# Structural insights into cellobiose dehydrogenases of a non-lignocellulolytic fungus and their transcriptional dynamics across different nutritional modes

## TL;DR

This paper explores the unique cellobiose dehydrogenase enzymes in Clonostachys rosea fungus and their roles in ecological interactions and biological control.

## Contribution

First characterization of the enriched CDH gene family in Clonostachys rosea and their ecological roles.

## Key findings

- Class-specific structural differences were found between CDHs in Clonostachys rosea.
- Transcriptome analysis shows CDH activation during host sensing and mycoparasitism.
- CDHs contribute to degradation of cellulosic compounds and plant root colonization.

## Abstract

Clonostachys rosea (Hypocreales, Bionectriaceae) is a ubiquitous fungus that colonizes living plants, digests organic material in soil (saprotroph) and parasitizes on or kills other fungi (necrotrophic mycoparasite) or nematodes and is thus of interest as a biological pest control agent. We recently found a high number of genes coding for cellobiose dehydrogenases (CDHs) in the C. rosea genome compared to significantly reduced or even missing gene content in saprotrophs such as Neurospora crassa, Trichoderma reesei, and other Trichoderma mycoparasites. Phylogenetic analysis of CDH enzymes in C. rosea revealed six members in AA3_1 class II and III, from which the enzymes of the class II group possess fungal carbohydrate-binding domains (CBM1). CDHs are extracellular enzymes produced by various wood-degrading fungi as they are proposed to be involved in cellulose, hemicellulose, and lignin biodegradation. Mass spectrometry analysis of class II CDHs confirmed the presence of oxidized products, consistent with the characteristics of single oxidized oligosaccharides, including glucose, mannose, maltose, cellobiose, lactose, and maltotriose. Structural and functional analysis revealed that class III CrCDHs exhibit significant differences in active site composition when compared to class II enzymes; however, no activity was found on the tested substrates. The functional transcript analysis in the mycoparasite C. rosea revealed activation of all cdh genes during self-recognition, sensing, and contact with the host fungus. The differential expression analysis indicated specific involvement of these CDHs in mycoparasitism and wheat root colonization and in degradation of simple and complex cellulosic compounds such as microcrystalline cellulose and wheat straw.

This study provides the first characterization of the unusually enriched cellobiose dehydrogenase (CDH) gene family in Clonostachys rosea, a mycoparasitic fungus of interest for biological control. We found class-specific structural differences between multiple CDHs belonging to distinct phylogenetic classes. Transcriptome profiling demonstrates their differential activation during host sensing, mycoparasitism, and plant root colonization, suggesting novel ecological roles beyond classical lignocellulose degradation. These findings establish a pioneering framework for understanding CDH diversification and its contribution to different fungal lifestyles.