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Investigation of the multifunctional gene AOP3 expands the regulatory network fine-tuning glucosinolate production in Arabidopsis.

Jensen LM, Kliebenstein DJ, Burow M - Front Plant Sci (2015)

Bottom Line: In this study, we use transgenic plants in combination with natural variation to investigate the regulatory role of the AOP3 gene found in GS-AOP locus previously suggested to contribute to the regulation of glucosinolate defense compounds.Phenotypic analysis and QTL mapping in F2 populations with different AOP3 transgenes support that the enzymatic function and the AOP3 RNA both play a significant role in controlling glucosinolate accumulation.Furthermore, we find different loci interacting with either the enzymatic activity or the RNA of AOP3 and thereby extend the regulatory network controlling glucosinolate accumulation.

View Article: PubMed Central - PubMed

Affiliation: DNRF Center DynaMo, Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen Frederiksberg, Denmark ; Copenhagen Plant Science Centre, Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen Frederiksberg, Denmark.

ABSTRACT
Quantitative trait loci (QTL) mapping studies enable identification of loci that are part of regulatory networks controlling various phenotypes. Detailed investigations of genes within these loci are required to ultimately understand the function of individual genes and how they interact with other players in the network. In this study, we use transgenic plants in combination with natural variation to investigate the regulatory role of the AOP3 gene found in GS-AOP locus previously suggested to contribute to the regulation of glucosinolate defense compounds. Phenotypic analysis and QTL mapping in F2 populations with different AOP3 transgenes support that the enzymatic function and the AOP3 RNA both play a significant role in controlling glucosinolate accumulation. Furthermore, we find different loci interacting with either the enzymatic activity or the RNA of AOP3 and thereby extend the regulatory network controlling glucosinolate accumulation.

No MeSH data available.


Position of the transgene(s) in the four populations. Four different positions were found for the active AOP3 FL6 (red, orange) by QTL mapping, however, co-segregation of the surrounding regions for the insert on chromosome 2, 3, and 5 (orange), suggests chromosomal rearrangements causing that what looks like three inserts are one found on any of three chromosomes. The AOP3 FL9 (green) and UT10 (blue) were found at chromosome 1, whereas the AOP3 UT2 (purple) is positioned on chromosome 3. The closest marker for each insertion site is indicated as well as all significant markers for a phenotype is depicted in italics.
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Figure 6: Position of the transgene(s) in the four populations. Four different positions were found for the active AOP3 FL6 (red, orange) by QTL mapping, however, co-segregation of the surrounding regions for the insert on chromosome 2, 3, and 5 (orange), suggests chromosomal rearrangements causing that what looks like three inserts are one found on any of three chromosomes. The AOP3 FL9 (green) and UT10 (blue) were found at chromosome 1, whereas the AOP3 UT2 (purple) is positioned on chromosome 3. The closest marker for each insertion site is indicated as well as all significant markers for a phenotype is depicted in italics.

Mentions: The four F2 populations have randomly shuffled background loci as a result of recombination in the F1 generation. This allows us to use QTL mapping to investigate the effect of the AOP3 enzyme and AOP3 RNA on glucosinolate accumulation as well as to identify other unidentified loci that vary between the parents and link to glucosinolates. To provide genotype information for QTL mapping, the four populations were genotyped for 100 SNPs using Sequenom MassARRAY® (Tables S9–S12). The physical position of the AOP3 transgene in each population was found by genotyping the plants for the transgene or based on accumulation of the AOP3 product, 3ohp. Mapping of the AOP3 transgene in three of the populations (FL9, UT2, and UT10) were in agreement with the presence of a single transgene. In the Col-0 × Gie-0 FL9 F2 population the insertion mapped to chromosome 1, in the UT2 to a position on chromosome 3 and in the UT10 at chromosome 1 (Figure 6). Genotyping of the Col-0 × Gie-0 FL6 population showed the same pattern of SNP markers on parts of chromosome 2, 3, and 5 across the population, which suggests that these chromosome parts co-segregated. This observation can be explained by a chromosomal rearrangement causing the three chromosome parts to be located at one chromosome. We identified these three regions by QTL mapping for the ratio of AOP3 product to C3 glucosinolate, which correlates with the presence of the AOP3 transgene on all three chromosomes, thus, suggesting chromosomal rearrangement associated with the AOP3 insertion. We also identified a position on chromosome 1 that contained a second copy of the transgene. For further analysis, we therefore included AOP3 as a marker for presence, when either position was positive for the insertion.


Investigation of the multifunctional gene AOP3 expands the regulatory network fine-tuning glucosinolate production in Arabidopsis.

Jensen LM, Kliebenstein DJ, Burow M - Front Plant Sci (2015)

Position of the transgene(s) in the four populations. Four different positions were found for the active AOP3 FL6 (red, orange) by QTL mapping, however, co-segregation of the surrounding regions for the insert on chromosome 2, 3, and 5 (orange), suggests chromosomal rearrangements causing that what looks like three inserts are one found on any of three chromosomes. The AOP3 FL9 (green) and UT10 (blue) were found at chromosome 1, whereas the AOP3 UT2 (purple) is positioned on chromosome 3. The closest marker for each insertion site is indicated as well as all significant markers for a phenotype is depicted in italics.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4585220&req=5

Figure 6: Position of the transgene(s) in the four populations. Four different positions were found for the active AOP3 FL6 (red, orange) by QTL mapping, however, co-segregation of the surrounding regions for the insert on chromosome 2, 3, and 5 (orange), suggests chromosomal rearrangements causing that what looks like three inserts are one found on any of three chromosomes. The AOP3 FL9 (green) and UT10 (blue) were found at chromosome 1, whereas the AOP3 UT2 (purple) is positioned on chromosome 3. The closest marker for each insertion site is indicated as well as all significant markers for a phenotype is depicted in italics.
Mentions: The four F2 populations have randomly shuffled background loci as a result of recombination in the F1 generation. This allows us to use QTL mapping to investigate the effect of the AOP3 enzyme and AOP3 RNA on glucosinolate accumulation as well as to identify other unidentified loci that vary between the parents and link to glucosinolates. To provide genotype information for QTL mapping, the four populations were genotyped for 100 SNPs using Sequenom MassARRAY® (Tables S9–S12). The physical position of the AOP3 transgene in each population was found by genotyping the plants for the transgene or based on accumulation of the AOP3 product, 3ohp. Mapping of the AOP3 transgene in three of the populations (FL9, UT2, and UT10) were in agreement with the presence of a single transgene. In the Col-0 × Gie-0 FL9 F2 population the insertion mapped to chromosome 1, in the UT2 to a position on chromosome 3 and in the UT10 at chromosome 1 (Figure 6). Genotyping of the Col-0 × Gie-0 FL6 population showed the same pattern of SNP markers on parts of chromosome 2, 3, and 5 across the population, which suggests that these chromosome parts co-segregated. This observation can be explained by a chromosomal rearrangement causing the three chromosome parts to be located at one chromosome. We identified these three regions by QTL mapping for the ratio of AOP3 product to C3 glucosinolate, which correlates with the presence of the AOP3 transgene on all three chromosomes, thus, suggesting chromosomal rearrangement associated with the AOP3 insertion. We also identified a position on chromosome 1 that contained a second copy of the transgene. For further analysis, we therefore included AOP3 as a marker for presence, when either position was positive for the insertion.

Bottom Line: In this study, we use transgenic plants in combination with natural variation to investigate the regulatory role of the AOP3 gene found in GS-AOP locus previously suggested to contribute to the regulation of glucosinolate defense compounds.Phenotypic analysis and QTL mapping in F2 populations with different AOP3 transgenes support that the enzymatic function and the AOP3 RNA both play a significant role in controlling glucosinolate accumulation.Furthermore, we find different loci interacting with either the enzymatic activity or the RNA of AOP3 and thereby extend the regulatory network controlling glucosinolate accumulation.

View Article: PubMed Central - PubMed

Affiliation: DNRF Center DynaMo, Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen Frederiksberg, Denmark ; Copenhagen Plant Science Centre, Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen Frederiksberg, Denmark.

ABSTRACT
Quantitative trait loci (QTL) mapping studies enable identification of loci that are part of regulatory networks controlling various phenotypes. Detailed investigations of genes within these loci are required to ultimately understand the function of individual genes and how they interact with other players in the network. In this study, we use transgenic plants in combination with natural variation to investigate the regulatory role of the AOP3 gene found in GS-AOP locus previously suggested to contribute to the regulation of glucosinolate defense compounds. Phenotypic analysis and QTL mapping in F2 populations with different AOP3 transgenes support that the enzymatic function and the AOP3 RNA both play a significant role in controlling glucosinolate accumulation. Furthermore, we find different loci interacting with either the enzymatic activity or the RNA of AOP3 and thereby extend the regulatory network controlling glucosinolate accumulation.

No MeSH data available.