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Identification of maize genes associated with host plant resistance or susceptibility to Aspergillus flavus infection and aflatoxin accumulation.

Kelley RY, Williams WP, Mylroie JE, Boykin DL, Harper JW, Windham GL, Ankala A, Shan X - PLoS ONE (2012)

Bottom Line: Sixteen of the candidate genes were found to be highly expressed in Mp313E and fifteen in Va35.A gene encoding glycine-rich RNA binding protein 2 was found to be associated with the host hypersensitivity and susceptibility in Va35.A nuclear pore complex protein YUP85-like gene was found to be involved in the host resistance in Mp313E.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Molecular Biology, Entomology, and Plant Pathology, Mississippi State University, Mississippi State, Mississippi, United States of America.

ABSTRACT

Background: Aspergillus flavus infection and aflatoxin contamination of maize pose negative impacts in agriculture and health. Commercial maize hybrids are generally susceptible to this fungus. Significant levels of host plant resistance have been observed in certain maize inbred lines. This study was conducted to identify maize genes associated with host plant resistance or susceptibility to A. flavus infection and aflatoxin accumulation.

Results: Genome wide gene expression levels with or without A. flavus inoculation were compared in two resistant maize inbred lines (Mp313E and Mp04:86) in contrast to two susceptible maize inbred lines (Va35 and B73) by microarray analysis. Principal component analysis (PCA) was used to find genes contributing to the larger variances associated with the resistant or susceptible maize inbred lines. The significance levels of gene expression were determined by using SAS and LIMMA programs. Fifty candidate genes were selected and further investigated by quantitative RT-PCR (qRT-PCR) in a time-course study on Mp313E and Va35. Sixteen of the candidate genes were found to be highly expressed in Mp313E and fifteen in Va35. Out of the 31 highly expressed genes, eight were mapped to seven previously identified quantitative trait locus (QTL) regions. A gene encoding glycine-rich RNA binding protein 2 was found to be associated with the host hypersensitivity and susceptibility in Va35. A nuclear pore complex protein YUP85-like gene was found to be involved in the host resistance in Mp313E.

Conclusion: Maize genes associated with host plant resistance or susceptibility were identified by a combination of microarray analysis, qRT-PCR analysis, and QTL mapping methods. Our findings suggest that multiple mechanisms are involved in maize host plant defense systems in response to Aspergillus flavus infection and aflatoxin accumulation. These findings will be important in identification of DNA markers for breeding maize lines resistant to aflatoxin accumulation.

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Related in: MedlinePlus

Biplots showing results of the principal component analysis (PCA) on log2 expression ratios (I/U) of 13,107 expressed genes.The distribution of the gene expression values shows evident trends represented by the vectors associated with resistant and susceptible maize inbred lines. Genes expressed toward the resistance trait were located close to the vectors for Mp313E and Mp04∶86. Genes contributing to susceptibility were located close to the vectors for Va35 and B73. 1A. Biplot of principal component analysis (PCA) on 13,107 expressed gene probes. 1B. Biplot of principal component analysis (PCA) on a subset of 500 expressed gene probes. (The arrows represent vectors. The direction and length represent the larger variance of the expression values).
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pone-0036892-g001: Biplots showing results of the principal component analysis (PCA) on log2 expression ratios (I/U) of 13,107 expressed genes.The distribution of the gene expression values shows evident trends represented by the vectors associated with resistant and susceptible maize inbred lines. Genes expressed toward the resistance trait were located close to the vectors for Mp313E and Mp04∶86. Genes contributing to susceptibility were located close to the vectors for Va35 and B73. 1A. Biplot of principal component analysis (PCA) on 13,107 expressed gene probes. 1B. Biplot of principal component analysis (PCA) on a subset of 500 expressed gene probes. (The arrows represent vectors. The direction and length represent the larger variance of the expression values).

Mentions: The maize oligonucleotide arrays (from NSF Maize Oligonucleotide Array Project) used in this experiment contained 57,452 maize gene probes. In our study, each microarray slide was hybridized with two samples (dual channel hybridization) that were the inoculated and uninoculated samples from the same genotype and the same plot. In such an array design we first evaluated the resistance and susceptibility responses in each genotype and then compared the differentially expressed genes between groups of two resistant and two susceptible maize inbred lines. We performed multiple analyses on the microarray data using different statistical algorithms to reveal the host plant specific responses to Aspergillus flavus infection and aflatoxin reduction. We first analyzed microarray data with SAS Version 9.1.3 [18] using the mixed model for a split plot design with four maize inbred lines as main unit and two treatments [inoculated (I) vs. uninoculated (U)] as subunit. Log transformed median expression values were used to obtain estimates of gene expression ratios (I/U) for each genotype. Then the log2 values of the expression ratios (I/U) of 13,107 expressed genes from all genotypes were analyzed by principal component analysis (PCA). Figure 1A and 1B are biplots showing the distribution of the log2 gene expression ratios (I/U) on a projected principal plane. The data appeared to be cloudy, however, we found that the vectors representing the larger variances associated with the two resistant maize inbred lines (Mp313E and Mp04∶86) were clustered together. Likewise, the vectors representing the larger variances associated with the two susceptible maize inbred lines (Va35 and B73) were clustered together. That means despite the differences in the genomes of these genotypes, trends of gene expression associated with host plant resistance or susceptibility were evident. For example, Mp04∶86 showed a distinguished expression pattern compared to its susceptible parent Va35 in response to the fungal infection (Figure 1A and 1B). The PCA analysis has revealed a separation of genes between the resistant and susceptible groups in response to Aspergillus flavus colonization. Genes that expressed toward the larger variances were considered as associated with the corresponding traits. Genes expressed toward the resistance trait located close to the vectors for Mp313E and Mp04∶86. Genes for susceptibility located close to the vectors for Va35 and B73 (Figure 1A and 1B). By this method, we grouped genes for possible candidates contributing to either the host resistance or the host susceptibility.


Identification of maize genes associated with host plant resistance or susceptibility to Aspergillus flavus infection and aflatoxin accumulation.

Kelley RY, Williams WP, Mylroie JE, Boykin DL, Harper JW, Windham GL, Ankala A, Shan X - PLoS ONE (2012)

Biplots showing results of the principal component analysis (PCA) on log2 expression ratios (I/U) of 13,107 expressed genes.The distribution of the gene expression values shows evident trends represented by the vectors associated with resistant and susceptible maize inbred lines. Genes expressed toward the resistance trait were located close to the vectors for Mp313E and Mp04∶86. Genes contributing to susceptibility were located close to the vectors for Va35 and B73. 1A. Biplot of principal component analysis (PCA) on 13,107 expressed gene probes. 1B. Biplot of principal component analysis (PCA) on a subset of 500 expressed gene probes. (The arrows represent vectors. The direction and length represent the larger variance of the expression values).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3351445&req=5

pone-0036892-g001: Biplots showing results of the principal component analysis (PCA) on log2 expression ratios (I/U) of 13,107 expressed genes.The distribution of the gene expression values shows evident trends represented by the vectors associated with resistant and susceptible maize inbred lines. Genes expressed toward the resistance trait were located close to the vectors for Mp313E and Mp04∶86. Genes contributing to susceptibility were located close to the vectors for Va35 and B73. 1A. Biplot of principal component analysis (PCA) on 13,107 expressed gene probes. 1B. Biplot of principal component analysis (PCA) on a subset of 500 expressed gene probes. (The arrows represent vectors. The direction and length represent the larger variance of the expression values).
Mentions: The maize oligonucleotide arrays (from NSF Maize Oligonucleotide Array Project) used in this experiment contained 57,452 maize gene probes. In our study, each microarray slide was hybridized with two samples (dual channel hybridization) that were the inoculated and uninoculated samples from the same genotype and the same plot. In such an array design we first evaluated the resistance and susceptibility responses in each genotype and then compared the differentially expressed genes between groups of two resistant and two susceptible maize inbred lines. We performed multiple analyses on the microarray data using different statistical algorithms to reveal the host plant specific responses to Aspergillus flavus infection and aflatoxin reduction. We first analyzed microarray data with SAS Version 9.1.3 [18] using the mixed model for a split plot design with four maize inbred lines as main unit and two treatments [inoculated (I) vs. uninoculated (U)] as subunit. Log transformed median expression values were used to obtain estimates of gene expression ratios (I/U) for each genotype. Then the log2 values of the expression ratios (I/U) of 13,107 expressed genes from all genotypes were analyzed by principal component analysis (PCA). Figure 1A and 1B are biplots showing the distribution of the log2 gene expression ratios (I/U) on a projected principal plane. The data appeared to be cloudy, however, we found that the vectors representing the larger variances associated with the two resistant maize inbred lines (Mp313E and Mp04∶86) were clustered together. Likewise, the vectors representing the larger variances associated with the two susceptible maize inbred lines (Va35 and B73) were clustered together. That means despite the differences in the genomes of these genotypes, trends of gene expression associated with host plant resistance or susceptibility were evident. For example, Mp04∶86 showed a distinguished expression pattern compared to its susceptible parent Va35 in response to the fungal infection (Figure 1A and 1B). The PCA analysis has revealed a separation of genes between the resistant and susceptible groups in response to Aspergillus flavus colonization. Genes that expressed toward the larger variances were considered as associated with the corresponding traits. Genes expressed toward the resistance trait located close to the vectors for Mp313E and Mp04∶86. Genes for susceptibility located close to the vectors for Va35 and B73 (Figure 1A and 1B). By this method, we grouped genes for possible candidates contributing to either the host resistance or the host susceptibility.

Bottom Line: Sixteen of the candidate genes were found to be highly expressed in Mp313E and fifteen in Va35.A gene encoding glycine-rich RNA binding protein 2 was found to be associated with the host hypersensitivity and susceptibility in Va35.A nuclear pore complex protein YUP85-like gene was found to be involved in the host resistance in Mp313E.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Molecular Biology, Entomology, and Plant Pathology, Mississippi State University, Mississippi State, Mississippi, United States of America.

ABSTRACT

Background: Aspergillus flavus infection and aflatoxin contamination of maize pose negative impacts in agriculture and health. Commercial maize hybrids are generally susceptible to this fungus. Significant levels of host plant resistance have been observed in certain maize inbred lines. This study was conducted to identify maize genes associated with host plant resistance or susceptibility to A. flavus infection and aflatoxin accumulation.

Results: Genome wide gene expression levels with or without A. flavus inoculation were compared in two resistant maize inbred lines (Mp313E and Mp04:86) in contrast to two susceptible maize inbred lines (Va35 and B73) by microarray analysis. Principal component analysis (PCA) was used to find genes contributing to the larger variances associated with the resistant or susceptible maize inbred lines. The significance levels of gene expression were determined by using SAS and LIMMA programs. Fifty candidate genes were selected and further investigated by quantitative RT-PCR (qRT-PCR) in a time-course study on Mp313E and Va35. Sixteen of the candidate genes were found to be highly expressed in Mp313E and fifteen in Va35. Out of the 31 highly expressed genes, eight were mapped to seven previously identified quantitative trait locus (QTL) regions. A gene encoding glycine-rich RNA binding protein 2 was found to be associated with the host hypersensitivity and susceptibility in Va35. A nuclear pore complex protein YUP85-like gene was found to be involved in the host resistance in Mp313E.

Conclusion: Maize genes associated with host plant resistance or susceptibility were identified by a combination of microarray analysis, qRT-PCR analysis, and QTL mapping methods. Our findings suggest that multiple mechanisms are involved in maize host plant defense systems in response to Aspergillus flavus infection and aflatoxin accumulation. These findings will be important in identification of DNA markers for breeding maize lines resistant to aflatoxin accumulation.

Show MeSH
Related in: MedlinePlus