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Solanum torvum responses to the root-knot nematode Meloidogyne incognita.

Bagnaresi P, Sala T, Irdani T, Scotto C, Lamontanara A, Beretta M, Rotino GL, Sestili S, Cattivelli L, Sabatini E - BMC Genomics (2013)

Bottom Line: GO term enrichment analyses with the 390 Torvum DEG revealed enhancement of several processes as chitin catabolism and sesquiterpenoids biosynthesis, while no GO term enrichment was found with eggplant DEG.The genes identified from S. torvum catalogue, bearing high similarity to known nematode resistance genes, were further investigated in view of their potential role in the nematode resistance mechanism.By combining 454 pyrosequencing and microarray technology we were able to conduct a cost-effective global transcriptome profiling in a non-model species.The expression profiling of S. torvum responses to nematode infection points to sesquiterpenoids and chitinases as major effectors of nematode resistance.

View Article: PubMed Central - HTML - PubMed

Affiliation: Consiglio per la Ricerca e la Sperimentazione in Agricoltura, Genomics Research Centre, via S Protaso 302, I-29107 Fiorenzuola d’Arda, PC, Italy.

ABSTRACT

Background: Solanum torvum Sw is worldwide employed as rootstock for eggplant cultivation because of its vigour and resistance/tolerance to the most serious soil-borne diseases as bacterial, fungal wilts and root-knot nematodes. The little information on Solanum torvum (hereafter Torvum) resistance mechanisms, is mostly attributable to the lack of genomic tools (e.g. dedicated microarray) as well as to the paucity of database information limiting high-throughput expression studies in Torvum.

Results: As a first step towards transcriptome profiling of Torvum inoculated with the nematode M. incognita, we built a Torvum 3' transcript catalogue. One-quarter of a 454 full run resulted in 205,591 quality-filtered reads. De novo assembly yielded 24,922 contigs and 11,875 singletons. Similarity searches of the S. torvum transcript tags catalogue produced 12,344 annotations. A 30,0000 features custom combimatrix chip was then designed and microarray hybridizations were conducted for both control and 14 dpi (day post inoculation) with Meloidogyne incognita-infected roots samples resulting in 390 differentially expressed genes (DEG). We also tested the chip with samples from the phylogenetically-related nematode-susceptible eggplant species Solanum melongena. An in-silico validation strategy was developed based on assessment of sequence similarity among Torvum probes and eggplant expressed sequences available in public repositories. GO term enrichment analyses with the 390 Torvum DEG revealed enhancement of several processes as chitin catabolism and sesquiterpenoids biosynthesis, while no GO term enrichment was found with eggplant DEG.The genes identified from S. torvum catalogue, bearing high similarity to known nematode resistance genes, were further investigated in view of their potential role in the nematode resistance mechanism.

Conclusions: By combining 454 pyrosequencing and microarray technology we were able to conduct a cost-effective global transcriptome profiling in a non-model species. In addition, the development of an in silico validation strategy allowed to further extend the use of the custom chip to a related species and to assess by comparison the expression of selected genes without major concerns of artifacts. The expression profiling of S. torvum responses to nematode infection points to sesquiterpenoids and chitinases as major effectors of nematode resistance. The availability of the long sequence tags in S. torvum catalogue will allow precise identification of active nematocide/nematostatic compounds and associated enzymes posing the basis for exploitation of these resistance mechanisms in other species.

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Heatmap of Torvum DEG (modulated) genes in class isoprenoid biosynthetic process. Expression patterns of the 10 modulated Torvum genes in GO class ‘isoprenoid biosynthetic process’ term (GO:0008299) are shown. Eggplant validated genes are boxed in red over the heatmap.
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Figure 7: Heatmap of Torvum DEG (modulated) genes in class isoprenoid biosynthetic process. Expression patterns of the 10 modulated Torvum genes in GO class ‘isoprenoid biosynthetic process’ term (GO:0008299) are shown. Eggplant validated genes are boxed in red over the heatmap.

Mentions: GO analysis of torvum DEGs revealed an enrichment for ‘isoprenoid biosynthetic process’ term (GO:0008299). Figure 6 provides an overview of expression pattern of 131 Torvum genes annotated with this term. Noteworthy, Torvum samples cluster together, while eggplant samples (despite several validated genes boxed in red) appear to respond less coherently, as control and infected samples do not cluster together. Figure 7 details expression patterns for the 10 modulated Torvum genes (eggplant validated counterparts boxed in red), while fold change data and the top blast hit for each of the modulated genes are reported in Table 4. Intriguingly, within this group of ‘isoprenoid biosynthetic process’ DEGs, sesquiterpene and diterpenoid biosynthetic genes undergo opposite modulation, i.e. induced the former and repressed the latter. In plants, diterpenes (20C, 4 isoprene units) are produced via the plastid pathway (MEP/DOXP pathway), whereas sesquiterpenes (15C, three isoprene units) derive from cytosolic mevalonate pathway [41]. Based on Blast2GO annotation and BlastX hits as reported in Table 5, tor5_c9415 encodes for a cytP450 enzyme with as best hits several CYP450, including epi-aristolochene 1,3 dihydroxylase and premnaspirodiene oxygenase (Cyt P450 71D55) [42]. This P450 enzyme catalyzes several hydroxylations for sesquiterpene substrates including phytoalexins as solavetivone. In turn, tor5_rep_c18585 and tor5_rep_c114 show both as best hit sesquiterpene synthase 2, while tor5_c8884 shows as best hit potato vetispiradiene synthase. The tor5_rep_c2244 (best hit 3-hydroxy-3-methylglutaryl CoAa reductase; HMGR) and tor5_c9415 (best hit cytochrome p450) are the only two genes within Torvum DEGs present in the validated eggplant dataset although they are not differentially expressed. HMGR plays a critical role in isoprenoid biosynthesis as catalyzes the first committed and rate limiting step in isoprenoid biosynthesis [43]. Thus, its presence in validated eggplant is a bona fide comparative clue towards the fact that the isoprenoid biosynthetic pathway is not up-regulated (at least at transcriptional level) following nematode infection in eggplant.


Solanum torvum responses to the root-knot nematode Meloidogyne incognita.

Bagnaresi P, Sala T, Irdani T, Scotto C, Lamontanara A, Beretta M, Rotino GL, Sestili S, Cattivelli L, Sabatini E - BMC Genomics (2013)

Heatmap of Torvum DEG (modulated) genes in class isoprenoid biosynthetic process. Expression patterns of the 10 modulated Torvum genes in GO class ‘isoprenoid biosynthetic process’ term (GO:0008299) are shown. Eggplant validated genes are boxed in red over the heatmap.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750854&req=5

Figure 7: Heatmap of Torvum DEG (modulated) genes in class isoprenoid biosynthetic process. Expression patterns of the 10 modulated Torvum genes in GO class ‘isoprenoid biosynthetic process’ term (GO:0008299) are shown. Eggplant validated genes are boxed in red over the heatmap.
Mentions: GO analysis of torvum DEGs revealed an enrichment for ‘isoprenoid biosynthetic process’ term (GO:0008299). Figure 6 provides an overview of expression pattern of 131 Torvum genes annotated with this term. Noteworthy, Torvum samples cluster together, while eggplant samples (despite several validated genes boxed in red) appear to respond less coherently, as control and infected samples do not cluster together. Figure 7 details expression patterns for the 10 modulated Torvum genes (eggplant validated counterparts boxed in red), while fold change data and the top blast hit for each of the modulated genes are reported in Table 4. Intriguingly, within this group of ‘isoprenoid biosynthetic process’ DEGs, sesquiterpene and diterpenoid biosynthetic genes undergo opposite modulation, i.e. induced the former and repressed the latter. In plants, diterpenes (20C, 4 isoprene units) are produced via the plastid pathway (MEP/DOXP pathway), whereas sesquiterpenes (15C, three isoprene units) derive from cytosolic mevalonate pathway [41]. Based on Blast2GO annotation and BlastX hits as reported in Table 5, tor5_c9415 encodes for a cytP450 enzyme with as best hits several CYP450, including epi-aristolochene 1,3 dihydroxylase and premnaspirodiene oxygenase (Cyt P450 71D55) [42]. This P450 enzyme catalyzes several hydroxylations for sesquiterpene substrates including phytoalexins as solavetivone. In turn, tor5_rep_c18585 and tor5_rep_c114 show both as best hit sesquiterpene synthase 2, while tor5_c8884 shows as best hit potato vetispiradiene synthase. The tor5_rep_c2244 (best hit 3-hydroxy-3-methylglutaryl CoAa reductase; HMGR) and tor5_c9415 (best hit cytochrome p450) are the only two genes within Torvum DEGs present in the validated eggplant dataset although they are not differentially expressed. HMGR plays a critical role in isoprenoid biosynthesis as catalyzes the first committed and rate limiting step in isoprenoid biosynthesis [43]. Thus, its presence in validated eggplant is a bona fide comparative clue towards the fact that the isoprenoid biosynthetic pathway is not up-regulated (at least at transcriptional level) following nematode infection in eggplant.

Bottom Line: GO term enrichment analyses with the 390 Torvum DEG revealed enhancement of several processes as chitin catabolism and sesquiterpenoids biosynthesis, while no GO term enrichment was found with eggplant DEG.The genes identified from S. torvum catalogue, bearing high similarity to known nematode resistance genes, were further investigated in view of their potential role in the nematode resistance mechanism.By combining 454 pyrosequencing and microarray technology we were able to conduct a cost-effective global transcriptome profiling in a non-model species.The expression profiling of S. torvum responses to nematode infection points to sesquiterpenoids and chitinases as major effectors of nematode resistance.

View Article: PubMed Central - HTML - PubMed

Affiliation: Consiglio per la Ricerca e la Sperimentazione in Agricoltura, Genomics Research Centre, via S Protaso 302, I-29107 Fiorenzuola d’Arda, PC, Italy.

ABSTRACT

Background: Solanum torvum Sw is worldwide employed as rootstock for eggplant cultivation because of its vigour and resistance/tolerance to the most serious soil-borne diseases as bacterial, fungal wilts and root-knot nematodes. The little information on Solanum torvum (hereafter Torvum) resistance mechanisms, is mostly attributable to the lack of genomic tools (e.g. dedicated microarray) as well as to the paucity of database information limiting high-throughput expression studies in Torvum.

Results: As a first step towards transcriptome profiling of Torvum inoculated with the nematode M. incognita, we built a Torvum 3' transcript catalogue. One-quarter of a 454 full run resulted in 205,591 quality-filtered reads. De novo assembly yielded 24,922 contigs and 11,875 singletons. Similarity searches of the S. torvum transcript tags catalogue produced 12,344 annotations. A 30,0000 features custom combimatrix chip was then designed and microarray hybridizations were conducted for both control and 14 dpi (day post inoculation) with Meloidogyne incognita-infected roots samples resulting in 390 differentially expressed genes (DEG). We also tested the chip with samples from the phylogenetically-related nematode-susceptible eggplant species Solanum melongena. An in-silico validation strategy was developed based on assessment of sequence similarity among Torvum probes and eggplant expressed sequences available in public repositories. GO term enrichment analyses with the 390 Torvum DEG revealed enhancement of several processes as chitin catabolism and sesquiterpenoids biosynthesis, while no GO term enrichment was found with eggplant DEG.The genes identified from S. torvum catalogue, bearing high similarity to known nematode resistance genes, were further investigated in view of their potential role in the nematode resistance mechanism.

Conclusions: By combining 454 pyrosequencing and microarray technology we were able to conduct a cost-effective global transcriptome profiling in a non-model species. In addition, the development of an in silico validation strategy allowed to further extend the use of the custom chip to a related species and to assess by comparison the expression of selected genes without major concerns of artifacts. The expression profiling of S. torvum responses to nematode infection points to sesquiterpenoids and chitinases as major effectors of nematode resistance. The availability of the long sequence tags in S. torvum catalogue will allow precise identification of active nematocide/nematostatic compounds and associated enzymes posing the basis for exploitation of these resistance mechanisms in other species.

Show MeSH
Related in: MedlinePlus