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Global expression analysis of nucleotide binding site-leucine rich repeat-encoding and related genes in Arabidopsis.

Tan X, Meyers BC, Kozik A, West MA, Morgante M, St Clair DA, Bent AF, Michelmore RW - BMC Plant Biol. (2007)

Bottom Line: Evidence for alternative splicing was found for at least 12 genes, 11 of which encode TIR-NBS-LRR proteins.Most were present at low levels and exhibited tissue-specific expression patterns.Expression data are consistent with most Arabidopsis NBS-LRR-encoding and related genes functioning in plant defense responses but do not preclude other biological roles.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Genome Center, University of California, Davis, California 95616, USA. xpitan@ucdavis.edu

ABSTRACT

Background: Nucleotide binding site-leucine rich repeat (NBS-LRR)-encoding genes comprise the largest class of plant disease resistance genes. The 149 NBS-LRR-encoding genes and the 58 related genes that do not encode LRRs represent approximately 0.8% of all ORFs so far annotated in Arabidopsis ecotype Col-0. Despite their prevalence in the genome and functional importance, there was little information regarding expression of these genes.

Results: We analyzed the expression patterns of approximately 170 NBS-LRR-encoding and related genes in Arabidopsis Col-0 using multiple analytical approaches: expressed sequenced tag (EST) representation, massively parallel signature sequencing (MPSS), microarray analysis, rapid amplification of cDNA ends (RACE) PCR, and gene trap lines. Most of these genes were expressed at low levels with a variety of tissue specificities. Expression was detected by at least one approach for all but 10 of these genes. The expression of some but not the majority of NBS-LRR-encoding and related genes was affected by salicylic acid (SA) treatment; the response to SA varied among different accessions. An analysis of previously published microarray data indicated that ten NBS-LRR-encoding and related genes exhibited increased expression in wild-type Landsberg erecta (Ler) after flagellin treatment. Several of these ten genes also showed altered expression after SA treatment, consistent with the regulation of R gene expression during defense responses and overlap between the basal defense response and salicylic acid signaling pathways. Enhancer trap analysis indicated that neither jasmonic acid nor benzothiadiazole (BTH), a salicylic acid analog, induced detectable expression of the five NBS-LRR-encoding genes and one TIR-NBS-encoding gene tested; however, BTH did induce detectable expression of the other TIR-NBS-encoding gene analyzed. Evidence for alternative mRNA polyadenylation sites was observed for many of the tested genes. Evidence for alternative splicing was found for at least 12 genes, 11 of which encode TIR-NBS-LRR proteins. There was no obvious correlation between expression pattern, phylogenetic relationship or genomic location of the NBS-LRR-encoding and related genes studied.

Conclusion: Transcripts of many NBS-LRR-encoding and related genes were defined. Most were present at low levels and exhibited tissue-specific expression patterns. Expression data are consistent with most Arabidopsis NBS-LRR-encoding and related genes functioning in plant defense responses but do not preclude other biological roles.

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GUS quantitative assays of enhancer trap lines with insertions into two NBS-LRR-encoding and one related gene. GUS expression levels of each enhancer trap line untreated or five days after application of JA or BTH. Treatments with ethanol (EtOH) or BTH carrier (carrier) were used as controls. Average GUS expression levels and standard deviations were based on three biological replicates. The background levels of GUS activity in untreated Col-0 plants were below 20 pmol 4-MU/μg protein/min.
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Figure 4: GUS quantitative assays of enhancer trap lines with insertions into two NBS-LRR-encoding and one related gene. GUS expression levels of each enhancer trap line untreated or five days after application of JA or BTH. Treatments with ethanol (EtOH) or BTH carrier (carrier) were used as controls. Average GUS expression levels and standard deviations were based on three biological replicates. The background levels of GUS activity in untreated Col-0 plants were below 20 pmol 4-MU/μg protein/min.

Mentions: The expression pattern of the corresponding NBS-LRR-encoding and related gene in each confirmed enhancer trap line was analyzed using GUS staining and quantitative GUS assays of seedlings and five-week old plants. No GUS activity was detected histochemically in whole seedling, leaves, roots, flowers and stems of any of the five NBS-LRR and one TIR-NBS enhancer trap lines. This indicated that the low levels of expression detected in the microarray and MPSS experiments for these genes were below the detection threshold of GUS assays in these enhancer trap lines. We found no evidence that any specific cell types exhibited localized, high levels of expression as has been observed for insertions into some other types of genes [81,87,88]. In the enhancer trap line with an insertion into a TIR-NBS-encoding gene, At1g72910, there was very faint blue color throughout the leaf after staining for GUS activity. This low level of GUS activity was confirmed using a GUS quantitative assay (Figure 4). At1g72910 was one of the more highly expressed genes detected in the MPSS libraries and microarray data in Col-0 leaves (Table 5). These data indicate the threshold necessary for histochemical detection of gene activity in gene trap lines.


Global expression analysis of nucleotide binding site-leucine rich repeat-encoding and related genes in Arabidopsis.

Tan X, Meyers BC, Kozik A, West MA, Morgante M, St Clair DA, Bent AF, Michelmore RW - BMC Plant Biol. (2007)

GUS quantitative assays of enhancer trap lines with insertions into two NBS-LRR-encoding and one related gene. GUS expression levels of each enhancer trap line untreated or five days after application of JA or BTH. Treatments with ethanol (EtOH) or BTH carrier (carrier) were used as controls. Average GUS expression levels and standard deviations were based on three biological replicates. The background levels of GUS activity in untreated Col-0 plants were below 20 pmol 4-MU/μg protein/min.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2175511&req=5

Figure 4: GUS quantitative assays of enhancer trap lines with insertions into two NBS-LRR-encoding and one related gene. GUS expression levels of each enhancer trap line untreated or five days after application of JA or BTH. Treatments with ethanol (EtOH) or BTH carrier (carrier) were used as controls. Average GUS expression levels and standard deviations were based on three biological replicates. The background levels of GUS activity in untreated Col-0 plants were below 20 pmol 4-MU/μg protein/min.
Mentions: The expression pattern of the corresponding NBS-LRR-encoding and related gene in each confirmed enhancer trap line was analyzed using GUS staining and quantitative GUS assays of seedlings and five-week old plants. No GUS activity was detected histochemically in whole seedling, leaves, roots, flowers and stems of any of the five NBS-LRR and one TIR-NBS enhancer trap lines. This indicated that the low levels of expression detected in the microarray and MPSS experiments for these genes were below the detection threshold of GUS assays in these enhancer trap lines. We found no evidence that any specific cell types exhibited localized, high levels of expression as has been observed for insertions into some other types of genes [81,87,88]. In the enhancer trap line with an insertion into a TIR-NBS-encoding gene, At1g72910, there was very faint blue color throughout the leaf after staining for GUS activity. This low level of GUS activity was confirmed using a GUS quantitative assay (Figure 4). At1g72910 was one of the more highly expressed genes detected in the MPSS libraries and microarray data in Col-0 leaves (Table 5). These data indicate the threshold necessary for histochemical detection of gene activity in gene trap lines.

Bottom Line: Evidence for alternative splicing was found for at least 12 genes, 11 of which encode TIR-NBS-LRR proteins.Most were present at low levels and exhibited tissue-specific expression patterns.Expression data are consistent with most Arabidopsis NBS-LRR-encoding and related genes functioning in plant defense responses but do not preclude other biological roles.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Genome Center, University of California, Davis, California 95616, USA. xpitan@ucdavis.edu

ABSTRACT

Background: Nucleotide binding site-leucine rich repeat (NBS-LRR)-encoding genes comprise the largest class of plant disease resistance genes. The 149 NBS-LRR-encoding genes and the 58 related genes that do not encode LRRs represent approximately 0.8% of all ORFs so far annotated in Arabidopsis ecotype Col-0. Despite their prevalence in the genome and functional importance, there was little information regarding expression of these genes.

Results: We analyzed the expression patterns of approximately 170 NBS-LRR-encoding and related genes in Arabidopsis Col-0 using multiple analytical approaches: expressed sequenced tag (EST) representation, massively parallel signature sequencing (MPSS), microarray analysis, rapid amplification of cDNA ends (RACE) PCR, and gene trap lines. Most of these genes were expressed at low levels with a variety of tissue specificities. Expression was detected by at least one approach for all but 10 of these genes. The expression of some but not the majority of NBS-LRR-encoding and related genes was affected by salicylic acid (SA) treatment; the response to SA varied among different accessions. An analysis of previously published microarray data indicated that ten NBS-LRR-encoding and related genes exhibited increased expression in wild-type Landsberg erecta (Ler) after flagellin treatment. Several of these ten genes also showed altered expression after SA treatment, consistent with the regulation of R gene expression during defense responses and overlap between the basal defense response and salicylic acid signaling pathways. Enhancer trap analysis indicated that neither jasmonic acid nor benzothiadiazole (BTH), a salicylic acid analog, induced detectable expression of the five NBS-LRR-encoding genes and one TIR-NBS-encoding gene tested; however, BTH did induce detectable expression of the other TIR-NBS-encoding gene analyzed. Evidence for alternative mRNA polyadenylation sites was observed for many of the tested genes. Evidence for alternative splicing was found for at least 12 genes, 11 of which encode TIR-NBS-LRR proteins. There was no obvious correlation between expression pattern, phylogenetic relationship or genomic location of the NBS-LRR-encoding and related genes studied.

Conclusion: Transcripts of many NBS-LRR-encoding and related genes were defined. Most were present at low levels and exhibited tissue-specific expression patterns. Expression data are consistent with most Arabidopsis NBS-LRR-encoding and related genes functioning in plant defense responses but do not preclude other biological roles.

Show MeSH
Related in: MedlinePlus