Regulatory Mechanism Of ABC Proteins In Iron Homeostasis And Pathogenesis Of Fusarium Graminearum

ATP-binding cassette(ABC)transporters family is the largest transporter gene family that is conserved present in living organisms.ABC transporters active transport various compounds via hydrolyzing ATP and play important roles in diverse biological processes,including heavy metal detoxification.lipid metabolism,signal transduction,multiple drug resistance,ribosome biogenesis and mRNA translation.To date,the study of fungal ABC proteins mainly focus on multiple drug resistance.However,the studies about ABC protein functions in iron homeostasis and pathogenesis are limited.Fusarium head blight(FHB)caused by Fusarium graminearumis one of the major diseases in wheat production.In addition to substantial yield losses.mycotoxin contamination poses a serious threat to food security and human health.In this study.we explored the functions of F.graminearum ABC proteins in iron homeostasis and virulence using bioinformatics,genetic and biochemical approaches.The study results show that:1.Generally,most fungi contain 30-45 ABC transporter genes in their genomes.However.F.graminearum genome has 62 ABC transporters that were divided into nine groups(ABC-A to-G,ABC-I and YDR061W-like)based on protein domain analysis.We obtained 60 ABC protein deletion mutants usin ghomologous recombination approach.For two ABC protein genes,Fg07101 and Fg01181,we failed to obtain knockout mutants after at least three independently transformation experiments,indicating that they may be essential for growth.The iron sensitivity screen of 60 ABC mutants shows that the ?FgAtm1 mutant is super sensitive to iron stress and contains a high level of intracellular iron.And the virulence assay of 60 ABC mutants shows that the ?FgArb1 mutant without transmembrane domain totally losts pathogenic ability on host tissues.2.The results of FgAtml regulating iron homeostasis mechanism are as follows:(1)The deletion of FgATM1 reduces the activity of cytosolic Fe-S proteins nitrite reductase FgNiiA and xanthine dehydrogenase FgXdh.(2)The reduced activities of FgNiiA and FgXdh activate the nitrogen metabolism regulator AreA.(3)FgAreA binds to the promoter of iron regulator FgHAPX and positively regulates its transcription.(4)FgHapX suppresses transcription of iron-consuming genes directly and also activates transcription of iron acquisition genes by repressing another transcription repressor FgSreA.(5)The function of FgHapX is dependent on phosphorylation by the Ser/Thr kinase FgYakl and its interaction with monothiol glutaredoxin FgGrx4.The study reveals that FgAtmk1 regulates iron homeostasis via the transcription factor cascade FgAreA-HapX in F.graminearum3.The results of FgArbl modulating virulence mechanism are as follows:(1)FgArbl interacts with the mitogen-activated protein kinase(MAPK)FgSte7,and partially modulates plant penetration by regulating the phosphorylation of FgGpmk1.(2)The FgArbl mutant exhibited dramatically reduced infective growth within wounded host tissues,likely resulting from its increased sensitivity to oxidative stresses,defective cell wall integrity and reduced deoxynivalenol(DON)production.(3)FgArb1 is importantfor the production of sexual and asexual spores.(4)FgArb1 is involved in the regulation of protein biosynthesis through impeding nuclear export of small ribosormal subunit.(5)acetylation modification at sites K28,K65,K341 and K525 in FgArb1 is required for its biological functions.This study uncovers that acetylated FgArb1 regulates the virulence of F.graminearum by regulating the infection structure formation and infective growth within host tissues.Results of this study promote the understanding of the functions of the ABC transporters in eukaryotes,and also provide theoretical basis for sustained management of FHB.