Identification of crucial modules and genes associated with backfat tissue development by WGCNA in Ningxiang pigs

Abstract

Fat deposition is an economically important trait in pigs. Ningxiang pig, one of the four famous indigenous breeds in China, is characterized by high fat content. The underlying gene expression pattern in different developmental periods of backfat tissue remains unclear, and the purpose of this investigation is to explore the potential molecular regulators of backfat tissue development in Ningxiang pigs. Backfat tissue (three samples for each stage) was initially collected from different developmental stages (60, 120, 180, 240, 300, and 360 days after birth), and histological analysis and RNA sequencing (RNA-seq) were then conducted. Fragments per kilobase of transcript per million (FPKM) method was used to qualify gene expressions, and differentially expressed genes (DEGs) were identified. Furthermore, strongly co-expressed genes in modules, which were named by color, were clustered by Weighted gene co-expression network analysis (WGCNA) based on dynamic tree cutting algorithm. Gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) enrichment were subsequently implemented, and hub genes were described in each module. Finally, QPCR analysis was employed to validate RNA-seq data. The results showed that adipocyte area increased and adipocyte number decreased with development of backfat tissue. A total of 1,024 DEGs were identified in five comparison groups (120 days vs. 60 days, 180 days vs. 120 days, 240 days vs. 180 days, 300 days vs. 240 days, and 360 days vs. 300 days). The turquoise, red, pink, paleturquoise, darkorange, and darkgreen module had the highest correlation coefficient with 60, 120, 180, 240, 300, and 360 days developmental stage, while the tan, black and turquoise module had strong relationship with backfat thickness, adipocyte area, and adipocyte number, respectively. Thirteen hub genes (ACSL1, ACOX1, FN1, DCN, CHST13, COL1A1, COL1A2, COL6A3, COL5A1, COL14A1, OAZ3, DNM1, and SELP) were recognized. ACSL1 and ACOX1 might perform function in the early developmental stage of backfat tissue (60 days), and FN1, DCN, COL1A1, COL1A2, COL5A1, COL6A3, and COL14A1 have unignorable position in backfat tissue around 120 days developmental stage. Besides, hub genes SELP and DNM1 in modules significantly associated with backfat thickness and adipocyte area might be involved in the process of backfat tissue development. These findings contribute to understand the integrated mechanism underlying backfat tissue development and promote the progress of genetic improvement in Ningxiang pigs.

Keywords: Ningxiang pig; WGCNA; backfat tissue; different developmental stage; hub gene; lipid metabolism.