Universität zu Köln

Lee, Yoon Joo (2024). Characterization of hypertrophy-related effectors in Ustilago maydis induced tumor formation. PhD thesis, Universität zu Köln.

PDF Yoon Joo Lee_PhD_Dissertation.pdf Download (5MB)

Abstract

Ustilago maydis, a biotrophic corn smut fungus, secretes a cocktail of effectors in a spatiotemporal regulated manner to induce nutrient-rich tumors in all aerial parts of maize. By down-regulating genes linked to photosynthesis and increasing hexose accumulation in developing tumors, it manipulates the host’s metabolic processes and alters the sink-source transition (Doehlemann et al., 2008a; Horst et al., 2008, 2010). Cell type-specific transcriptome profiling of U. maydis has revealed that a set of effectors are highly and specifically up-regulated in hypertrophy, leading to the enlargement of tumor cells through endoreduplication (Matei et al., 2018). This study focused on identifying and functionally characterizing U. maydis effectors involved in hypertrophy induction. Through various screening and selection methods, we have identified hypertrophy-associated proteins 1-3 (Hap1-3) which are highly expressed at 2 days post infection (dpi) as potential hypertrophic mesophyll tumor cell (HTT) effectors. Pull-down of Hap effectors followed by mass-spectrometry analysis revealed that Hap effectors interact within the host cell, suggesting potential formation of effector complex. To gain insights into the cooperative mechanisms of Hap effectors, double and triple frameshift knockout mutants of hap1, hap2, and hap3 were generated, and a large-scale mass spectrometry analysis was conducted. These analyses revealed that Hap1 is the dominant HTT-related virulence factor. Furthermore, Hap1 interacted with maize Snf1-related protein kinase 1 (SnRK1), a key regulator of cellular energy and nutrient homeostasis, which is activated by phosphorylation under energy deprivation. To explore Hap1’s influence on the global gene expression within the host, RNA-seq analysis was performed. Hap1 showed high expression levels of AGPase3, SBEI, and AeI involved in starch biosynthesis. Conversely, the hap1 frameshift knockout mutant exhibited high expression of WRKY transcription factors, wall-associated kinases, and lectin domain-containing receptor kinases, indicating that the deletion of hap1 induces a plant defense response. Quantitative phosphoproteomics analysis comparing Hap1 frameshift knockout mutants to wild-type infected plants revealed that SnRK1 and key metabolic enzymes are increased in phosphorylation in response to infection by the wild-type. Our findings support a model where U. maydis, in the presence of Hap1, targets the SnRK1α subunit. This targeting prevents SnRK1 inhibition by high levels of T6P, thereby disrupting the antagonistic relationship between T6P and SnRK1 and reprogramming transcription required for starch metabolism and inhibiting sugar-induced immune signaling.

Item Type:	Thesis (PhD thesis)
Creators:	Creators Email ORCID ORCID Put Code Lee, Yoon Joo ylee4@uni-koeln.de UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-748684
Date:	November 2024
Language:	English
Faculty:	Faculty of Mathematics and Natural Sciences
Divisions:	Faculty of Mathematics and Natural Sciences > Department of Biology > Botanical Institute
Subjects:	Natural sciences and mathematics
Uncontrolled Keywords:	Keywords Language Ustilago English effector English SnRK1 English
Date of oral exam:	11 July 2024
Referee:	Name Academic Title Doehlemann, Gunther Professor Thomma, Bart Professor Hofmann, Kay Professor
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/74868