Whole-genome Porcine RNA Editing Sites Identification And Difference Between The High And Low Backfat Pigs

RNA editing is an important posttranscriptional modification mechanism that increases the diversity of transcripts,with potential functional consequences,such as recoding amino acids,affecting alternative splicing,influencing RNA stability,and modulating the nuclear retention of RNAs.However,our knowledge of RNA editing in swine is far less than that in humans and mice.In this study,RNA editing sites were detected in seven tissues of pigs based on matched strand-specific RNA sequencing data and whole-genome resequencing data.Differential RNA editing analysis was carried out between three full-sib pairs with opposite backfat thickness phenotypes.In addition,ADAR1 and ADAR2 cDNAs were cloned by rapid-amplification of cDNA ends(RACE).1.Identification of RNA editing sites in seven porcine tissuesWe utilized RES-Scanner to identify RNA editing sites in the brain,subcutaneous fat,heart,liver,muscle,lung and ovary from three pigs based on matched strand-specific RNA sequencing and whole-genome resequencing data.In total,we identified 74863 RNA editing sites,and 92.1% of which caused adenosine-to-guanosine(A-to-G)conversion.Most of the A-to-G editing sites were located in noncoding regions.Further analyses found that 97.4% of the A-to-G editing sites were located in repeat sequences,and 88.6% of which occurred in PRE-1.The expression levels of ADAR1 and ADAR2 were significantly correlated with the number and the global editing level of A-to-G editing sites.The number of A-to-G editing sites ranged from 4155(muscle)to 25001(brain)across the seven tissues.Functional enrichment analyses of the genes with tissue-specific editing sites in each tissue showed that these genes were significantly enriched in biological processes and pathways related to their respective tissue functions.2.Differential RNA editing analysis between high and low backfat pigsRES-Scanner and REDItools were used to identify RNA editing sites based on the whole genome and transcriptome sequencing data of the high and low groups composed of three full-sib pairs with opposite backfat phenotypes.A total of 439 intergroup differential RNA editing sites were found in 334 genes.Functional enrichment analysis showed that these genes were significantly enriched into biological processes and signaling pathways related to adipose deposition.Among the 439 differential RNA editing sites,three sites could lead to amino acid changes and seven sites could regulate gene expression by affect the interaction of miRNA and mRNA.These ten sites are located in nine genes,and six of which are potentially related to adipose deposition.3.Cloning and analysis of ADAR1 and ADAR2The full-length cDNA sequences of RNA editing enzymes ADAR1 and ADAR2 were cloned by RACE.The length of ADAR1 and ADAR2 cDNAs was 6259 bp and 6305 bp.A 47 bp insertion possibly caused by a self-editing site was found in ADAR2 cDNA.A complementary sequence of the 47 bp insertion plus an additional adjacent 70 nucleotides were identified in intron 4.The DNA genotype of this editing site was identical across different pig species,and its editing level first increased and then decreased with individual development in multi-tissues.This study systematically identified and analyzed RNA editing in different tissues of pigs,which largely extends the profile of RNA editing sites and enhances our understanding of RNA editing in swine.The findings of this research provide a foundation for further investigating the regulation mechanism of RNA editing and its function in adipose deposition.