Glycosylphosphatidylinositol (GPI) anchoring controls cell wall integrity, immune evasion and surface localization of ChFEM1 for infection of Cochlibolus heterostrophus

Glycosylphosphatidylinositol (GPI) anchoring represents a fundamental post-translational modification in eukaryotic cells. In fungi, this modification facilitates diverse biological functions through protein targeting to the cell wall, yet research on its roles in plant pathogenic fungi remains limited. This study elucidates the function of GPI anchoring in the maize fungal pathogen Cochlibolus heterostrophus. The research demonstrates widespread accumulation of GPI-anchored proteins in hyphae, appressorium and infection hyphae of C. heterostrophus. Deletion of ChGPI7, encoding a crucial enzyme in GPI anchor biosynthesis, substantially reduced vegetative growth, conidiation, and virulence through impaired appressorium formation and invasive growth. The ΔChgpi7 mutants exhibited marked deficiencies in cell wall integrity, leading to decreased stress resistance. Both ChGPI7 deletion and hydro fluoric acid (HF) pyridine treatment eliminated cell wall GPI-anchored proteins and exposed chitin, indicating that GPI-anchored proteins shield chitin from host immune recognition. Analysis identified 124 predicted GPI-anchored proteins in C. heterostrophus, including the putative cell wall glycoprotein ChFEM1. The deletion of ChFEM1 similarly reduced virulence and compromised infection structures and cell wall integrity. Additionally, ChGPI7 influenced both the cell wall localization and protein abundance of ChFEM1. These findings demonstrate that GPI anchoring mediates cell wall integrity and immune evasion during C. heterostrophus infection.