Functional analysis of Sec14-family proteins in the filamentous fungus aspergillus nidulans.

Filamentous fungi rely on polarized hyphal growth, a process fundamentally driven by vesicular transport. Sec14, a conserved phosphatidylinositol/phosphatidylcholine transfer protein first identified in Saccharomyces cerevisiae, and its Sec14-like SFH proteins have been implicated in multiple trafficking- and/or lipid-linked processes in diverse yeasts and pathogenic fungi. Despite substantial progress in Sec14 research in yeasts and a limited number of other fungi, systematic functional studies of Sec14/SFH proteins in filamentous fungi remain scarce. Here, using Aspergillus nidulans as a model filamentous fungus, we investigated the physiological function of Sec14-family genes. We identified eight orthologs of S. cerevisiae SEC14 and SFH genes in A. nidulans: secN (SEC14 ortholog) and seven sfh genes (sfhA-sfhG). Phenotypic analysis of single-deletion strains revealed that the ΔsecN strain exhibited severe growth retardation, accompanied by increased hyphal branching, defective conidiophore development, and extremely low conidia germination rate. Scanning electron microscopy further revealed that secN deletion impairs conidiophore morphogenesis during conidiation. Lipidomic analysis revealed that loss of secN altered the mycelial phospholipid composition. Analysis of lipid species recovered with affinity-purified SecN-V5 identified various phosphatidylinositol species, along with selected phosphatidylcholine species, as candidate SecN-associated lipids. In contrast, most sfh deletion strains showed no major growth defects under normal growth conditions, although they exhibited reduced growth sensitivity to calcofluor white. These findings indicate that SecN is a major Sec14-family protein required for normal growth, asexual development, and phospholipid homeostasis in A. nidulans, while suggesting that other Sfh proteins may have redundant or condition-specific functions. (244 words).