RESUMO
The conidia produced by Fusarium oxysporum f. sp. cubense (Foc), the causative agent of Fusarium Wilt of Banana (FWB), play central roles in the disease cycle, as the pathogen lacks a sexual reproduction process. Until now, the molecular regulation network of asexual sporogenesis has not been clearly understood in Foc. Herein, we identified and functionally characterized thirteen (13) putative sporulation-responsive genes in Foc, namely FocmedA(a), FocmedA(b), abaA-L, FocflbA, FocflbB, FocflbC, FocflbD, FocstuA, FocveA, FocvelB, wetA-L, FocfluG and Foclae1. We demonstrated that FocmedA(a), abaA-L, wetA-L, FocflbA, FocflbD, FocstuA, FocveA and Foclae1 mediate conidiophore formation, whereas FocmedA(a) and abaA-L are important for phialide formation and conidiophore formation. The expression level of abaA-L was significantly decreased in the ΔFocmedA(a) mutant, and yeast one-hybrid and ChIP-qPCR analyses further confirmed that FocMedA(a) could bind to the promoter of abaA-L during micro- and macroconidiation. Moreover, the transcript abundance of the wetA-L gene was significantly reduced in the ΔabaA-L mutant, and it not only was found to function as an activator of micro- and macroconidium formation but also served as a repressor of chlamydospore production. In addition, the deletions of FocflbB, FocflbC, FocstuA and Foclae1 resulted in increased chlamydosporulation, whereas FocflbD and FocvelB gene deletions reduced chlamydosporulation. Furthermore, FocflbC, FocflbD, Foclae1 and FocmedA(a) were found to be important regulators for pathogenicity and fusaric acid synthesis in Foc. The present study therefore advances our understanding of the regulation pathways of the asexual development and functional interdependence of sporulation-responsive genes in Foc.
RESUMO
Apical secretion at hyphal tips is important for the growth and development of filamentous fungi. In this study, we analyzed the role of the Rab GTPases FoSec4 involved in the secretion of the banana wilt fungal pathogen Fusarium odoratissimum. We found that the deletion of FoSEC4 affects the activity of extracellular hydrolases and protein secretion, indicating that FoSec4 plays an important role in the regulation of protein secretion in F. odoratissimum. As a typical Rab GTPase, Sec4 participates in the Rab cycle through the conversion between the active GTP-bound state and the inactive GDP-bound state, which is regulated by guanine nucleate exchange factors (GEFs) and GTPase-activating proteins (GAPs). We further found that FoSec2 can interact with dominant-negative FoSec4 (GDP-bound and nucleotide-free form, FoSec4DN), and that FoGyp5 can interact with dominant active FoSec4 (GTP-bound and constitutively active form, FoSec4CA). We evaluated the biofunctions of FoSec4, FoSec2 and FoGyp5, and found that FoSec4 is involved in the regulation of vegetative growth, reproduction, pathogenicity and the environmental stress response of F. odoratissimum, and that FocSec2 and FoGyp5 perform biofunctions consistent with FoSec4, indicating that FoSec2 and FoGyp5 may work as the GEF and the GAP, respectively, of FoSec4 in F. odoratissimum. We further found that the amino-terminal region and Sec2 domain are essential for the biological functions of FoSec2, while the carboxyl-terminal region and Tre-2/Bub2/Cdc16 (TBC) domain are essential for the biological functions of FoGyp5. In addition, FoSec4 mainly accumulated at the hyphal tips and partially colocalized with Spitzenkörper; however, FoGyp5 accumulated at the periphery of Spitzenkörper, suggesting that FoGyp5 may recognize and inactivate FoSec4 at a specific location in hyphal tips.
