Abstract
Type 2C protein phosphatases (PP2Cs) play important roles in regulating many biological processes in eukaryotes. Currently, little is known about functions of PP2Cs in filamentous fungi. The causal agent of wheat head blight, Fusarium graminearum, contains seven putative PP2C genes, FgPTC1, -3, -5, -5R, -6, -7 and -7R. In order to investigate roles of these PP2Cs, we constructed deletion mutants for all seven PP2C genes in this study. The FgPTC3 deletion mutant (ΔFgPtc3-8) exhibited reduced aerial hyphae formation and deoxynivalenol (DON) production, but increased production of conidia. The mutant showed increased resistance to osmotic stress and cell wall-damaging agents on potato dextrose agar plates. Pathogencity assays showed that ΔFgPtc3-8 is unable to infect flowering wheat head. All of the defects were restored when ΔFgPtc3-8 was complemented with the wild-type FgPTC3 gene. Additionally, the FgPTC3 partially rescued growth defect of a yeast PTC1 deletion mutant under various stress conditions. Ultrastructural and histochemical analyses showed that conidia of ΔFgPtc3-8 contained an unusually high number of large lipid droplets. Furthermore, the mutant accumulated a higher basal level of glycerol than the wild-type progenitor. Quantitative real-time PCR assays showed that basal expression of FgOS2, FgSLT2 and FgMKK1 in the mutant was significantly higher than that in the wild-type strain. Serial analysis of gene expression in ΔFgPtc3-8 revealed that FgPTC3 is associated with various metabolic pathways. In contrast to the FgPTC3 mutant, the deletion mutants of FgPTC1, FgPTC5, FgPTC5R, FgPTC6, FgPTC7 or FgPTC7R did not show aberrant phenotypic features when grown on PDA medium or inoculated on wheat head. These results indicate FgPtc3 is the key PP2C that plays a critical role in a variety of cellular and biological functions, including cell wall integrity, lipid and secondary metabolisms, and virulence in F. graminearum.
Highlights
Reversible phosphorylation of proteins controlled by protein kinases and phosphatases is an important mechanism for regulating numerous biological processes in eukaryotes
Sequence analysis of FgPP2Cs Search for PP2C homologs in F. graminearum genome showed that this fungus contains seven putative PP2C genes: FgPTC1, -3, 5, -5R (FgPTC5-related family), -6, -7 and -7R (FgPTC7-related family)
Disruption of PP2Cs in F. graminearum To investigate the role of FgPP2Cs, we generated single gene deletion mutants of FgPTC1, -3, -5, -5R, -6, -7 and -7R using a homologous recombination strategy
Summary
Reversible phosphorylation of proteins controlled by protein kinases and phosphatases is an important mechanism for regulating numerous biological processes in eukaryotes. Protein phosphorylation and dephosphorylation generally occurs at tyrosine, serine, or threonine residues. Based on the substrate specificity, protein phosphatases are classed in two major groups: serine/threonine (Ser/Thr) phosphatases and tyrosine phosphatases (PTPs) [1]. Ser/Thr protein phosphatases have been classically categorized into two superfamilies: PPPs (phosphoprotein phosphatases) and PPMs (metal-dependent protein phosphatases). The PPP family consists of PP1, PP2A, and PP2B phosphatases. The PPM family contains type 2C protein phosphatases (PP2Cs) and pyruvate dehydrogenase phosphatase. PP2Cs are normally monomeric enzymes and require metal cation Mg2+ or Mn2+ for their dephosphorylation activities [2,3,4,5]
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