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Functional characterization of Fusarium verticillioides CPP1, a gene encoding a putative protein phosphatase 2A catalytic subunit

Department of Plant Pathology and Microbiology, Program for the Biology of Filamentous Fungi, Texas A&M University, College Station, TX 77843-2132, USA

Correspondence
Won-Bo Shim
wbshim{at}tamu.edu

Fusarium verticillioides produces the mycotoxin fumonisin B₁ (FB₁) on maize kernels. In this study, we identified a putative protein phosphatase gene CPP1 in F. verticillioides, and investigated its role in FB₁ regulation. Previous work has shown that CPP1 expression is elevated in an FB₁-suppressing genetic background. Thus, we hypothesized that CPP1 is negatively associated with FB₁ production. To test this hypothesis, we generated a CPP1 knockout mutant, PP179, and studied the effects of gene deletion on FB₁ biosynthesis and fungal development. PP179 showed elevated expression of FUM genes, and in turn produced higher levels of FB₁ than the wild-type progenitor. Other significant mutant phenotypes included reduced radial growth on agar plates, reduced conidia germination rates, significantly increased macroconidia formation, and hyphal swelling. To verify that these phenotypes were directly due to CPP1 deletion, we complemented PP179 with the wild-type CPP1 gene. The complemented strain PPC4 showed FUM1 expression and FB₁ production similar to that of the wild-type, providing evidence that CPP1 is negatively associated with FB₁ biosynthesis. Other PP179 phenotypes, such as macroconidiation and hyphal swelling, were also restored to that of wild-type progenitor. Furthermore, we complemented F. verticillioides PP179 strain with Neurospora crassa wild-type ppe-1 gene, demonstrating that Cpp1 and PPE-1 proteins are functionally conserved. Pleiotropic effects of CPP1 deletion led us to hypothesize that CPP1 is associated with multiple downstream signalling pathways in F. verticillioides. Identification and functional characterization of downstream Cpp1-interacting proteins are necessary to better understand the complex regulatory mechanisms associated with Cpp1.

The GenBank/EMBL/DDBJ accession number for the nucleotide sequence of CPP1 is DQ924537.

A description of PP179 complementation using the YEC3 construct (section 1) and a description of how exogenous application of protein phosphatase inhibitor okadaic acid did not induce hyphal swelling in Fusarium verticillioides (section 2) are available as supplementary data with the online version of this paper.