Abstract
Cassava, Manihot esculenta Crantz, is one of the major crops in developing countries. One of the main limitations in cassava production is Cassava Bacterial Blight (CBB), a disease caused by the gram-negative bacterium Xanthomonas axonopodis pv. manihotis (Xam). Although some resistant varieties have been identified, the mechanisms underlying cassava resistance to Xam remain largely unclarified. In this study, we characterized gene expression changes induced by Xam in resistant cassava plants along three different time points. Clustering analysis of differentially expressed genes (DEGs) induced by Xam inoculation showed four main groups of genes: early upregulated genes, late upregulated genes, genes constantly repressed and genes with small changes. Based on the data generated, a co-expression network was constructed, allowing the identification of hub genes associated to immune responses (Coiled Coil-Nucleotide Binding-Leucine Rich Repeat (CC-NB-LRR protein) and co-regulated with the phenolic metabolism (WD40 repeat-like protein). We characterized four transcription factor (TF) families that have been widely associated to the immune response in plants such as NAC (NAM (no apical meristem)/ATAF (Arabidopsis transcription activation factor)/CUC2 (cup-shaped cotyledon), bZIP (basic leucine zipper), WRKY and TCP (Teosinte branched1 (TB1) /Cincinnata (CIN)/ proliferating cell factor (PCF). In total 111, 86, 102 and 36 non-redundant genes were assigned to these TF families. Among these, seven WRKY, seven NAC, two bZIP and one TCP TFs, changed their expression in cassava plants inoculated with Xam, and were localized in the network as being co-expressed with other 41 DEGs. In addition four differentially expressed genes co-localized with QTLs associated to CBB resistance. This result shows the importance of combining gene expression, network reconstruction and mapping to identify proteins involved in plant immunity.
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Acknowledgements
We thank members of the Manihot Biotec and LAMFU groups for fruitful discussions of the manuscript.
Funding
This study was funded by COLCIENCIAS (Colombian Administrative Department for the Advancement of Science) 521–2011 and 0794–2013. JS received a PhD scholarship from Colciencias (call 528) and FG was supported by the Faculty of Sciences at Universidad de los Andes.
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The conceptualization of the research was done by CL, LLK, AB and SR. JCSS and FGC conducted the inoculation experiments under the guidance of CL and AB. LLK conducted the statistical analysis. FGC conducted the bioinformatics analysis. CL and FGC wrote the paper. All authors read, edited and approved the manuscript.
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Author FG declares that he has no conflict of interest. Author JS declares that she has no conflict of interest. Author SR declares that she has no conflict of interest Author AB declares that she has no conflict of interest. Author LL declares that she has no conflict of interest Author CL declares that he has no conflict of interest.
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Supplemental Fig. 1
Distribution of transcriptome expression (log2 CPM) of DEGs by cluster and days tested. (XLSX 224 kb) (PDF 1215 kb)
Supplemental Fig. 2
Co-expression network. a. Degree distribution of whole network b. Physical characteristics of each of the networks shown in a. (XLSX 137 kb) (PDF 186 kb)
Supplemental Fig. 3
Co-expression network associated to immune responses. a. Toy model describing the way as the different co-expression networks were built. Green nodes represent DEGs. b. Degree distribution of reduced sub-network. C. Physical characteristics of each of the subnetworks of DEG including their direct connectors. (XLSX 152 kb) (PDF 391 kb)
Supplemental Fig. 4
Comparison of NAC, bZIP, WRKY, and TCP candidates families to be involved in cassava’s resistance to Xam. TFs reported previously (black circles), and TF identified in the current study (green circles). (XLSX 14 kb) (PDF 10 kb)
Supplemental Fig. 5
Cassava resistance and susceptible responses genes expression profile. a. Comparison of susceptible and resistance cassava genes. Groups 5 and 7 represent Each group represent b. Expression profile of each of the gene groups in A. (PDF 1215 kb) (PNG 159 kb)
Supplemental table 1
(PDF 186 kb) (XLSX 46 kb)
Supplemental table 2
(XLSX 106 kb)
Supplemental tables 3, 4, 5 and 6
: GOs overrepresented in DEGs from cluster 1, cluster 2, cluster 3 and cluster 4, respectively. (XLSX 51 kb)
Supplemental table 7
NAC protein sequences. Sheet Summary: Gene cassava IDs and protein sequences (cassava genome V6.1) associated with previous report. Sheet Mochida et al. 2013: Protein sequences and gene IDs reported in Mochida et al. 2013, along with results of the blastp search carried out to associate previous protein sequences reported to current genome version. Sheet Hu et al. 2015: Protein sequences and gene IDs reported in Hu et al. 2015, along with results of the blastp search carried out to associate previous protein sequences reported to current genome version. (XLSX 197 kb)
Supplemental table 8
bZIP protein sequences. Sheet Summary: Gene cassava IDs and protein sequences (cassava genome V6.1) associated with previous report. Sheet Mochida et al. 2013: Protein sequences and gene IDs reported in Mochida et al. 2013, along with results of the blastp search carried out to associate previous protein sequences reported to current genome version. Sheet Hu et al. 2016: Protein sequences and gene IDs reported in Hu et al. 2016, along with results of the blastp search carried out to associate previous protein sequences reported to current genome version. (XLSX 224 kb)
Supplemental table 9
WRKY protein sequences. Sheet Summary: Gene cassava IDs and protein sequences (cassava genome V6.1) associated with previous report. Sheet Mochida et al. 2013: Protein sequences and gene IDs reported in Mochida et al. 2013, along with results of the blastp search carried out to associate previous protein sequences reported to current genome version. Sheet Wei et al. 2016: Protein sequences and gene IDs reported in Wei et al. 2016, along with results of the blastp search carried out to associate previous protein sequences reported to current genome version. (XLSX 137 kb)
Supplemental table 10
CTP protein sequences. Sheet Summary: Gene cassava IDs and protein sequences (cassava genome V6.1) associated with previous report. Sheet Mochida et al. 2013: Protein sequences and gene IDs reported in Mochida et al. 2013, along with results of the blastp search carried out to associate previous protein sequences reported to current genome version. (XLSX 152 kb)
Supplemental table 11
(XLSX 14 kb)
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Gómez-Cano, F., Soto, J., Restrepo, S. et al. Gene co-expression network for Xanthomonas-challenged cassava reveals key regulatory elements of immunity processes. Eur J Plant Pathol 153, 1083–1104 (2019). https://doi.org/10.1007/s10658-018-01628-4
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DOI: https://doi.org/10.1007/s10658-018-01628-4