords: Plant tissue infection, Plant defense response, cDNA macroarray
Project description:Microarray expression profiling was carried out in poplar leaves upon infection with rust in order to identify genes expressed during tree defense response. For this purpose, we inoculated detached leaves of the interamerican hybrid poplar Populus trichocarpa x Populus deltoides 'Beaupré' grown in greenhouse with spores of avirulent strain 93ID6 of the pathogenic rust fungus Melampsora larici-populina (incompatible interaction, I48). Besides, we mock-inoculated 'Beaupré' leaves with water (control condition, T48). Detached leaves were maintained in vitro in controled conditions to allow fungal infection and colonization of plant tissue. Leaves were sampled 48 hours post-inoculation after that the fungus attempt to penetrate plant cells in mesophyll. Competitive hybridization between transcripts of incompatible interaction (I48) and control condition (T48) was done on Populus PICME 28K cDNA microarray. Keywords: Time-course infection of plant tissue, defense response, cDNA microarray
Project description:The wheat gene Lr1 imparts resistance to certain races of the leaf rust pathogen Puccinia triticina. However, the biological basis underlying the complex resistance mechanisms remains largely unknown. Gene expression profiling analyses using the Affymetrix GeneChip® Wheat Genome Array were performed with six independent Lr1 transgenic lines sampled before inoculation (0h) and at 6 and 24 h after inoculation with the avirulent P. triticina race CCDS. For the 6 and 24 h time points, 3 384 probe sets corresponding to 2 978 transcripts were differentially expressed as compared to the plants before inoculation. Genes involved in defence signalling were triggered as early as 6 h post inoculation (HPI) and included COI1, the primary component of jasmonic acid signalling. By 24 HPI, a shift in plant cellular metabolism, namely carbon conservation, was notable. The enzymes of glycolysis and gluconeogenesis were down-regulated, however, enzymes linked to alternative carbon sequestration processes such as the glyoxylate cycle were up-regulated. Also at 24 HPI, various secondary signalling molecules belonging to the various hormone signalling pathways were triggered. These microarray data suggest that the Lr1 mediated leaf rust resistance is achieved by early onset of defence signalling followed by co-ordinated interplay of metabolism alteration and elicitation of defence responses. We used microarrays to detail the early gene expression events underlying Lr1-mediated resistance to Puccinia triticina pathogen in wheat.