西农萨能奶山羊ATF4基因克隆及其功能初步研究

doi:10.11843/j.issn.0366-6964.2023.06.015

Abstract

Abstract: As a binding protein of ATF/cAMP response element, Transcriptional activator 4 (ATF4) play a pivotal role in mammalian growth and development. The present study aimed to obtain the ATF4 gene sequence of Xinong Saanen dairy goats (Capra hircus), elucidate its structure and investigate its function on lipid metabolism and milk protein synthesis through siRNA interference of the ATF4 gene in goat mammary epithelial cells(GMECs). In order to clarify the expression differences of the ATF4 in different tissues of Xinong Saanen dairy goats, the breast tissues of Xinong Saanen dairy goats at different lactation stages were collected, and the CDS region of the ATF gene was cloned and analyzed by bioinformatics and tissue expression profile. The results showed that the complete CDS region of the ATF4 gene was 1 059 bp, encoding 352 amino acids, the ATF4 protein is acidic protein without transmembrane structure, with a molecular weight of 38.545 24 ku, a theoretical isoelectric point of 4.61, and 35 serine phosphorylation sites. The advanced structure of ATF4 protein was composed of α-helix, extension chain irregularly coiled and β-turn were 39.77%, 7.10%, 51.14% and 1.99%, respectively. ATF4 interacted with CEBPB, CEBPG, ATF3, FOSL2, ATF1, DDIT3, TRIB3, BTRC, CREB1 and DISC. Homology analysis showed that sequence homologies of Xinong Saanen dairy goat with sheep (Ovis aries) and cattle (Bos taurus) were 97.45% and 91.48% respectively. Tissue expression analysis revealed that the highest expression of ATF4 gene was in the rumen, followed by muscle. The expression level of the ATF4 gene was the highest in early lactation, followed by that in peak lactation. After interfering with ATF4 gene expression in mammary epithelial cells of dairy goats, the results showed that mRNA levels of FASN, FABP3 and BLG were down-regulated significantly. Moreover, the mRNA levels of PPARA, PPARG and LALBA were significantly up-regulated. The above results suggest that ATF4 might be involved in the regulation of lipid metabolism and milk protein synthesis in GMECs. The present study provides basic data for further investigation of the molecular mechanism of ATF4 on milk fat metabolism and milk protein synthesis in dairy goats.

Key words: ATF4, gene cloning, milk fat metabolism, milk protein, Xinong Saanen dairy goat

CLC Number:

S827.2

SHAO Yuexin, ZHANG Xinyu, GE Liyan, SHI Huaiping. Cloning and RNA Interference Analysis of ATF4 Gene in Xinong Saanen Dairy Goat[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(6): 2353-2364.

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Cloning and RNA Interference Analysis of ATF4 Gene in Xinong Saanen Dairy Goat

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