Localizing gene regulation reveals a staggered wood decay mechanism for the brown rot fungus Postia placenta

Significance Wood-decomposing fungi are key players in the carbon cycle and are models for making energy from lignocellulose, sustainably. Our study focuses on brown rot fungi that selectively remove carbohydrates, leaving most lignin behind. These fungi often decompose wood faster than their lignin-degrading white rot ancestors, despite losses in genes involved in plant cell wall hydrolysis. To explain brown rot, many have implicated reactive oxygen species (ROS) in facilitating hydrolysis, with microenvironmental gradients partitioning ROS from enzymes. By spatially colocalizing gene expression and enzyme activities as Postia placenta colonizes wood, we provide evidence of an oxidative-hydrolytic two-step mechanism controlled by differential expression, not microenvironments, and we highlight 549 genes (∼4% of the genome) that are upregulated during this unique pretreatment. Wood-degrading brown rot fungi are essential recyclers of plant biomass in forest ecosystems. Their efficient cellulolytic systems, which have potential biotechnological applications, apparently depend on a combination of two mechanisms: lignocellulose oxidation (LOX) by reactive oxygen species (ROS) and polysaccharide hydrolysis by a limited set of glycoside hydrolases (GHs). Given that ROS are strongly oxidizing and nonselective, these two steps are likely segregated. A common hypothesis has been that brown rot fungi use a concentration gradient of chelated metal ions to confine ROS generation inside wood cell walls before enzymes can infiltrate. We examined an alternative: that LOX components involved in ROS production are differentially expressed by brown rot fungi ahead of GH components. We used spatial mapping to resolve a temporal sequence in Postia placenta, sectioning thin wood wafers colonized directionally. Among sections, we measured gene expression by whole-transcriptome shotgun sequencing (RNA-seq) and assayed relevant enzyme activities. We found a marked pattern of LOX up-regulation in a narrow (5-mm, 48-h) zone at the hyphal front, which included many genes likely involved in ROS generation. Up-regulation of GH5 endoglucanases and many other GHs clearly occurred later, behind the hyphal front, with the notable exceptions of two likely expansins and a GH28 pectinase. Our results support a staggered mechanism for brown rot that is controlled by differential expression rather than microenvironmental gradients. This mechanism likely results in an oxidative pretreatment of lignocellulose, possibly facilitated by expansin- and pectinase-assisted cell wall swelling, before cellulases and hemicellulases are deployed for polysaccharide depolymerization.