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Novel candidate genes influencing natural variation in potato tuber cold sweetening identified by comparative proteomics and association mapping.

Fischer M, Schreiber L, Colby T, Kuckenberg M, Tacke E, Hofferbert HR, Schmidt J, Gebhardt C - BMC Plant Biol. (2013)

Bottom Line: The molecular basis of CIS in potato tubers is of interest not only in basic research on plant adaptation to environmental stress but also in applied research, since high amounts of reducing sugars affect negatively the quality of processed food products such as potato chips.We compared by two-dimensional polyacrylamide gel electrophoresis the tuber proteomes of cultivars highly diverse for CIS.Novel SNP's diagnostic for increased tuber starch content, starch yield and chip quality were identified, which are useful for selecting improved potato processing cultivars.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Plant Breeding and Genetics, Max-Planck-Institute for Plant Breeding Research, Cologne, Germany. fischer@mpipz.mpg.de

ABSTRACT

Background: Higher plants evolved various strategies to adapt to chilling conditions. Among other transcriptional and metabolic responses to cold temperatures plants accumulate a range of solutes including sugars. The accumulation of the reducing sugars glucose and fructose in mature potato tubers during exposure to cold temperatures is referred to as cold induced sweetening (CIS). The molecular basis of CIS in potato tubers is of interest not only in basic research on plant adaptation to environmental stress but also in applied research, since high amounts of reducing sugars affect negatively the quality of processed food products such as potato chips. CIS-tolerance varies considerably among potato cultivars. Our objective was to identify by an unbiased approach genes and cellular processes influencing natural variation of tuber sugar content before and during cold storage in potato cultivars used in breeding programs. We compared by two-dimensional polyacrylamide gel electrophoresis the tuber proteomes of cultivars highly diverse for CIS. DNA polymorphisms in genomic sequences encoding differentially expressed proteins were tested for association with tuber starch content, starch yield and processing quality.

Results: Pronounced natural variation of CIS was detected in tubers of a population of 40 tetraploid potato cultivars. Significant differences in protein expression were detected between CIS-tolerant and CIS-sensitive cultivars before the onset as well as during cold storage. Identifiable differential proteins corresponded to protease inhibitors, patatins, heat shock proteins, lipoxygenase, phospholipase A1 and leucine aminopeptidase (Lap). Association mapping based on single nucleotide polymorphisms supported a role of Lap in the natural variation of the quantitative traits tuber starch and sugar content.

Conclusions: The combination of comparative proteomics and association genetics led to the discovery of novel candidate genes for influencing the natural variation of quantitative traits in potato tubers. One such gene was a leucine aminopeptidase not considered so far to play a role in starch sugar interconversion. Novel SNP's diagnostic for increased tuber starch content, starch yield and chip quality were identified, which are useful for selecting improved potato processing cultivars.

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Effect of the SNP allele StLapN-A2746on average RSC of 40 cultivars during cold storage. Cultivars were grouped according to presence (triangles) or absence (squares) of SNP allele A2746. The genotypic groups were tested by ANOVA for significant differences between means of RSC (log transformed) before and after 1, 2, 4 and 12 weeks of cold storage. The mean RSC was different between the genotypic groups at all time points (**: 0.01 > p <0,001; ***: p < 0.001). The amount of variance explained by StLapN-SNP2746 (R2) at the different time points is given as percentage. R2 values increased during cold storage.
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Figure 4: Effect of the SNP allele StLapN-A2746on average RSC of 40 cultivars during cold storage. Cultivars were grouped according to presence (triangles) or absence (squares) of SNP allele A2746. The genotypic groups were tested by ANOVA for significant differences between means of RSC (log transformed) before and after 1, 2, 4 and 12 weeks of cold storage. The mean RSC was different between the genotypic groups at all time points (**: 0.01 > p <0,001; ***: p < 0.001). The amount of variance explained by StLapN-SNP2746 (R2) at the different time points is given as percentage. R2 values increased during cold storage.

Mentions: BLAST searches with the cDNA sequence X77015 corresponding to the differential Lap protein against the potato genome sequence [30] identified superscaffold PGSC0003DMB000000116 (chr12:910581..2552409), which contained a single gene model annotated as ‘neutral Lap’ (StLapN). The same superscaffold contained the marker loci AGPaseB-b (PGSC0003DMG400046891) and GP34[32], which anchored the superscaffold to the distal part of the long arm of potato chromosome XII. The sequence X77015 clearly matched to two regions (> 95% sequence identity), indicating a tandem repeat of two Lap genes with ten exons each, located within a 17 kbp region of superscaffold PGSC0003DMB000000116 (chr12:910581..2552409). The genomic organization was confirmed by aligning tomato Lap sequences U50151 (acidic Lap, SlLapA) and AF510743 (neutral Lap, SlLapN) with the superscaffold. The cDNA sequence of accession X77015 was more similar to SlLapA (94.6%) than to SlLapN (86.2%) and StLapN (87.2%), indicating that the differential protein annotated as ‘chloroplastic leucine aminopeptidase’ corresponded to StLapA. A specific amplicon suitable for sequencing and SNP detection could only be obtained from the physically closely linked StLapN gene. Seventeen SNPs and one indel (Additional file 6: Figure S1) were tested for association with the tuber traits in the CHIPS-ALL population. Four SNPs were associated with one or more tuber trait, two of which showed only very small effects (data not shown). SNP2746 showed highly significant associations with tuber starch content (TSC), tuber starch yield (TSY), chip quality after harvest (CQA) and chip quality after cold storage (CQS). Cultivars containing the minor frequency allele A2746 showed significant increased average trait values (Table 3). Furthermore, when scored in the 40 cultivars evaluated for RSC during cold storage, the SNP allele A2746 was detected in 16 cultivars, preferentially in cultivars with low RSC and particularly after four weeks cold storage (Figure 1). This observation was confirmed when the 40 cultivars were grouped based on presence and absence of the SNP allele A2746 and tested for difference between the RSC group means. At all time-points, the genotypic group having at least one dosage of the A2746 allele accumulated significantly lower amounts of reducing sugars than the group homozygous for the allele G2746 (Figure 4). The positive effect of SNP2746 increased during cold storage, consistent with the result obtained in the CHIPS-ALL population.


Novel candidate genes influencing natural variation in potato tuber cold sweetening identified by comparative proteomics and association mapping.

Fischer M, Schreiber L, Colby T, Kuckenberg M, Tacke E, Hofferbert HR, Schmidt J, Gebhardt C - BMC Plant Biol. (2013)

Effect of the SNP allele StLapN-A2746on average RSC of 40 cultivars during cold storage. Cultivars were grouped according to presence (triangles) or absence (squares) of SNP allele A2746. The genotypic groups were tested by ANOVA for significant differences between means of RSC (log transformed) before and after 1, 2, 4 and 12 weeks of cold storage. The mean RSC was different between the genotypic groups at all time points (**: 0.01 > p <0,001; ***: p < 0.001). The amount of variance explained by StLapN-SNP2746 (R2) at the different time points is given as percentage. R2 values increased during cold storage.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750364&req=5

Figure 4: Effect of the SNP allele StLapN-A2746on average RSC of 40 cultivars during cold storage. Cultivars were grouped according to presence (triangles) or absence (squares) of SNP allele A2746. The genotypic groups were tested by ANOVA for significant differences between means of RSC (log transformed) before and after 1, 2, 4 and 12 weeks of cold storage. The mean RSC was different between the genotypic groups at all time points (**: 0.01 > p <0,001; ***: p < 0.001). The amount of variance explained by StLapN-SNP2746 (R2) at the different time points is given as percentage. R2 values increased during cold storage.
Mentions: BLAST searches with the cDNA sequence X77015 corresponding to the differential Lap protein against the potato genome sequence [30] identified superscaffold PGSC0003DMB000000116 (chr12:910581..2552409), which contained a single gene model annotated as ‘neutral Lap’ (StLapN). The same superscaffold contained the marker loci AGPaseB-b (PGSC0003DMG400046891) and GP34[32], which anchored the superscaffold to the distal part of the long arm of potato chromosome XII. The sequence X77015 clearly matched to two regions (> 95% sequence identity), indicating a tandem repeat of two Lap genes with ten exons each, located within a 17 kbp region of superscaffold PGSC0003DMB000000116 (chr12:910581..2552409). The genomic organization was confirmed by aligning tomato Lap sequences U50151 (acidic Lap, SlLapA) and AF510743 (neutral Lap, SlLapN) with the superscaffold. The cDNA sequence of accession X77015 was more similar to SlLapA (94.6%) than to SlLapN (86.2%) and StLapN (87.2%), indicating that the differential protein annotated as ‘chloroplastic leucine aminopeptidase’ corresponded to StLapA. A specific amplicon suitable for sequencing and SNP detection could only be obtained from the physically closely linked StLapN gene. Seventeen SNPs and one indel (Additional file 6: Figure S1) were tested for association with the tuber traits in the CHIPS-ALL population. Four SNPs were associated with one or more tuber trait, two of which showed only very small effects (data not shown). SNP2746 showed highly significant associations with tuber starch content (TSC), tuber starch yield (TSY), chip quality after harvest (CQA) and chip quality after cold storage (CQS). Cultivars containing the minor frequency allele A2746 showed significant increased average trait values (Table 3). Furthermore, when scored in the 40 cultivars evaluated for RSC during cold storage, the SNP allele A2746 was detected in 16 cultivars, preferentially in cultivars with low RSC and particularly after four weeks cold storage (Figure 1). This observation was confirmed when the 40 cultivars were grouped based on presence and absence of the SNP allele A2746 and tested for difference between the RSC group means. At all time-points, the genotypic group having at least one dosage of the A2746 allele accumulated significantly lower amounts of reducing sugars than the group homozygous for the allele G2746 (Figure 4). The positive effect of SNP2746 increased during cold storage, consistent with the result obtained in the CHIPS-ALL population.

Bottom Line: The molecular basis of CIS in potato tubers is of interest not only in basic research on plant adaptation to environmental stress but also in applied research, since high amounts of reducing sugars affect negatively the quality of processed food products such as potato chips.We compared by two-dimensional polyacrylamide gel electrophoresis the tuber proteomes of cultivars highly diverse for CIS.Novel SNP's diagnostic for increased tuber starch content, starch yield and chip quality were identified, which are useful for selecting improved potato processing cultivars.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Plant Breeding and Genetics, Max-Planck-Institute for Plant Breeding Research, Cologne, Germany. fischer@mpipz.mpg.de

ABSTRACT

Background: Higher plants evolved various strategies to adapt to chilling conditions. Among other transcriptional and metabolic responses to cold temperatures plants accumulate a range of solutes including sugars. The accumulation of the reducing sugars glucose and fructose in mature potato tubers during exposure to cold temperatures is referred to as cold induced sweetening (CIS). The molecular basis of CIS in potato tubers is of interest not only in basic research on plant adaptation to environmental stress but also in applied research, since high amounts of reducing sugars affect negatively the quality of processed food products such as potato chips. CIS-tolerance varies considerably among potato cultivars. Our objective was to identify by an unbiased approach genes and cellular processes influencing natural variation of tuber sugar content before and during cold storage in potato cultivars used in breeding programs. We compared by two-dimensional polyacrylamide gel electrophoresis the tuber proteomes of cultivars highly diverse for CIS. DNA polymorphisms in genomic sequences encoding differentially expressed proteins were tested for association with tuber starch content, starch yield and processing quality.

Results: Pronounced natural variation of CIS was detected in tubers of a population of 40 tetraploid potato cultivars. Significant differences in protein expression were detected between CIS-tolerant and CIS-sensitive cultivars before the onset as well as during cold storage. Identifiable differential proteins corresponded to protease inhibitors, patatins, heat shock proteins, lipoxygenase, phospholipase A1 and leucine aminopeptidase (Lap). Association mapping based on single nucleotide polymorphisms supported a role of Lap in the natural variation of the quantitative traits tuber starch and sugar content.

Conclusions: The combination of comparative proteomics and association genetics led to the discovery of novel candidate genes for influencing the natural variation of quantitative traits in potato tubers. One such gene was a leucine aminopeptidase not considered so far to play a role in starch sugar interconversion. Novel SNP's diagnostic for increased tuber starch content, starch yield and chip quality were identified, which are useful for selecting improved potato processing cultivars.

Show MeSH
Related in: MedlinePlus