The Molecular Mechanism For The T-2 Toxin-induced Transcriptional Regulation Of Chicken CYP1A5

T-2 toxin belongs to type-A trichothecenes,which is mainly produced by Fusariumgenus.Itis one of the major toxins that naturally occur in contaminated feed and food.Compared with humans,it is more harmful to poultry due to its higher probability of exposure to T-2 toxin.C ytochrome P450(C YP)is a group of structurally and functionally related genes encoding membrane proteins.P450 occurs in most oforganisms,mainly participating the metabolism of various endogenous and exogenous compounds.Previous studies in our laboratoryhave shownthat chicken CYP1A5 was involved in the metabolism of T-2 toxin,meanwhile T-2 toxin could induce chicken CYP1A5 up-regulation at transcriptional level.To date,the molecular mechanism of T-2 toxin and its metabolites regulatingthe expression of chicken CYP1A5 remains unclear.Therefore,our present study will be helpful to illustratethe mechanisms of the formation of harmful substances in poultry products,residuesandtoxic effects.To obtain the different expression due to T-2 toxin treatment in primary chicken embryo hepatocytes,RNA-seq was used.In the results,we found that CYP1A4 and CYP1A5 were the first and the third most up-regulated genes.At the same time,we found that a variety of mycotoxins can induce chicken CYP1A4 and CYP1A5 up-regulated at transcriptional level,but only T-2 toxin can induce thousand times expression of chicken CYP1A4 and hundred times expression of CYP1A5.We verified that T-2 toxin can induce the up-regulated expression of chicken CYP1A5 at the protein level by Western blot.To compare the induction of the CYP1 A family with other trichothecenes thatare similar to the T-2 toxin molecular structure,we found that the acetyl group at the 15-position C atom are related to this phenomenon.In order to clarify the molecular mechanism of T-2 toxin regulation of chicken CYP1A5 expression,we cloned approximatly 2.4 kb upstream sequence of chicken CYP1A5.Using Dula-Luciferase Reporter System combined with promoter deletion analysis,we found an important region involved in the basal and drug-induction regulation of C YP1A5,the promoter upstream-311 bp~-250 bp region.Combined promoter mutation analysis with Mat Inspector,we found the key transcription factors in the basal regulation which mediate the drug-induction effect of C YP1A5,aryl hydrocarbon receptor(AHR).Overexpression of transcription factor AHR can increase the expression of CYP1A5 at the transcriptional level and increase the activity of the promoter.We validated that the CYP1A5 transcriptional level was associated with AHR via AHR inhibitor and si RNAs.In conclusion,this study found that T-2 toxin can specifically up-regulate the expression of chicken CYP1A4 and C YP1A5 at the transcriptional level,which is related to the acetyl group at the 15-position C atom.Moreover,we also demonstratedthat the transcription factor AHR was involved in the basal and drug-induction regulation of CYP1A5.