Screening And Identification Of Genes For Aflatoxin Biosynthesis Under Oxidative Stress By Transcriptomics Analyses

Aflatoxin(AF)is a carcinogenic secondary metabolite produced by Aspergillus flavus and A.parasiticus and can seriously endanger the health of humans and animals.The prevention and control of AF contamination is very significant for food safety and human health.It is particularly prominent to clarify AF biosynthesis and its molecular regulation.The regulation of transcription factors relative to AF biosynthetic synthesis was the research emphasis in this study.The main conclusions of this study are as follows:1.Here,growth and aflatoxin production by the different A.flavus strains(TF-12,TF-7,XinZ-16)was monitored for 8 days.The maximum aflatoxin production with respect to mycelial dry weight on the fifth day was observed.2.The mycelia were collected on the 5th day for transcriptome analysis.Compared to toxigenic strain,24 of 29 AF-biosynthesis genes showed reduced expression in atoxigenic strain XinZ-16,and 10 genes exhibited>2-fold.Additionally,the expression of 6 genes during the later stage of AF synthesis was significantly lower or not observed in atoxigenic strain,despite the nonexistence of gene deletions.Global transcription factors and other transcription factors which closely relate to AF biosynthesis and stress response were downregulated in the atoxigenic strain.Among them,one bZIP transcription factor Afap1,a YAP 1 homolog in response to oxidative-stress,was significantly downregulated in the atoxigenic strain.3.By NCBI BLAST analysis,Afap1,a basic leucine zipper(bZIP)transcription factor,was presumed to be a transcription factor associated with oxidative stress.4.The treatment of different oxidative stress on A.flavus strain TF-12 and XinZ-16 demonstrated that the growth of strain was significantly inhibited by the increased H2O2 and was completely inhibited at 40 mM.However,for quantitative analysis of aflatoxin production,HPLC showed that AFB1 concentration was increased by the increased H2O2 concentration until 10mM in TF-12.5.Based on the contruction of afapl mutants,treatment of 5 mM H2O2 completely inhibited the growth of afapl mutants but did not affect the growth of CA14PTs.In addition,it showed that AFB1 concentration in Aafapl was only about one fourth that of CA14PTs.Based on Real Time-PCR,the down-stream genes of aflatoxin biosynthesis gene cluster are significant decline,but the transtription factors related to oxidative stress are significant increase.This study revealed that the significantly differential transcriptional level of genes related to AF biosynthesis and stress response occurred between the toxigenic and atoxigenic strains and Afapl is a critical transcription factor in the oxidative-stress-response system and toxin production in A.flavus.