Differential Expression of Small RNAs and mRNAs in Bovine Skeletal Muscle with Extreme Saturated Fatty Acid Levels

Understanding the genetic control of fatty acid metabolism may allow for the production of healthier beef.  Twelve extreme skeletal muscle samples (high =6; low =6) for saturated to unsaturated fatty acid ratio were analyzed using Illumina RNA-seq technology to identify differentially expressed (DE) mRNAs.  The same samples, including an additional 10 samples (N=22), were also sequenced using the Illumina small RNA library preparation kit.  Sequencing runs were checked for quality parameters using FASQC and mapped to the UMD 3.1 reference genome using Bowtie2.  Aligned reads were quantitated using HTseq (small RNAs) or Cufflinks (mRNA).  Statistical analyses were conducted using the QuasiSeq R package (small RNAs) or a two-stage Poisson/ Quasi-Poisson model (mRNAs), where animal age, contemporary group and sex were included as covariates.  The residual saturated fatty acid ratio was also included as a covariate to determine it’s impact on differential expression of transcripts.  A total of 90 mRNAs were identified as DE (q < 0.15).  A total of 176 small RNAs were identified as DE (q < 0.15).  The small RNAs included 21 miRNA, 4 miscRNA, 11 snoRNAs, 5 snRNA, 3 unannotated small RNAs, and 132 5s rRNAs.   Targets of the DE miRNAs were identified using the expressed genes from the RNA-seq study and miRANDA software.  Enrichment analysis with DAVID software identified targets of mir-628 as enriched for terms related to fatty acid metabolism (Benjamini p< 0.10).  These results indicate that small RNA levels may play an important role in skeletal muscle fatty acid metabolism.