鸡肝脏活化黄曲霉毒素B1代谢的关键CYPs基因的转录调控机制研究

Dietary exposure to aflatoxin B1 (AFB1) is harmful to the health and performance of domestic animals. Poultry are relatively sensitive to the toxic actions of AFB1. Previous studies have shown that hepatic cytochrome P450 (CYPs), CYP1A1 and CYP2A6, are primarily enzymes responsible for the bioactivation of AFB1 in chicks. Meanwhile, AFB1 can also induce the high expression of these CYPs genes. However, the transcriptional regulation mechanism of these CYPs genes in the liver of chicks during AFB1 metabolism is remained unknown. This project intends to use bioinformatics, point (or deletion) mutation, luciferase reporter gene and chromatin immunoprecipitation (ChIP) assay, and RNA interference (RNAi) technologies to identify the pivotal cis-acting elements and transcription factors in the promoter area of CYP1A1 and CYP2A6. Moreover, the function of these identified transcription factors involved in highly upregulation of the CYPs and hepatotoxity by AFB1 will be analyzed by RNAi technology in chick primary hepatocytes. Furthermore, we will explore whether the nutrients could alleviate aflatoxicosis in chicks through the regulation of the crucial transcription factors of these CYPs. These studies will reveal the transcriptional regulation mechanism of these crucial CYPs genes in chick liver during AFB1 metabolism, which can provide the new regulatory targets for the development of nutritional strategies to prevent aflatoxicosis in chicks.

黄曲霉毒素B1（AFB1）对畜禽生产危害极大，家禽对其尤为敏感。前人和申请人研究表明鸡肝脏中细胞色素P450（CYPs）酶系CYP1A1和CYP2A6在活化AFB1毒性代谢中起关键作用，同时，AFB1还能促进上述CYPs基因的高表达。但是，上述CYPs基因在鸡肝脏AFB1代谢中的转录调控机制尚不清楚。本项目拟应用生物信息学、点（或缺失）突变、双荧光素酶报告基因、染色质免疫沉淀（ChIP）和RNA干扰等技术在鸡肝细胞中鉴定上述CYPs基因启动子区的关键顺式作用元件及与其互作的转录因子；然后，应用RNA干扰等技术验证上述关键转录因子在AFB1促进上述CYPs基因高表达及致鸡原代肝细胞损伤中的作用；最后，探究营养素通过调控上述CYPs基因关键转录因子拮抗AFB1致鸡肝毒性的可行性。本研究旨在阐明鸡肝脏活化AFB1代谢的关键CYPs基因的转录调控机制，可为营养素防治鸡AFB1中毒提供新的调控靶点。

黄曲霉毒素B1（AFB1）对畜禽生产危害极大，家禽对其尤为敏感。研究表明鸡肝脏中细胞色素P450（CYPs）酶系CYP1A1和CYP2A6在活化AFB1毒性代谢中起关键作用，同时，AFB1还能促进上述CYPs基因的高表达。但是，上述CYPs基因在鸡肝脏AFB1代谢中的转录调控机制尚不清楚。本项目应用生物信息学、点（或缺失）突变、双萤光素酶报告基因、染色质免疫沉淀（ChIP）和圈套寡核苷酸等技术在鸡肝细胞中鉴定出了CYP1A1及CYP2A6基因启动子区的关键转录因子主要为Sp1和Ap-1。同时，发现AFB1促进鸡肝细胞CYP2A6基因的表达，与其激活CYP2A6基因的转录因子Ap-1的表达相关。然而，AFB1促进鸡肝细胞CYP1A1基因的表达与其调控转录因子Sp1和Ap-1的表达无关。本研究阐明了鸡肝脏活化AFB1代谢的关键基因CYP1A1和CYP2A6的转录调控机制，可为营养素防治鸡AFB1中毒提供新的调控靶点。