Effects of Overexpression of BCAT1 Gene on Milk Protein Synthesis in Mammary Epithelial Cells of Dairy Goats (Capra hircus)
SONG Yi-Min1, LI Hong-Qiang1, WANG Dong-Xian1, LIU Si-Jiang1, GUO Zi-Long1, LUO Yu2, LI Xiao-Yi-Yan1, SHAO Yue-Xin1, LIU Yan-Xin2, SHI Huai-Ping3, LIU Zheng-Zhu1,*, ZHANG Hui-Wen2,*
1 College of Animal Science and Technology/Hebei Key Laboratory of Specialty Animal Germplasm Resources Exploration and Innovation, Hebei Normal University of Science & Technology, Qinhuangdao 066600, China; 2 Chengde Veterinary Drug Administration, Chengde 067032, China; 3 College of Animal Science and Technology, Northwest A&F University, Yangling 712100, China
Abstract：Branched-chain amino acid aminotransferase 1 (BCAT1) is a key enzyme in the catabolism of branched chain amino acids (BCAAs). This study aimed to obtain the CDS sequence of the BCAT1 gene and preliminarily investigate the effect of this gene on milk protein synthesis in mammary epithelial cells of dairy goats (Capra hircus). Mammary epithelial cells of dairy goats were used as materials, the CDS region of the BCAT1 gene was cloned, and bioinformatics analysis of the CDS sequence was performed using online software. The pcDNA3.1-BCAT1 overexpression vector was constructed, and the effect of the BCAT1 gene on milk protein synthesis in mammary epithelial cells was explored. The results showed that the full-length CDS region of the BCAT1 gene was 1 161 bp, encoding 386 amino acids. The molecular weight of the BCAT1 protein was 43 173.42 D, and the theoretical pI value was 5.05. No transmembrane domain or signal peptide was detected, and it was identified as a negatively charged, hydrophobic and stable protein. Protein-protein interaction analysis showed that the BCAT1 protein interacted with branched chain keto acid dehydrogenase E1 subunit alpha (BCKDHA), BCKDHB and other proteins. Tissue expression profile analysis indicated that the expression level of the BCAT1 gene was relatively high in mammary gland tissue during the dry period. After transfection of the BCAT1 gene recombinant overexpression vector into mammary epithelial cells, the expression levels of milk protein-related genes, including beta-lactoglobulin (BLG) (P<0.01), casein alpha s1 (CSN1S1) (P<0.05), casein beta (CSN2) (P<0.05), and casein kappa (CSN3) (P<0.05), were significantly increased. It was indicated that the BCAT1 gene played a regulatory role in milk protein synthesis in goat mammary epithelial cells. This study provides a theoretical foundation for further research on the regulatory mechanism of the BCAT1 gene in goat milk protein synthesis.
宋伊敏, 李宏强, 王东贤, 刘泗江, 郭子龙, 罗妤, 李肖依颜, 邵钺馨, 刘艳新, 史怀平, 刘铮铸, 张会文. 过表达BCAT1基因对奶山羊乳腺上皮细胞乳蛋白合成的影响[J]. 农业生物技术学报, 2026, 34(9): 1940-1949.
SONG Yi-Min, LI Hong-Qiang, WANG Dong-Xian, LIU Si-Jiang, GUO Zi-Long, LUO Yu, LI Xiao-Yi-Yan, SHAO Yue-Xin, LIU Yan-Xin, SHI Huai-Ping, LIU Zheng-Zhu, ZHANG Hui-Wen. Effects of Overexpression of BCAT1 Gene on Milk Protein Synthesis in Mammary Epithelial Cells of Dairy Goats (Capra hircus). 农业生物技术学报, 2026, 34(9): 1940-1949.
[1] 李玲. 2009. CSN1S2、CSN2、CSN3基因在西农萨能奶山羊乳腺组织中的表达分析[D]. 硕士学位论文, 西北农林科技大学, 导师: 曹斌云, pp. 41-42. (Li L.2009. Expression analysis of CSN1S2, CSN2 and CSN3 genes in mammary gland tissue of Xinong Saanen dairy goat[D]. Master's thesis, Northwest A&F University, Supervisor: Cao B Y, pp. 41-42. ) [2] 潘坛, 吕明, 罗军, 等. 2019. 西农萨能奶山羊TRIB3基因克隆及RNA干扰分析[J]. 农业生物技术学报, 27(08): 1392-1400. (Pan T, Lv M, Luo J, et al.2019. Cloning and RNA interference analysis of TRIB3 gene in Xinong Saanen dairy goat[J]. Journal of Agricultural Biotechnology, 27(08): 1392-1400.) [3] 石新芳, 王育民, 张凯秀, 等. 2023. 沉默BCAT1对大鼠腹腔感染性脓毒症急性肾损伤的干预作用[J]. 中华医院感染学杂志, 33(23): 3527-3531. (Shi X F, Wang Y M, Zhang K X, et al.2023. Intervention effect of silencing BCAT1 on acute kidney injury in rats with abdominal infectious sepsis[J]. Chinese Journal of Nosocomiology, 33(23): 3527-3531.) [4] 谭力力, 卢杰. 2025. BCAT1通过调节巨噬细胞泡沫化影响老年动脉粥样硬化[J]. 沈阳医学院学报, 27(05): 507-513. (Tan L L, Lu J.2025. BCAT1 affects atherosclerosis in the elderly by regulating macrophage foam cell formation[J]. Journal of Shenyang Medical College, 27(05): 507-513.) [5] 唐继鹏. 2025. 新型亚临床低钙血症对奶牛代谢、健康和生产性能的影响及其血清蛋白组学分析[D]. 硕士学位论文, 黑龙江八一农垦大学, 导师: 夏成, pp. 43-44. (Tang J P.2025. Effects of novel subclinical hypocalcemia on metabolism, health and production performance of dairy cows and its serum proteomics analysis[D]. Thesis for M.S., Heilongjiang Bayi Agricultural University, Supervisor: Xia C, pp. 43-44.) [6] 王影. 2022. 荷斯坦奶牛高、低乳脂差异代谢物筛选与PDGFD基因功能验证[D]. 硕士学位论文, 宁夏大学, 导师: 顾亚玲, pp. 22-29. (Wang Y.2022. Screening of differential metabolites in high and low milk fat Holstein cows and functional verification of PDGFD gene[D]. Thesis for M.S. Ningxia University, Supervisor: Gu Y L, pp. 22-29.) [7] 张幸开, 袁耀明. 2006. 影响奶牛乳蛋白生产的因素及应对措施[J]. 中国奶牛,(12): 5. (Zhang X K, Yuan Y M. 2006. Factors affecting milk protein production in dairy cows and countermeasures [J]. China Dairy Cattle, (12): 5.) [8] 宗学阳. 2023. 奶山羊β-酪蛋白及其消化产物的功能研究[D]. 硕士学位论文, 西北农林科技大学, 导师: 史怀平, pp. 22. (Zong X Y. 2023. Functional study on β-casein and its digestive products in dairy goats[D].Thesis for M.S., Northwest A&F University, Supervisor: Shi H P, pp. 4.) [9] Amalfitano N, Macedo Mota L F, Rosa G M, et al.2022. Role of CSN2, CSN3, and BLG genes and the polygenic background in the cattle milk protein profile[J]. Journal of Dairy Science, 105(7): 6001-6020. [10] Bruhat A, Chérasse Y, Chaveroux C, et al.2009, Amino acids as regulators of gene expression in mammals: Molecular mechanisms[J]. BioFactors, 35(3): 249-257. [11] Goto M, Shinno H, Ichihara A.1977. Isozyme patterns of branched-chain amino acid transaminase in human tissues and tumors[J]. Japanese Journal of Cancer Research, 68(5): 663-667. [12] Han L, Dong L, Leung K, et al.2023. METTL16 drives leukemogenesis and leukemia stem cell self-renewal by reprogramming BCAA metabolism[J]. Cell Stem Cell, 30(1): 52-68.e13. [13] Harris R A, Hawes J W, Popov K M, et al.1997. Studies on the regulation of the mitochondrial α-ketoacid dehydrogenase complexes and their kinases[J]. Advances in Enzyme Regulation, 37: 271-293. [14] Huang L, Li G, Zhang Y, et al.2025. Small-molecule targeting BCAT1-mediated BCAA metabolism inhibits the activation of SHOC2-RAS-ERK to induce apoptosis of triple-negative breast cancer cells[J]. Journal of Advanced Research, 75: 723-738. [15] Lemosquet S, Guinard-Flament J, Raggio G, et al.2010. How does increasing protein supply or glucogenic nutrients modify mammary metabolism in lactating dairy cows[C]. Energy and Protein Metabolism and Nutrition. Wageningen Academic Publishers, 127: 175-186. [16] Moioli B, Pilla F, Tripaldi C.1998. Detection of milk protein genetic polymorphisms in order to improve dairy traits in sheep and goats: A review[J]. Small Ruminant Research, 27(3): 185-195. [17] Mu T, Hu H, Feng X, et al.2022. Screening and conjoint analysis of key lncRNAs for milk fat metabolism in dairy cows[J]. Frontiers in Genetics, 13: 772115. [18] Nichols K, Dijkstra J, Van Laar H, et al.2019. Expression of genes related to energy metabolism and the unfolded protein response in dairy cow mammary cells is affected differently during dietary supplementation with energy from protein and fat[J]. Journal of Dairy Science, 102(7): 6603-6613. [19] Nongonierma A B, Fitzgerald R J.2016. Strategies for the discovery, identification and validation of milk protein-derived bioactive peptides[J]. Trends in Food Science & Technology, 50: 26-43. [20] Pimentel E C, Bauersachs S, Tietze M, et al.2011. Exploration of relationships between production and fertility traits in dairy cattle via association studies of SNPs within candidate genes derived by expression profiling[J]. Animal Genetics, 42(3): 251-262. [21] Qian L, Li N, Lu X-C, et al.2023. Enhanced BCAT1 activity and BCAA metabolism promotes RhoC activity in cancer progression[J]. Nature Metabolism, 5(7): 1159-1173. [22] Sawicka-Zugaj W, Chabuz W, Barłowska J, et al.2025. Analysis of the frequency of the A1 and A2 alleles in the beta-casein gene and the A, B and E alleles in the kappa-casein gene in local cattle breeds: Polish red and polish white-backed[J]. International Journal of Molecular Sciences, 26(5): 2212. [23] Shekar P C, Goel S, Rani S D, et al.2006. Kappa-casein-deficient mice fail to lactate[J]. Proceedings of the National Academy of Sciences of the USA, 103(21): 8000-8005. [24] Sivanand S, Vander Heiden M G.2020. Emerging roles for branched-chain amino acid metabolism in cancer[J]. Cancer Cell, 37(2): 147-156. [25] Treweek T M, Thorn D C, Price W E, et al.2011. The chaperone action of bovine milk alpha(S1)-and alpha(S2)-caseins and their associated form alpha(S)-casein[J]. Archives of Biochemistry and Biophysics, 510(1): 42. [26] Webb L A, Sadri H, Von Soosten D, et al.2019. Changes in tissue abundance and activity of enzymes related to branched-chain amino acid catabolism in dairy cows during early lactation[J]. Journal of dairy science, 102(4): 3556-3568. [27] Zhang Q, Calus M P L, Bosse M, et al.2018. Human-mediated introgression of haplotypes in a modern dairy cattle breed[J]. Genetics, 209(4): 1305-1317.
