Long non-coding RNAs (lncRNAs) are key regulators of gene expression, playing crucial roles in biological processes across many species, tissues and diseases. The liver is a highly responsive organ in which large changes in gene expression are perpetuated by a myriad of internal and external stimuli; as such, the liver makes an ideal system in which to study lncRNAs. Global patterns of expression, maturation and localization were established for both lncRNA and protein-coding gene (PCG) transcripts across five subcellular compartments in male and female mouse liver, both with and without exposure to TCPOBOP, a direct agonist of the nuclear receptor CAR. In contrast to PCGs, lncRNAs showed very strong enrichment for tight chromatin binding, which increased the sensitivity for lncRNA detection and facilitated discovery of many novel sex-biased and xenobiotic-responsive lncRNAs. These findings helped identify candidate regulatory lncRNAs based on their co-localization within topologically associating domains, or their transcription divergent or antisense to PCGs associated with pathways linked to liver physiology and disease. The liver cell type-specific expression of lncRNAs and PCGs was assessed by single nucleus RNA-seq (snRNA-seq). Liver sexual dimorphism was largely restricted to hepatocyte populations, where many sex-biased genes exhibited zonated expression. Changes in lncRNA and PCG expression following exposure to endogenous hormones (growth hormone) and exogenous chemicals (TCPOBOP) was assessed, identifying cell cluster-specific perturbations to native sex-bias and hepatocyte zonation-dependent gene expression, and highlighting the interconnectedness between liver sexual dimorphism and zonation of the hepatic lobule at the single nuclei level. Finally, an in vivo method for epigenetic reprogramming of lncRNAs using a dual adeno-associated virus delivery system was utilized to knockdown two TCPOBOP-inducible lncRNAs in mouse liver. The knockdown phenotype of one of these lncRNAs, established by snRNA-seq, suggests it plays a functional role in regulating cholesterol metabolism and transport, triglyceride catabolism, and pyruvate metabolism in mouse liver. Together, these studies characterize hepatic lncRNA expression patterns, on both the sub-cellular and single cell levels, and present a strategy for interrogating the roles of specific lncRNAs in liver tissue in vivo. / 2023-01-18T00:00:00Z