Comparative analysis of temporal transcriptome reveals the relationship between pectin degradation and pathogenicity of defoliating Verticillium dahliae to Upland cotton (Gossypium hirsutum)

Zhang, F.; Zhang, J.; Chen, W.; Liu, X.; Li, C.; Cao, Y.; Zhao, T.; Lu, D.; Hui, Y.; Zhang, Y.; Chen, J.; Pepper, A. E.; Yu, J. Z.; Zhu, S.

doi:10.1101/2020.10.02.323402

bioRxiv · 10.1101/2020.10.02.323402

Comparative analysis of temporal transcriptome reveals the relationship between pectin degradation and pathogenicity of defoliating Verticillium dahliae to Upland cotton (Gossypium hirsutum)

Zhang, F.·Zhang, J.·Chen, W.·Liu, X.·Li, C.·Cao, Y.·Zhao, T.·Lu, D.·Hui, Y.·Zhang, Y.·Chen, J.·Pepper, A. E.·Yu, J. Z.·Zhu, S.

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Abstract

Verticillium wilt (VW), caused by Verticillium dahliae Kleb., is a major plant disease that causes heavy annual losses around the world, especially in Upland cotton (Gossypium hirsutum). The disease-causing pathogen can be classified into defoliating (D) and non-defoliating (ND) pathotypes based on the induced symptoms. At present, little is known about the complex mechanisms of fungal pathogenicity and cotton resistance to it. Comparative analysis of temporal transcriptome was performed on two V. dahliae strains, Vd_086 (D) and Vd_BP2 (ND), at key development stages (hyphal growth, microsclerotia production, and spore germination) to reveal the functional process on plant defoliation and death. Differentially expressed gene (DEG) analysis revealed a strong correlation between cell wall protein kinase activities and the early pathogenicity of defoliating Vd_086. With weighted gene co-expression network analysis (WGCNA), six specific gene modules were correlated with the biological traits of the fungal samples. Functional enrichment with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways together with DEG analysis revealed six pectin degrading enzymes including Polygalacturonase gene 1 (PG1), Pectate lyase gene (PEL) and Pectinesterase gene 1 (PME1) expressed in the early development of Vd_086 that may be related to the robust pathogenicity of this strain during the early invasion. The expression of four of these genes was verified by real-time quantitative reverse transcription PCR (qRT-PCR). In addition, we identified Mitogen-Activated Protein Kinase (MAPK) signaling "hub" genes that may regulate these pectinases. In a word, enhanced expression of pectin degradation enzymes is associated with the stronger pathogenicity of Vd_086 than Vd_BP2, especially at early infection stages. The disease-causing capability is likely regulated by MAPK signaling genes. This study provides new insight into molecular mechanisms of the plant-pathogen interaction on the VW disease, facilitating more effective control measures against this pathogen, including molecular breeding for the VW-resistant cotton cultivars. Author summaryVerticillium wilt (VW), caused by fungal pathogen Verticillium dahliae (Vd), is arguably the most devastating disease in cotton production for decades. Molecular biologists and plant breeders have been working hard to identify host plant resistant genes for many years but have met with little success due to the large complex genome of cotton. The V. dahliae strains are grouped in two pathotypes, of which defoliating (D) strains cause total leaf loss of infected cotton plants and non-defoliating (ND) strains do not. Comparative transcriptome analysis of D strain Vd_086 and ND strain Vd_BP2 identified the candidate genes and molecular mechanisms related to the Vd pathogenicity. Besides the difference in pathogenicity, these strains are distinguishable by the rate of hyphal elongation, microsclerotia production, and spore germination. With these phenotypes, transcriptome sequencing of both strains was performed at the three growth phases. By the combination of comparative transcriptomic differentially expressed gene (DEG) analysis and weighted gene correlation network analysis (WGCNA), cell wall-associated pectinase genes were found to be active at hyphal elongation stage of the V. dahliae pathogen and ribosome-related processes were activated for microsclerotia production. Gene modification processes were activated with many protein kinases at spore germination stage that for the next infection cycle. Furthermore, four pectinases in the pentose and glucuronate interconversion (PGI) pathway were identified and verified as highly expressed in the D strain with strong pathogenicity to Upland cotton (Gossypium hirsutum). Our results provided evidence in support of the hypothesis that stronger early pathogenicity of the D strain is resulted from greater plant cell wall pectin degradability. Transcription factors (TFs) and "hub" module genes were identified in searching of protein interaction for possible regulators of the recognized pectinases. TFs involved in mitogen-activated protein kinase (MAPK) signaling pathway were shown to regulate not only hyphal processes but also the entire growth period of V. dahliae. This is the first study known to use module extraction techniques of WGCNA to identify differentially co-expressed genes between two fungal pathotypes of V. dahliae strains. The study provides new insights into molecular mechanisms of the plant-pathogen interaction and may lead to molecular breeding for resistant cotton cultivars to effectively control this devastating disease.

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Zhang, F., Zhang, J., Chen, W., Liu, X., Li, C., Cao, Y., Zhao, T., Lu, D., Hui, Y., Zhang, Y., Chen, J., Pepper, A. E., Yu, J. Z., Zhu, S.. 2020-10-03. Comparative analysis of temporal transcriptome reveals the relationship between pectin degradation and pathogenicity of defoliating Verticillium dahliae to Upland cotton (Gossypium hirsutum). https://doi.org/10.1101/2020.10.02.323402

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