Identification and analysis of the expression of microRNA from lactating and nonlactating mammary glands of the Chinese swamp buffalo

Abstract

To study the role of microRNA (miR) in the lactation physiology of water buffalo, 2 multiparous dairy buffaloes (including an 8-yr-old buffalo that had been lactating for 3 mo, as well as a 10-yr-old nonlactating, nonpregnant buffalo) were used for miR library construction. The profile of differentially expressed miR in lactating and nonlactating mammary gland tissues of these water buffalo were investigated using Illumina-Solexa high-throughput sequencing technology (Illumina, San Diego, CA). The data identified 259 miR families, 359 mature miR, 363 pre-miR, 230 novel buffalo miR, and 5 buffalo-specific miR that were expressed in mammary tissues. Some highly significantly differentially expressed miR were explored, including bbu-miR-497, bbu-miR-30a-5p, bbu-miR-148a, bbu-miR-29a, bbu-miR-125a, bbu-miR-125b, and bbu-miR-103. The expression patterns of 18 miR were confirmed by quantitative real-time PCR in both tissues, and the expression of bbu miR-103 and novel miR-57 constituted the largest differences between lactating and nonlactating tissues. Further functional analysis indicated that the overexpression or suppression of miR-103 in buffalo mammary epithelial cells downregulated or upregulated the expression of pantothenate kinase 3, and also significantly increased the transcription factor steroid regulatory element binding protein, followed by the acceleration of de novo synthesis of fatty acids by upregulation of acetyl coenzyme A carboxylase α expression. The expression levels of 34 predicted target genes of novel-miR-57 in lactating and nonlactating mammary gland tissues were all analyzed by quantitative real-time PCR. Finally, only the expression of docking protein 4 could be upregulated or downregulated selectively by bbu-novel-miR-57 in buffalo mammary epithelial cells and the Bcap-37 cell line. This study provides an overview of the miR expression profile of water buffalo and the interaction between some key miR and their target genes, which may improve understanding of the important roles of miR in buffalo milk fat synthesis.