Two eisosome proteins play opposite roles in autophagic control and sustain cell integrity, function and pathogenicity in Beauveria bassiana.

Pil1A and Pil1B are two core eisosome proteins that are homologous to yeast Pil1/Lsp1 or filamentous fungal Pil1A/Pil1B but have been unexplored in entomopathogenic fungi. Here we examined subcellular localization and functions of Pil1A and Pil1B in Beauveria bassiana, a fungal insect pathogen. Either localization or co-localization experiments of the two proteins demonstrated that Pil1A and Pil1B were simultaneously localized at the periphery of hyphal cells for formation of stable, punctuate spots in B. bassiana. This is different from a reliance of proper Lsp1/Pil1B localization upon Pil1/Pil1A in other fungi. Deletions of pil1A and pil1B caused opposite changes in expression of many autophagy-related genes and formation of intravacuolar autophagosomes. Such opposite changes were restored to nearly normal status by exogenous rapamycin, implicating a link of Pil1A/B to the target of rapamycin signalling pathway. All single/double deletion mutants of pil1A and pil1B lost almost all pathogenicity due to reduced ability to secrete Pr1 proteases for cuticle degradation. They also showed differential changes in cell wall integrity and multiple stress responses. These findings unveil opposite roles for Pil1A and Pil1B in autophagic regulation and an essentiality of both for cell integrity, function and pathogenicity of the fungal entomopathogen.