Hepatic Long Intergenic Noncoding RNAs: High Promoter Conservation and Dynamic, Sex-Dependent Transcriptional Regulation by Growth Hormone

Abstract

Long intergenic noncoding RNAs (lincRNAs) are increasingly recognized as key chromatin regulators, yet few studies have characterized lincRNAs in a single tissue under diverse conditions. Here, we analyzed 45 mouse liver RNA sequencing (RNA-Seq) data sets collected under diverse conditions to systematically characterize 4,961 liver lincRNAs, 59% of them novel, with regard to gene structures, species conservation, chromatin accessibility, transcription factor binding, and epigenetic states. To investigate the potential for functionality, we focused on the responses of the liver lincRNAs to growth hormone stimulation, which imparts clinically relevant sex differences to hepatic metabolism and liver disease susceptibility. Sex-biased expression characterized 247 liver lincRNAs, with many being nuclear RNA enriched and regulated by growth hormone. The sex-biased lincRNA genes are enriched for nearby and correspondingly sex-biased accessible chromatin regions, as well as sex-biased binding sites for growth hormone-regulated transcriptional activators (STAT5, hepatocyte nuclear factor 6 [HNF6], FOXA1, and FOXA2) and transcriptional repressors (CUX2 and BCL6). Repression of female-specific lincRNAs in male liver, but not that of male-specific lincRNAs in female liver, was associated with enrichment of H3K27me3-associated inactive states and poised (bivalent) enhancer states. Strikingly, we found that liver-specific lincRNA gene promoters are more highly species conserved and have a significantly higher frequency of proximal binding by liver transcription factors than liver-specific protein-coding gene promoters. Orthologs for many liver lincRNAs were identified in one or more supraprimates, including two rat lincRNAs showing the same growth hormone-regulated, sex-biased expression as their mouse counterparts. This integrative analysis of liver lincRNA chromatin states, transcription factor occupancy, and growth hormone regulation provides novel insights into the expression of sex-specific lincRNAs and their potential for regulation of sex differences in liver physiology and disease.

Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.00861-15.

ACKNOWLEDGMENTS

We thank the following laboratory members who contributed tissue samples and/or RNA-Seq samples: Nicholas J. Lodato, Dana Lau-Corona, Alexander Suvorov, Jeannette Connerney, and Tara L. Conforto. We also thank Gracia Bonilla, Andy Rampersaud, and George F. Steinhardt IV of our laboratory for initial ChIP-seq peak discovery for (i) FOXA1 and FOXA2 , (ii) BCL6, CEBPA, and CEBPB, and (iii) HNF6, respectively.

We declare that we have no competing interests.

T.M. conceived of the study jointly with D.J.W., carried out all of the computational and bioinformatics analysis, and drafted the manuscript together with D.J.W. P.H. participated in early-stage experimental and computational efforts to identify sex-specific lincRNAs and contributed to the revision of the manuscript. F.Y. carried out the qPCR studies and associated data analysis. D.J.W. contributed to study design, coordinated the overall project, helped T.M. draft the manuscript, and edited the manuscript. We all read and approved the final manuscript.