A Gβ protein and the TupA Co-Regulator Bind to Protein Kinase A Tpk2 to Act as Antagonistic Molecular Switches of Fungal Morphological Changes

PloS One
Thamarai K JangananAdrian R Walmsley

Abstract

The human pathogenic fungus Paracoccidioides brasiliensis (Pb) undergoes a morphological transition from a saprobic mycelium to pathogenic yeast that is controlled by the cAMP-signaling pathway. There is a change in the expression of the Gβ-protein PbGpb1, which interacts with adenylate cyclase, during this morphological transition. We exploited the fact that the cAMP-signaling pathway of Saccharomyces cerevisiae does not include a Gβ-protein to probe the functional role of PbGpb1. We present data that indicates that PbGpb1 and the transcriptional regulator PbTupA both bind to the PKA protein PbTpk2. PbTPK2 was able to complement a TPK2Δ strain of S. cerevisiae, XPY5a/α, which was defective in pseudohyphal growth. Whilst PbGPB1 had no effect on the parent S. cerevisiae strain, MLY61a/α, it repressed the filamentous growth of XPY5a/α transformed with PbTPK2, behaviour that correlated with a reduced expression of the floculin FLO11. In vitro, PbGpb1 reduced the kinase activity of PbTpk2, suggesting that inhibition of PbTpk2 by PbGpb1 reduces the level of expression of Flo11, antagonizing the filamentous growth of the cells. In contrast, expressing the co-regulator PbTUPA in XPY5a/α cells transformed with PbTPK2, but not untransfo...Continue Reading

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Methods Mentioned

BETA
GTPase
two-hybrid
fluorescence microscopy
light microscopy
PCR
protein assay
pulldown
pull-down
Y2H
electron-microscopy

Software Mentioned

Image J
PbTupA
Informax
LSM
Vector NTI
gene
Rotor

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