The role of the multi-KH domain mRNA regulator Khd4 during pathogenic development of Ustilago maydis

• Applicant: Professor Dr. Michael Feldbrügge
• Subject Area: Metabolism, Biochemistry and Genetics of Microorganisms
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 539119480

Pathogenic fungi cause severe diseases and reduce crop yield substantially. In order to defeat fungi, we need to uncover their infection strategy. In recent years, substantial progress has been made in understanding host invasion using fungal effector proteins that counteract plant defense and alter host metabolism to support fungal proliferation. This sophisticated pathogenic programme is intensively regulated at the level of gene expression and depends on complex membrane trafficking such as protein secretion. However, it has become apparent that the underlying spatiotemporal control of protein expression is determined at the level of RNA biology. Importantly, the RNA regulatory processes involved are tightly linked with membrane trafficking including effector secretion. We study the model fungus Ustilago maydis causing corn smut disease. We discovered that the RNA-binding protein Kdh4 is crucial for infection since loss-of-function mutants are severely affected in pathogenicity. Khd4 deletion mutants still form infections hyphae but upon entry of the plant tissue a strong defense response is elicited, resulting in inhibition of fungal growth. To elucidate the function of Khd4 we have recently established HyperTRIBE for use in fungi to obtain a transcriptome-wide view of Khd4 target mRNAs in vivo. This revealed that during the formation of infectious hyphae, Khd4 regulates a defined set of mRNAs encoding regulatory proteins linked to GTPase signalling and membrane trafficking. One important function of Khd4 is the precise regulation of mRNA stability via its binding site AUACCC positioned in the 3´ UTR. Studying target mRNAs disclosed a novel link to the regulation of vacuole biogenesis as well as a potential link to Endoplasmic reticulum (ER) homeostasis. Based on these preliminary results we are now able to expand our approach and unravel the role of Khd4 during plant infection. We follow two main questions: How is Khd4 linked to ER functions, for example, during effector protein secretion and which mRNA targets are bound by Khd4 during plant invasion? During the stage when the fungus is in intimate contact with the plant, the HyperTRIBE methodology is perfectly suited and has clear advantages in comparison to other biochemical techniques identifying protein-bound mRNAs. In essence, studying target mRNAs and their mode of regulation in planta will clarify Khd4 functions during pathogenic development. We expect to find new regulatory principles and derive novel fungicide targets.

• DFG Programme: Research Grants