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Time-course Investigation of Phytophthora infestans infection of potato leaf from three cultivars by quantitative proteomics

ABSTRACT: Potato Late blight is one the most important crop diseases worldwide. Even though potato has been studied for many years, the potato disease late blight still has a huge negative effect on the potato production. A total of three commercially available field potato cultivars of different resistance to late blight infection: Kuras (moderate), Sarpo Mira (highly resistant) and Bintje (very suseptable) under controlled green house growing conditions innoculated with a diversity of P. infestans populations. We used label-free quantitative proteomics to investigate the infection with P. infestans in a time-course study over 258 hours. Several key issues limits proteome analysis of potato leaf tissue4–6. Firstly, the immense complexity of the plant proteome which is further complicated by the presence of highly abundant proteins, such as ribulose bisphosphate carboxylase/oxygenase (RuBisCO). Secondly, plant leaf and potato in particular contain abundant levels amounts of phenols and polyphenols which hinder or, unless precautions are taken, completely prevent a successful protein extraction.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Solanum Tuberosum (potato)

SUBMITTER: Allan Stensballe

LAB HEAD: Allan Stensballe

PROVIDER: PXD002767 | Pride | 2016-01-05

REPOSITORIES: Pride

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Time-course investigation of Phytophthora infestans infection of potato leaf from three cultivars by quantitative proteomics.

Larsen Mia Kruse Guldstrand MK Jørgensen Malene Møller MM Bennike Tue Bjerg TB Stensballe Allan A

Data in brief 20151217

Potato late blight is one the most important crop diseases worldwide. Even though potato has been studied for many years, the potato disease late blight still has a vast negative effect on the potato production [1], [2], [3]. Late blight is caused by the pathogen Phytophthora infestans (P. infestans), which initiates infection through leaves. However, the biological activities during different stages of infection are poorly described, and could enable novel or improved ways of defeating late bli ...[more]

PMID: 26862565

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Project description:To gain novel molecular insights into quantitative late blight resistance, we performed a high-resolution quantitative analysis of gene expression using potato cultivars with contrasting SNP alleles at the StAOS2 locus associated with maturity corrected resistance (MCR). SuperSAGE samples were generated from uninfected and infected plants of the selected genotypes under controlled environmental conditions. Genotypes were pooled to reduce the influence of the genetic background on the transcriptome. Nine SuperSAGE samples were prepared from artificially inoculated plants in a growth chamber using total RNA of the pooled 14, 6 and 9 genotypes in groups A1, A2 and B2, respectively, at three infection time points T0, T1 and T2. Combining the tag counts of both NlaIII and DpnII libraries resulted in 1.1 to 6.2 million tags per sample. Of total 266361 unique tags (unitags), 52.6% matched to the potato genome sequence when up to three mismatches per 26 base pairs were allowed, and 23.3% matched without mismatch. Fifteen pair wise comparisons were performed between the nine SuperSAGE samples to identify transcripts that were differentially expressed in response to infection (six comparisons) or between three genotype pools at the infection time points T0, T1 and T2 (nine comparisons). The number of unitags per comparison ranged from 127 000 to 182 000 (average 158 000). Between 2100 and 11800 tags were differentially expressed in pair wise comparisons, depending on arbitrary cut-off p-values for a significant difference. The highest number of differences was observed for the comparison between genotype pools A1 and A2 one day after infection (A1-T1 vs A2-T1), and the lowest between genotype pools A2 and B2 two days after infection (B2-T2 vs A2-T2). The number of differences in response to infection and between genotype pools even before infection (T0) was in the same order of magnitude. Based on the annotations in the DFCI potato gene index, in the potato genome and in few cases by BLASTX searches against the protein database at NCBI, transcripts that showed reproducible differential expression over the infection time course or between genotype pools A1, A2 and B2 were grouped in 16 functional categories, with overlaps between categories. Genes with genotype dependent, constitutive differential expression provide excellent targets for developing novel diagnostic markers for breeding cultivars with improved quantitative resistance to late blight and possibly other biotic and abiotic stresses. Relevant in this respect appear, besides numerous genes of unknown or ill-defined function, genes with known function involved in stress responses, photosynthesis, protein biosynthesis, protein degradation via the 26S proteasome, transport of proteins, lipids, ions and other small molecules, cell wall structure and many others. Eighteen SuperSAGE libraries were constructed based on nine leaf samples from one infection experiment. For each time point (T0, T1 and T2) one leaflet each of 14, 6 and 9 SL genotypes in genotypic groups A1, A2 and B2, respectively, were pooled. Frozen pooled leaf tissue was powdered in a CryoMill. SuperSAGE libraries were generated at GenXPro GmbH (Frankfurt, Germany) essentially as described (Matsumura et al. 2010). To prevent amplification biases the TrueQuant technology was applied as described by B. Rotter (Patent application Nr. WO2009152928). Besides NlaIII (recognition site: 5M-bM-^@M-^Y-CATG-3M-bM-^@M-^Y), DpnII (recognition site: 5M-bM-^@M-^Y-GATC-3M-bM-^@M-^Y) was used as second anchoring enzyme, to capture transcripts without a NlaIII site. Therefore, the 26 bp tags carry either CATG or GATC at their 5M-bM-^@M-^Y end. The libraries were pooled and sequenced by Solexa/Illumina technology (Illumina, Inc., USA).

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	BIn_K258_1.raw	Raw
	BIn_K258_1b.raw	Raw
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	BIn_K258_3.raw	Raw