Long noncoding RNAs (lncRNAs) regulate an array of biological processes in cells and organ systems. Less is known about their expression and function in lymphocyte lineages. Here we have identified >2000 lncRNAs expressed in human T cell cultures and those which display a TH lineage specific pattern of expression and are intragenic or adjacent to TH lineage specific genes encoding proteins with immunologic functions. One lncRNA cluster selectively expressed by the effector TH2 lineage consists of four alternatively spliced transcripts that regulate expression of TH2 cytokines, IL-4, IL-5 and IL-13. Genes encoding this lncRNA cluster in humans overlap the RAD50 gene and thus are contiguous with the previously described TH2 locus control region (LCR) in the mouse. Given its genomic synteny with the TH2 LCR, we refer to this lncRNA cluster as TH2-LCR lncRNA.

Objective. We sought to determine interrelationships among expression of lincRNA-p21, a long intergenic non-coding RNA, activity of NF-κB, and responses to methotrexate in rheumatoid arthritis (RA) by analyzing patient samples and cell culture models. Methods. Expression levels of long non-coding RNAs and messenger RNAs were determined by quantitative reverse transcription-polymerase chain reaction. Western blotting and flow cytometry were used to quantify levels of intracellular proteins. Intracellular NF-κB was determined using an NF-κB luciferase reporter plasmid. Results. RA patients expressed reduced basal levels of lincRNA-p21 and increased basal levels of phosphorylated p65 (RelA), a marker of NF-κB activation. RA subjects not receiving MTX expressed lower levels of lincRNA-p21 and higher levels of phosphorylated p65 compared to RA subjects receiving low-dose MTX. In cell culture using primary cells and transformed cell lines, we found that MTX induced lincRNA-p21 through a DNA-PKcs-dependent mechanism. Deficiencies of PRKDC mRNA levels in RA subjects were also corrected by MTX, in vivo. Further, MTX lowered NF-κB activity in TNF-α treated cells through a DNA-PKcs-dependent mechanism via induction of lincRNA-p21. Finally, we found that depressed levels of TP53 and lincRNA-p21 increased NF-κB activity in cell lines. Decreased levels of lincRNA-p21 did not alter NFKB1 or RELA transcripts. Rather, lincRNA-p21 physically bound to RELA mRNA. Conclusion. Our findings support a model whereby depressed levels of lincRNA-p21 in RA contribute to increased NF-κB activity. MTX decreases basal levels of NF-κB activity by increasing lincRNA-p21 through a DNA-PKcs dependent mechanism.

Long non-coding RNAs (lncRNAs) possess a diverse array of regulatory functions including activation and silencing of gene transcription, regulation of splicing, and coordinating epigenetic modifications. GATA3-AS1 is a divergent lncRNA gene neighboring GATA3. GATA3 is considered the master regulator of TH2 lineage commitment enabling TH2 effector cells to efficiently transcribe genes encoding cytokines IL-4, IL-5, and IL-13. Here, we show that the GATA3-AS1 lncRNA is selectively expressed under TH2 polarizing conditions and is necessary for efficient transcription of GATA3, IL5, and IL13 genes, while being sufficient for GATA3 transcription. GATA3-AS1 is required for formation of permissive chromatin marks, H3K27 acetylation and H3K4 di/tri-methylation, at the GATA3-AS1-GATA3 locus. Further, GATA3-AS1 binds components of the MLL methyltransferase and forms a DNA-RNA hybrid (R-loop) thus tethering the MLL methyltransferase to the gene locus. Our results indicate a novel regulatory function for a divergent lncRNA and provide new insight into the function of lncRNAs in T helper cell differentiation.

Long noncoding RNAs (lncRNAs) represent a newly discovered class of regulatory molecules that impact a variety of biological processes in cells and organ systems. In humans, it is estimated that there may be more than twice as many lncRNA genes than protein-coding genes. However, only a handful of lncRNAs have been analyzed in detail. In this review, we describe expression and functions of lncRNAs that have been demonstrated to impact innate and adaptive immunity. These emerging paradigms illustrate remarkably diverse mechanisms that lncRNAs utilize to impact the transcriptional programs of immune cells required to fight against pathogens and maintain normal health and homeostasis.

Genome-wide association studies have identified numerous genetic variants conferring autoimmune disease risk. Most of these genetic variants lie outside protein-coding genes hampering mechanistic explorations. Numerous mRNAs are also differentially expressed in autoimmune disease but their regulation is also unclear. The majority of the human genome is transcribed yet its biologic significance is incompletely understood. We performed whole genome RNA-sequencing [RNA-seq] to categorize expression of mRNAs, known and novel long non-coding RNAs [lncRNAs] in leukocytes from subjects with autoimmune disease and identified annotated and novel lncRNAs differentially expressed across multiple disorders. We found that loci transcribing novel lncRNAs were not randomly distributed across the genome but co-localized with leukocyte transcriptional enhancers, especially super-enhancers, and near genetic variants associated with autoimmune disease risk. We propose that alterations in enhancer function, including lncRNA expression, produced by genetics and environment, change cellular phenotypes contributing to disease risk and pathogenesis and represent attractive therapeutic targets.