RNA-Seq and Pathway Analysis of Chicken Breast Muscle Revealed Important Genes Associated with Metabolism Under Heat Stress

Heat stress is a major problem for the poultry industry; it affects broiler performance decreasing growth rate, increasing mortality, and causing negative changes in meat quality attributes. This research aims to identify genes and biological pathways associated with heat stress in the breast muscle of broilers using the technology of total RNA next generation sequencing (RNA-Seq). At 28 days of age, the chickens were assigned to two groups: heat stress (32 °C) and thermo neutral (24 °C). Four samples of Pectoralis major muscle per treatment collected 16 days after the start of heat stress were used for RNA-Seq. Total RNA was extracted from muscle samples; the libraries preparation was carried out using the TruSeq Stranded mRNA LT Sample Prep Kit and sequenced using the HiSeq2500 equipment. The quality of raw reads was analyzed with FastQC software; the reads were mapped to the Gallus gallus-4.75 reference genome by Bowtie2 and TopHat2 softwares. To obtain raw reads counts mapped for each annotated gene the HTSeq-count package was employed. Comparisons between treatment groups were made using the DESeq2 software, a package of R/Bioconductor. A total of 5,590 DEG were identified (5% FDR) between the two groups, from which 2,741 were up-regulated and 2,849 were down-regulated. Ingenuity Pathways Analysis (IPA) revealed important pathways related to metabolism such as Apoptosis, mTOR, and PPAR signaling, and also the Regulation of Cellular Mechanics by Calpain Protease. The mTOR signaling Pathway may mediate most of the negative impact of heat stress on growth performance, given its important role in mass accumulation and responsiveness to environmental cues.