Abstract
ABSTRACTTrehalose biosynthesis is found in fungi but not humans. Proteins involved in trehalose biosynthesis are essential for fungal pathogen virulence in humans and plants through multiple mechanisms. Loss of canonical trehalose biosynthesis genes in the human pathogen Aspergillus fumigatus significantly alters cell wall structure and integrity, though the mechanistic link between these virulence-associated pathways remains enigmatic. Here we characterize genes, called tslA and tslB, which encode proteins that contain domains similar to those corresponding to trehalose-6-phosphate phosphatase but lack critical catalytic residues for phosphatase activity. Loss of tslA reduces trehalose content in both conidia and mycelia, impairs cell wall integrity, and significantly alters cell wall structure. To gain mechanistic insights into the role that TslA plays in cell wall homeostasis, immunoprecipitation assays coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) were used to reveal a direct interaction between TslA and CsmA, a type V chitin synthase enzyme. TslA regulates not only chitin synthase activity but also CsmA sub-cellular localization. Loss of TslA impacts the immunopathogenesis of murine invasive pulmonary aspergillosis through altering cytokine production and immune cell recruitment. In conclusion, our data provide a novel model whereby proteins in the trehalose pathway play a direct role in fungal cell wall homeostasis and consequently impact fungus-host interactions.
Highlights
IMPORTANCE Human fungal infections are increasing globally due to HIV infections and increased use of immunosuppressive therapies for many diseases
Protein domain analyses revealed that TslA and TslB share domains similar to those of the trehalose-6-phosphate phosphatase (TPP) OrlA, such as the glycosyl transferase domain (GT1-TPS) and the halogen-associated dehydrogenase-like domain (HAD-TPP), as previously reported in A. niger [16]
ΔtslA showed restored cell wall phenotypes on both SMM and Sabouraud dextrose agar (SDA) in the presence of calcofluor white (CFW) (Fig. 2B). As both Congo red (CR) and CFW bind to chitin on the cell wall and inhibit growth, while CPG inhibits -1,3-glucan synthase, these results suggest that loss of TslA affects the chitin component of the fungal cell wall
Summary
IMPORTANCE Human fungal infections are increasing globally due to HIV infections and increased use of immunosuppressive therapies for many diseases. Components of the trehalose biosynthesis pathway are important for the virulence of human-pathogenic fungi, including Aspergillus fumigatus. We discovered a novel moonlighting role for a regulatory-like subunit of the trehalose biosynthesis pathway in A. fumigatus that regulates cell wall homeostasis through modulation of chitin synthase localization and activity. Characterization of the unstudied A. fumigatus trehalose regulatory subunits tslA and tslB revealed a surprising role for TslA in modulating fungal cell wall homeostasis. For the first time, our results provide novel insights into mechanisms through which the canonical fungal trehalose biosynthesis pathway directly impacts fungal cell wall homeostasis and the hostpathogen interaction
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