LNCing RNA to Immunity

Abstract

Despite an ever-increasing appreciation for how protein-coding genes shape immune responses, the molecular underpinnings of immune regulation remain incompletely understood. This incomplete picture impedes the development of more precise therapeutics and diagnostics for immune-mediated diseases. Long noncoding RNAs (lncRNA) are versatile cell- and context-specific regulators of gene expression and cellular function. The number of lncRNA genes rivals that of protein-coding genes; however, comparatively little is known about their function. Even though the functions of most lncRNA genes are unknown, multiple lncRNAs have recently emerged as important immune regulators. Therefore, further unlocking the role of lncRNAs in the immune system coupled with their tissue-specific expression might lead to more precise therapeutics and diagnostics for immune disorders in general.

LncRNAs are Cell- and Context-Specific Immune Regulators

The mammalian immune system has a powerful capacity to protect and promote healing. However, when dysregulated, it can also cause damage manifesting as inflammatory and autoimmune diseases. Current approaches to enhance or inhibit immune responses are imprecise and have many side effects. A richer and more precise understanding of immune response regulation may reveal more specific therapeutic targets.

Long noncoding RNAs (see Glossary) are a subclass of regulatory noncoding RNAs (ncRNAs) that do not code for proteins^1-4. They are distinct from housekeeping ncRNAs and are arbitrarily divided into small (less than 200 nucleotides) and long (greater than 200 nucleotides) noncoding RNAs^5,6. Previously thought to represent merely “aberrant” transcription¹, lncRNAs are now known to impact a wide array of cellular and physiologic processes including the cell cycle, metabolism, muscle function, carcinogenesis, and hematopoiesis^2-4. The number of human lncRNA genes rivals that of protein-coding genes with a recent annotation identifying 18,962 protein-coding genes and 25,738 lncRNAs^7-9. Unlike protein-coding genes, the function of only a small fraction of lncRNA genes has been determined^2,10,11. Prediction of lncRNA function is difficult because primary sequence and evolutionary conservation are typically less reliable than for protein coding genes^8,12-14. Further, lncRNAs are often redundant or cross regulate each other making it hard to elucidate a phenotype¹⁵. Regardless, it is useful to describe some lncRNA functions that may inform further studies. In addition, lncRNA have greater cell- and context-specific expression relative to protein-coding genes^12,16,17, which might make lncRNAs useful as potential therapeutic and diagnostic targets for certain immune system disorders and in certain specific instances^18,19.

Here we discuss lncRNA biogenesis (see Box 1), explore the diversity of lncRNA molecular mechanisms (see Box 2), discuss lncRNA conservation (see Box 3), and illustrate how lncRNAs have recently emerged as cell- and context-specific versatile regulators of immune cell development, differentiation, and function (Table 1). We highlight those lncRNAs that have been suggested to regulate protective antimicrobial immune responses as well as pathogenic immune responses such as acute graft-versus-host disease (aGVHD) and systemic lupus erythematosus (SLE)^20,21.

Box 1. LncRNA Biogenesis.

Like protein-coding genes, mammalian lncRNA genes are transcribed by RNA Pol II, spliced, 5’ capped, and 3’ polyadenylated². However, compared with mRNA, lncRNAs generally have lower expression with greater variation, less efficient splicing, shorter length, lower stability, greater nuclear localization, enrichment of transposable elements, and fewer exons^{2,7,8,12,16,17,110,111}. Promoters of lncRNAs have fewer transcription factor binding sites that likely contributes to the greater tissue-specific expression of human lncRNAs¹¹², while active lncRNAs have greater repressive H3K9me3 marks^110,111. The nuclear retention of lncRNAs is mediated by binding to U1 small nuclear ribonucleoprotein¹¹³, while the nuclear export of lncRNAs is similar to that of mRNA¹¹⁴. The most widely used lncRNA classification system is based on the orientation of lncRNAs to protein coding genes¹¹⁵. However, systems based on biogenesis and biochemistry have also been proposed¹⁷.

Box 2. LncRNA Molecular Mechanisms are Diverse.

The molecular mechanisms of lncRNAs are varied and depend on their interactions with partner macromolecules². These interactions occur in both cis or in trans and in multiple subcellular locations. In fact, the subcellular location of lncRNAs is linked to their molecular mechanism¹¹⁵. For example, nuclear lncRNAs tend to regulate transcription whereas those in the cytoplasm influence protein function or regulate gene expression by impacting a variety of regulatory mechanisms such as translation and signal transduction².

The diverse interactions of nuclear-localized lncRNAs with their macromolecular partners regulate gene expression at the epigenetic, transcriptional, and post-transcriptional levels. For example, lncRNAs can interact with and recruit or sequester chromatin modifiers from target-gene promoters (Figure 1A and B)², or form R-loops at target genes, either in cis or trans, resulting in altered expression of protein-coding genes^2,71 (Figure 1C).

Additional ways in which lncRNAs can regulate gene expression include controlling looping of transcriptionally active and inactive chromatin domains^53,116 and organizing membraneless nuclear compartments important for transcription, splicing, and stress-induced-RNA and -protein sequestration^44,117-119. Promoter antisense (PAS) lncRNAs activate ligand-induced, paused, protein-coding genes by recruiting H3K9me3 demethylases via their stem-loop RNA structures that release inhibitory transcription complexes from the adjacent protein-coding gene¹²⁰ (Figure 1D). Intriguingly, a few lncRNAs regulate the expression of target genes independent of the lncRNA transcript. For example, enhancers in the promoters of some lncRNAs are co-transcriptionally activated, which then regulate the transcription of nearby target genes^1,2 (Figure 1E).

Extranuclear lncRNAs interact with a variety of proteins and alter their activity and localization. These lncRNA-protein interactions impact translation, RNA stability, metabolism, genomic stability, and signal transduction^2,6 (Figure 1F-H). For example, the mouse and human lncRNA NORAD induces the liquid-liquid phase separation and sequestration of Pumilio RNA-binding proteins in the cytosol thereby protecting against Pumilio-driven genomic instability¹²¹. Through RNA-RNA interactions, lncRNAs can regulate the stability and translation of mRNAs² or function as micro-RNA “sponges” termed competing endogenous RNAs (ceRNA) (Figure 1I)⁶.

By definition, lncRNAs do not code for proteins. However, systematic analyses demonstrate that some lncRNAs harbor small open reading frames that encode functional micro-peptides^122,123 (Figure 1J). The presence of a functional micro-peptide does not exclude a separate RNA-dependent molecular function of a lncRNA^124-128. For example, the human lncRNA MIR155HG encodes a pro-inflammatory micro-RNA, miR-155, but also encodes a micro-peptide that suppresses autoimmune inflammation by inhibiting antigen presentation in DCs and ameliorates murine experimental autoimmune encephalomyelitis and psoriatic-like skin inflammation¹²⁹.

Box 3. LncRNA Conservation and Function.

Sequence conservation helps identify lncRNAs that are functional¹³⁰. However, lncRNA sequence conservation is poor compared to protein coding genes^8,12. Despite this, lncRNAs with low sequence conservation often retain functional conservation^31,131-133. For instance, two different lncRNAs, Pair in mice and HULC in humans, augment the activity of phenylalanine hydroxylase (PAH, deficiency causes phenylketonuria), and importantly, a lncRNA mimic of the HULC functional region rescues phenylalanine accumulation in Pair^−/− mice demonstrating functional equivalence across species¹³⁴. Micro-peptides may also function across species barriers particularly if the interaction sites on their protein-binding partners are conserved. An example of this is the miro-peptide encoded by human MIR155HG that binds both mouse and human HSP70 to inhibit antigen presentation in DCs¹²⁹. Low sequence conservation with preservation of function may also be a feature of lncRNA-encoded micro-peptides. For example, the SPAR polypeptide encoded by LINC00961 in humans has just 65% amino acid identity with its murine orthologue yet murine SPAR retains its function in human HEK293 cells¹³⁵. Collectively, these studies suggest lncRNA linear nucleotide sequence is a poor predictor of function.

Positional and structural conservation provide import clues to aid in identifying functional lncRNAs when sequence conservation is poor^136,137. Thus, validation of function remains necessary in most if not all cases, which impedes high throughput functional classification of lncRNAs. However, the similar abundance of short RNA motifs, called k-mers, from the linear sequence of lncRNAs may allow for categorization of lncRNAs by analogous function¹³⁸. Altogether, these studies highlight the need for a better understanding of how lncRNA sequence might impart function.

Table 1.

Characteristics of selected immune-regulatory lncRNAs.

lncRNA	Immune Cell Expression	Subcellular Localization	Mouse or Human	In Vivo Role	Ref. #
Innate Immunity
LIMIT	Dendritic cells, Macrophages, and Tumor cells	Nucleus	Both	Promotes anti-tumor immunity and enhances checkpoint inhibitor responses by increasing expression of MHC-I	[104]
LincRNA-Cox2	Macrophages and Dendritic cells	Nucleus	Mouse	Regulates innate PRR pathways	[40]
LincRNA-EPS	Macrophages and Dendritic cells	Nucleus	Mouse	Negatively regulates inflammatory phenotype, protective against endotoxemia and Listeria monocytogenes infection in mice	[41]
Lnc13	Bone marrow derived macrophages	Nucleus	Both	Negatively regulates inflammatory phenotype, decreased expression and hypofunctional polymorphism seen in patients with Celiac disease	[42]
LncHSC1	Hematopoietic stem cells	Nucleus	Mouse	Restrains myeloid lineage commitment	[24]
Lnc-CD56	Natural killer cells	Nucleus	Human	Positively regulates CD56 expression, promotes NK cell development	[54]
LncKdm2b	Group 3 innate lymphoid cells	Nucleus	Mouse	Promotes ILC3 maintenance and proliferation, enhances effector functions of ILC3s, protects against intestinal bacterial infection in a murine model	[51]
LncRNA-GM	Macrophages	Cytoplasm	Both	Promotes antiviral innate immune response, prevents viral infection	[28]
LncRNA-ISIR	Myeloid cells, T cells, and fibroblasts following viral infection	Cytoplasm and nucleus	Both	Protects against viral infection, expression is increased in PBMCs of patients with SLE and correlates with disease severity	[37]
Lnc-UC	Macrophages	Nucleus	Both	Decreases inflammatory signaling, prevents colitis in mouse models	[48]
Lnczc3h7a	Macrophages	Cytoplasm	Mouse	Promotes antiviral innate immune response, prevents viral infection	[36]
LOUP	Myeloid cells	Cytoplasm and nucleus	Both	Induces myeloid differentiation	[29]
NAIL	Macrophages	Nucleus	Both	Expression is increased in patients with ulcerative colitis, exacerbates murine model of colitis	[47]
Rroid	Group 1 innate lymphoid cells	Nucleus	Mouse	Controls lineage identity, homeostasis, and function of group 1 ILCs	[53]
Spehd	Hematopoietic progenitors	Cytoplasmic	Both	Promotes multilineage differentiation and homeostasis	[25]
Adaptive Immunity
BCALM	B cells	Cytoplasm	Human	Negatively regulates B cell receptor-mediated calcium signaling	[78]
EPIC1	Malignant cells	Nucleus	Human	Promotes tumor immune evasion and resistance to checkpoint inhibitor therapy	[103]
FIRRE	Hematopoietic stem cells	Nucleus	Both	Promotes an increase in frequency of common lymphoid progenitors and peripheral blood CD4+ and CD8+ T cells	[63, 133]
Flatr	Regulatory T cells (Treg)	Unknown	Both	Promotes Treg differentiation	[73]
Flicr	Mature Tregs	Nucleus	Both	Reduces Treg activity, inhibits murine model of autoimmune diabetes	[83]
IFNG-AS1 (NeST or TMEVPG1)	Cytotoxic T cells (CTL) and Th1 cells	Nucleus	Both	Promotes Th1-lineage specific expression of IFNγ, regulates susceptibility to viral and bacterial pathogens, associated with UC risk locus, expression is increased in patients with UC and aGVHD	[80,81,97,98]
INCR1	Tumor cells	Nucleus	Human	Inhibits T cell-mediated tumor killing, promotes tumor cell growth and division	[101]
LINC00402 (ReLoT)	T cells	Both	Both	Augments T cell proliferation and TCR signaling, expression correlates with aGVHD development	[27]
Linc-MAF-4	Th1 cells	Nucleus	Human	Promotes Th1 phenotype, elevated in PBMCs of patients with aGVHD	[59, 96, 98]
LncCSRIgA	B cells	Nucleus	Both	Promotes class switch recombination to IgA and IgG2b, prevents intestinal dysbiosis and mucosal inflammation	[26]
LncDDIT4	Type 17 helper T cells	Cytoplasm	Human	Inhibits Th17 polarization, expression is elevated in PBMCs of patients with multiple sclerosis	[70]
Lnc-EGFR	Tregs	Cytoplasm	Human	Promotes Treg differentiation, inhibits cytotoxic T lymphocyte-mediated killing, increased expression correlates with increased hepatocellular carcinoma tumor size	[72]
LncHSC2	Hematopoietic stem cells	Nucleus	Mouse	Restrains lymphoid lineage commitment, involved in HSC self-renewal	[24]
Lnc-Smad3	Tregs	Nucleus	Both	Negatively regulates Treg differentiation	[74]
NKILA	CTLs and Th1 cells	Both	Human	Enhances T cell activation induced cell death, TIL apoptosis and tumor size are increased in patients with high expression	[82]
NRON	CTLs and Th1 cells	Cytoplasm	Both	Prevents the activation of T cells, promotes HIV latency	[75,86,87,154]
PVT1	Tregs	Nucleus	Mice	Enhances Treg function, prolongs allograft survival and alleviates graft rejection	[84]
Snhg1	CTLs	Nucleus	Both	Promotes CD8+ memory T cell differentiation, inhibits CD8+ effector differentiation in response to viral infection	[67]
TH2-LCR lncRNA cluster	Th2 cells	Nucleus	Both	Regulates the expression of Th2 cytokines	[62]
XIST	B cells	Nuclear	Human	Inactivates one X chromosome in early female development, maintains X chromosome inactivation of X-linked immune genes that may contribute to pathogenic atypical B cell development in SLE and rheumatoid arthritis	[21]
Both Innate and Adaptive Immunity
Malat1	Macrophages, Dendritic cells, Th1 cells, Th2 cells	Nucleus	Both	Myeloid: Negatively regulates innate antiviral responses, overexpression in DCs ameliorates cardiac allograft rejection, expression is lower in PBMCs of patients with SLE Lymphoid: Negatively regulates CD4+ cell function, promotes immune tolerance and tolerogenic dendritic cells that augment Treg expansion and anti-inflammatory cytokine production, delays clearance of Leishmania	[38,79,88]
MORRBID	Eosinophils, Neutrophils, Monocytes, and CTLs	Myeloid: Nucleus; Lymphoid: Cytoplasm	Both	Myeloid: Promotes myeloid cell survival by preventing apoptosis, expression is elevated in patients with hypereosinophilic syndromes Lymphoid: Promotes apoptotic death of infected cells, regulates CD8+ T cell survival and function	[22,23]

LncRNAs and Immunity

Immune regulation achieves a balance between host protection and damage from an overly aggressive immune response. LncRNAs are now known to be key for fine-tuning certain aspects of immune cell development, differentiation, and functions (Figure 2) as well as modifying protective and pathogenic immune responses (Table 1)^22-28.

Figure 2. Modulatory LncRNAs in immune cells.

Selected lncRNAs that regulate immune cell development, differentiation, activation, and function are shown. LncRNAs that have been reported to promote (blue typeset), inhibit (red typeset), or both inhibit and promote (pink) the developmental step or function of their indicated cell type are denoted. LncRNAs with an “*” have orthologs in mice and humans. LncRNAs in all capital letters without an asterisk are present in humans but do not have a known mouse ortholog. For further functional details about each lncRNA, please refer to the text. CLP: common lymphoid progenitor, CMP: common myeloid progenitor, ILC: innate lymphoid cell, NK: natural killer cell, Th: CD4⁺ helper T cell. This figure was created using BioRender (https://biorender.com/).

LncRNAs Modulate Innate Immunity

Innate immunity is a rapid inflammatory response that shapes the subsequent adaptive immune response. It is mainly mediated by hematopoietic-derived myeloid and lymphoid innate immune cells. Recent data point to a key cell-specific role for lncRNAs in innate myeloid and lymphoid cell development, differentiation, and function. Consistent with their often species- and cell-specific expression, some lncRNAs appear to be functional in only one species while conserved lncRNAs appear to share function in cells across species^12,16,17.

LncRNAs are now recognized as critical regulators of hematopoietic innate myeloid cell (e.g., monocytes, macrophages, neutrophils, and dendritic cells) development and survival. For example, Spehd is expressed in mouse and human hematopoietic stem and progenitor cells and is necessary for murine multilineage differentiation and maintaining mitochondrial function in myeloid progenitors²⁵. The myeloid-specific lncRNA LOUP promotes expression of the master myeloid transcription factor PU.1 during human myeloid differentiation by recruiting the transcription factor RUNX1 to the PU.1 promoter and enhancer²⁹. By contrast, transcription of an antisense lncRNA from an alternative promoter in intron 3 of PU.1 induced during lymphoid differentiation blocked myeloid differentiation in mice and humans³⁰. Similarly, cell type-specific regulation by another conserved lncRNA, MORRBID, promotes innate myeloid cell survival²². It achieves this by recruiting inhibitory chromatin marks to the pro-apoptotic Bcl2l11 locus in cis²². Thus, lncRNAs can provide a lineage-specific layer of regulation on myeloid cell development and survival, and their molecular mechanisms appear to be diverse ranging from regulating transcription to regulating metabolism.

Innate immune myeloid cell differentiation, specifically macrophage differentiation, is now understood to be governed by lncRNAs. For instance, pro-inflammatory macrophage differentiation is inhibited by Mirt2, which in mice is induced by LPS stimulation and downregulated by the anti-inflammatory polarizing cytokine IL-4 in vitro³¹. Mirt2 inhibits inflammatory macrophage polarization by binding TRAF6 and blocking TRAF6’s ability to promote a pro-inflammatory phenotype³¹. Similar to Mirt2, PTPRE-AS1 also inhibits inflammatory macrophage differentiation³². It is specifically induced by IL-4 in murine and human macrophages where it promotes expression of PTPRE by increasing histone lysine trimethylation (H3K4me3) of the PTPRE promoter through its interaction with WDR5, a component of histone H3 Lys4 methyltransferase complexes³². PTPRE then favors inflammatory differentiation by blocking the MAPK/ERK pathway³². Consistent with its favoring inflammatory differentiation, PTPRE-AS1 expression is lower in peripheral blood mononuclear cells (PBMCs) of patients with asthma versus healthy controls, and Ptpre-as1^−/− mice are more susceptible to allergic pulmonary inflammation than wild-type animals³². However, future studies will need to clarify the in vivo dependence of Ptpre-as1 deficiency in macrophages using conditional models and further exploring its expression and function in other cell types. These data so far show that lncRNA-mediated effects on cell signaling and transcription are key mechanisms for their regulation of myeloid cell differentiation.

In addition to myeloid cell development and survival, lncRNAs can also regulate myeloid cell function³³. Innate myeloid cell functional responses are often triggered by danger-associated molecular patters (DAMPs) or pathogen-associated molecular patterns (PAMPS) that initiate signaling cascades mediated by viral (e.g. RIG-I and MDA5), bacterial (e.g. TLR4 and the NLRP3 inflammasome), and fungal (e.g TLR2 and NOD2) pattern recognition receptors (PRRs)³⁴. These signaling cascades induce cell-intrinsic immune responses and release antimicrobial peptides, cytokines, and chemokines that limit the spread of infection and also activate the adaptive immune response. LncRNAs are integral regulators of innate viral, bacterial, and fungal PRR pathways in myeloid innate immune cells.

One viral PRR pathway is triggered by cytosolic viral RNA binding the broadly expressed PRR RIG-I. In turn, RIG-I signals through TANK binding kinase 1 (TBK1) leading to interferon regulatory factor 3 (IRF3) activation and release of anti-viral type-I interferons (type-I IFN)²⁸. Several lncRNAs have been shown to regulate this pathway in macrophages. For instance, murine Lnc-Lsm3b³⁵ negatively regulates this pathway by binding RIG-I and stabilizing it in an inactive conformation³⁵. By contrast, Lnczc3h7a in mouse macrophages promotes RIG-I signaling by serving as a molecular scaffold for the RIG-I activator TRIM25³⁶. LncRNA-GM in murine and human macrophages promotes antiviral innate immune responses by binding to glutathione S-transferase M1 (GSTM1) and impairing its ability to inactivate TBK-1²⁸. Whilst, LncRNA-ISIR is a conserved positive regulator of IRF3 in mouse and human macrophages³⁷, in murine macrophages, Malat1 is an indirect negative regulator of IRF3 by interacting with the RNA binding protein TDP43³⁸. These studies and others reviewed elsewhere³⁹, demonstrate the extensive regulation of viral PRR pathways by lncRNAs. In addition, emerging evidence indicates viruses employ lncRNA-based strategies of circumventing antiviral PRR signaling further illustrating the intricate regulation of these pathways by both host and pathogen lncRNAs³⁹.

Regulation of bacterial and fungal PRR pathways is also mediated by lncRNAs⁴⁰. For instance, LincRNA-EPS acts as a negative regulator of murine macrophage and dendritic cell inflammatory cytokine production by binding hnRNPL⁴¹. It is downregulated following stimulation with a broad array of in vitro TLR ligands, Listeria monocytogenes infection, and Sendai virus infection thereby promoting inflammatory/innate immune response gene expression⁴¹. Similarly, Lnc13 is a negative regulator of inflammatory gene production, and its expression in human and murine macrophages decreases following LPS stimulation in vitro as a result of reduced stability from greater decapping by DCP2⁴². These studies collectively indicate that lncRNAs can be key negative and positive regulators of anti-viral, anti-bacterial, and anti-fungal PRR pathways.

Inflammasomes are cytoplasmic PRR complexes that induce innate myeloid cell inflammatory and anti-pathogen responses⁴³, and they too can be regulated by lncRNAs. For instance, Neat1 is well-known for its role in nuclear paraspeckles⁴⁴; however, it also serves as an enhancer of in vitro inflammasome assembly when comparing wild-type to Neat1^−/− murine macrophages⁴³. Neat1 also promotes inflammatory dendritic cell responses (DC) by regulating the Nlrp3 inflammasome as evidenced by decreased in vitro murine DC inflammatory cytokine production following Neat1 siRNA-mediated depletion⁴⁵. Several other lncRNAs may regulate inflammasomes and are reviewed elsewhere⁴⁶. Like Neat1, the majority of these are widely expressed, thus a major challenge for translating these findings into improved outcomes for inflammasome-mediated disorders will be identifying tissue- and context- specific lncRNAs regulating inflammasomes.

Myeloid cell functions beyond PRR pathway signaling can also be regulated by lncRNAs. For example, the conserved lncRNA NAIL positively regulates inflammatory cytokine production (TNFα and IL-1β), primarily in bone marrow derived macrophages, in a mouse model of colitis⁴⁷. NAIL promotes both p38 MAPK and p65 NF-κB activation by sequestering and inhibiting the phosphatase Wip1⁴⁷. Like NAIL, Lnc-UC is conserved between mice and humans and is induced by NF-κB⁴⁸. It decreased inflammatory signaling in a diurnal manner by inducing the circadian clock gene Rev-erbα, which in turn, inhibited NF-κB and the Nlrp3 inflammasome in macrophages⁴⁸. However, both of these lncRNAs are broadly expressed limiting their specificity as biomarkers or therapeutic targets. In addition, these functions and mechanisms may be variable in different cells.

Metabolic pathways influence myeloid cell function, and lncRNAs regulate these pathways. As an example, expression of LncRNA-ACOD1 is induced by viral infection in mouse macrophages where it binds and enhances the enzyme glutamic-oxaloacetic transaminase (GOT2)⁴⁹. The GOT2 metabolites, aspartate and α-ketoglutarate, enhance viral replication⁴⁹. By contrast, Lnc-Dpf3 inhibits HIF1α-dependent glycolysis in mouse dendritic cells and restrains inflammation⁵⁰. While these studies showcase the importance of lncRNAs in macrophage and DC metabolism, further studies are needed to understand how lncRNAs influence metabolism and function in macrophage and DC subsets. Such granular understanding may reveal highly specific translational targets.

Similar to myeloid innate immune cells, lncRNAs are also necessary for innate lymphoid cell (ILC) development, differentiation, and function. LncKdm2b is required for type 3 innate lymphoid cell (ILC3) maintenance and proliferation as demonstrated by deficiency of LncKdm2b in the hematopoietic compartment (LncKdm2b^f/fl; Vav-Cre⁺) or the ILC3 compartment (LncKdm2b^f/fl; Rorc-Cre⁺)^51,52. Rroid specifically controlled the identity, homeostasis, and function of murine group 1 ILCs by regulating the expression of the key ILC1 transcription factory Id2⁵³. Global analysis of lncRNA expression in human NK cells identified many novel NK cell-specific lncRNAs that may regulate their differentiation and function⁵⁴. For example, Lnc-CD56 positively regulated CD56 expression of primary NK cells⁵⁴. These studies suggest lncRNAs are pervasive regulators of innate lymphoid cell biology; however, further studies need to focus on lncRNAs required for each developmental stage and clinically relevant functions of each subset. This level of understanding may lead to improved innate lymphoid cell-based therapies, which have recently shown promising anti-leukemia activity⁵⁵.

Role of LncRNAs in Adaptive Immunity

Adaptive lymphoid cell development can also be shaped by lncRNAs. For example, lncRNA expression patterns are highly stage specific in mouse and human B and T cells and are more specific than protein-coding genes^56-62. These lncRNAs are often co-expressed with neighboring protein-coding genes enriched for ontologies pertaining to B or T cell development, differentiation, and activation, respectively^56-62. In addition, adoptive transfer of murine HSCs depleted of LncHSC2 or Lnc6689 restrained lymphoid commitment or B cell development, respectively^24,25. Deletion of the conserved, broadly expressed, trans-acting lncRNA Firre locus in mice reduced common lymphoid progenitors and peripheral blood T cells whereas inducible overexpression of Firre increased peripheral NK cells in vivo⁶³. Consistent with their developmental-specific expression, lncRNA expression in murine B and T cell subsets was controlled by subset-defining transcription factors^57,60. With regard to B cell maturation, lncCSR^IgA promoted class switch recombination to IgA and IgG2b by regulating local and long-range chromosomal topology and interactions in cis as demonstrated in lncCSR^IgA−/− mice²⁶. While these studies imply lncRNAs are important for adaptive lymphoid cell development, the precise mechanism(s) and developmental stage(s) where they act remain largely unknown. Establishing conditional and inducible deletion and overexpression murine models will help refine these mechanisms. In addition, many HSC-specific lncRNAs remain uncharacterized suggesting that much remains unknown regarding lncRNA regulation of lymphopoeisis^24,25,64. In this regard, CRISPR-Cas9-based high-throughput approaches may be well suited to detect lncRNAs with non-redundant functions. However, determining those that are redundant may remain difficult. Further, recent reports suggest lncRNAs are important for leukemia stem cell function^65,66, thus a thorough understanding of lncRNAs biology in normal and dysregulated hematopoiesis may reveal new insights and potential translational strategies for improving leukemia therapy.

Following antigen activation, naïve T cells differentiate into CD8⁺ or CD4⁺ memory and effector subsets⁶¹. LncRNA Snhg1 promoted both human and mouse CD8⁺ memory T cell differentiation while inhibiting CD8⁺ effector differentiation in response to viral infection⁶⁷. These studies were carried out using an in vivo murine adoptive transfer model of lymphocytic choriomeningitis virus- (LCMV) specific T cells transduced with shRNAs to deplete Snhg1 or constructs to overexpress Snhg1⁶⁷. The authors further showed that lncRNA Snhg1 interacted with the vesicle trafficking protein Vps13D to promote IL-7Rα surface expression and signaling thereby enhancing CD8⁺ memory T cell differentiation⁶⁷. Given the importance of memory T cells to anti-tumor and vaccine responses^68,69, further exploring lncRNA regulation of memory T cell development may lead to translational strategies for improving memory T cell responses.

CD4⁺ T cell helper subsets include pro-inflammatory (Th1 and Th17), pro-autoimmune (Th2), and regulatory (Treg) cells (among others). Linc-MAF-4 promotes Th1 in vitro polarization⁵⁹ of primary human T cells by recruiting the repressive chromatin modifiers EZH2 and LSD1 to the adjacent locus encoding the Th2-promoting transcription factor Maf⁵⁹. By contrast, LncDDIT4 is induced under in vitro Th17-polarizing conditions and inhibits Th17 polarization of human CD4⁺ T cells, suggesting it may act as a Th17-specific negative feedback regulator⁷⁰. Human in vitro Th2 differentiation is also supported by lncRNAs expressed from the Th2 locus control region (LCR), which promotes expression of Th2 cytokines IL-4, IL-5, and IL-13 in a manner that may depend on WDR5 promoting the expression of the Th2 master transcription factor GATA3^62,71.

Human Treg differentiation is supported by Lnc-EGFR binding to the EGFR receptor and increasing downstream signaling⁷². Enhanced Treg differentiation by lentivirus overexpression of Lnc-EGFR inhibited cytotoxic T lymphocyte-mediated killing⁷². Similar to Lnc-EGFR, the lncRNA Flatr also promoted mouse Treg differentiation in vitro and in vivo using a murine knockout model; however, its molecular mechanism has not yet been determined⁷³. In contrast to Lnc-EGFR and Flatr, Lnc-Smad3 is a negative regulator of in vitro murine Treg differention⁷⁴. Lnc-Smad3 silences the Treg-promoting transcription factor Smad3 by recruiting histone deacetylase 1 (HDAC1) and silencing Smad3 transcription⁷⁴. These studies suggest that lncRNAs control helper T cell differentiation at various levels including epigenetic, transcriptional, and post-transcriptional.

Apart from contributing to the regulation of lymphoid cell development and maturation, lncRNAs are integral regulators of lymphoid cell function^75-78. Malat1 regulates CD4⁺ helper T cell function and is decreased upon helper T cell activation⁷⁹. Despite being decreased upon activation, it promotes expression of the transcription factor Maf and the anti-inflammatory Maf target IL-10⁷⁹. LINC00402 (ReLoT) is a T cell-enriched lncRNA that augments T cell receptor (TCR) signal transduction²⁷ when overexpressed in human T cell lines. IFNG-AS1 (NeST or TMEVPG1) is a nuclear-localized lncRNA that promotes IFNγ expression in mouse and human Th1 cells by recruiting the permissive chromatin modifier WDR5 to the IFNG locus^80,81. The lncRNA NKILA enhances activation-induced cell death (AICD) in human Th1 and CD8⁺ cytotoxic breast and lung tumor infiltrating lymphocytes (TILs) by preventing nuclear translocation of the pro-survival T cell activation transcription factor NF-κB⁸². Like NKILA, Morrbid promotes apoptotic death of antigen-specific murine CD8⁺ T cells during the contraction phase of an in vivo T cell response to LCMV by increasing expression of the anti-apoptotic protein Bim²³. Morrbid also inhibits PI3K-AKT signaling downstream of the type-I IFN receptor required for optimal effector function (i.e., IFNγ production)²³. Taken together, these studies demonstrate a wide array of potentially translational functions for lncRNAs in T cells. However, whether these functions, demonstrated in model systems, will translate to improvements in diagnostics or therapeutics is an open question.

Treg function can also be modulated by lncRNAs. Flicr is a negative regulator of the key Treg transcription factor Foxp3⁸³. In a murine knock-out model, it decreased Foxp3 expression by modifying chromatin accessibility of the Foxp3 conserved noncoding sequence 3⁸³. As evidence of its biologic relevance, Flicr deletion promoted autoimmune diabetes in a murine model⁸³. In contrast to Flicr, PVT1 enhances murine in vitro Treg function by augmenting autophagy⁸⁴. Treg adoptive transfer has shown promise for treating or preventing many inflammatory and autoimmune disorders⁸⁵. Therefore, understanding the role of lncRNAs in Treg development and function may reveal targeted strategies for improving Treg-based therapeutics.

LncRNAs Modulating Immunity In Vivo During Disease

Consistent with their pervasive effects on the immune system, lncRNAs shape both protective and pathologic immune responses. Below we highlight those lncRNAs that influence outcomes of immune responses to infection, inflammation, autoimmunity, and malignancy (Table 1). How lncRNAs might be used to improve the treatment and diagnosis of immune-mediated disorders is discussed in Box 4.

Box 4. LncRNAs as Specific Translational Targets for Modulating Immunity.

LncRNAs might be useful candidate biomarkers for certain immune-mediated pathologies. The superb cell type-specific expression and surprising stability of lncRNAs in body fluids makes them well suited for this purpose. Indeed, lncRNAs have been extensively explored as biomarkers for the diagnosis of kidney allograft rejection^139-141 and malignancy^142,143. Promising lncRNA biomarkers are now emerging for the diagnosis and risk-stratification of infection, autoimmunity, and inflammatory disorders including sepsis^144,145, SLE^146-148, and GVHD^97,98,149, but evidently remain to be validated. LncRNAs might also be able to predict certain anti-tumor responses to immuno-therapy, although this remains conjectural at present^77,103,104.

The recent technical advances in RNA-based medicines and vaccines along with the FDA approval of antisense oligonucleotide-based (ASO) therapeutics^18,150,151 suggest that targeting immune-related lncRNAs might be feasible, particularly for tissue- and context-specific lncRNAs. In addition to ASO-based approaches, lncRNA mimetics with or without tissue-targeting linker molecules are also exciting possibilities that remain to be assessed¹³⁴. However, the development of lncRNA therapeutics, will need to be thoughtfully pursued, particularly for broadly expressed lncRNAs with cell type-specific functions¹³³ that may have opposing phenotypes. For example, Neat1 is pro-inflammatory in macrophages and DCs^45,50. However, in a Neat1^−/− myocardial infarction model, myocardial inflammation was increased, and a multitude of opposing effects on the function of various myeloid and lymphoid immune cell subsets were seen¹⁵². Similarly, NKILA is both oncogenic in human breast cancer cells and inhibitory toward tumor infiltrating lymphocytes in a breast cancer patient-derived xenograft model^82,153. Thus, systemic depletion of NKILA may have competing outcomes on tumor growth. Although, adoptive transfer of NKILA-depleted tumor-specific CTLs might be a promising approach for enhancing tumor immunotherapy, which is presently speculative. Before any lncRNA is targeted therapeutically, a detailed understanding of host expression and function is imperative.

LncRNAs Regulate Infectious Disease Outcomes

LncRNA regulation of innate PRRs pathways, myeloid cell function, and lymphoid cell function is crucial for antimicrobial immune responses. Consistent with their ability to either inhibit or augment antiviral PRR pathways, LncLsm3b^−/− and Malat1^−/− mice are resistant to vesicular stomatitis virus (VSV) infection whereas LncRNA-GM^−/− and LncRNA-ISIR^−/− mice are more susceptible to VSV infection in vivo^28,35,37,38. The lncRNA NRON promotes HIV-1 latency by restricting the host HIV-1 transactivator, NFAT, to the cytoplasm in human T cell lines, whereas in primary human CD4⁺ T cells it depletes the HIV-1 transactivator Tat^86,87. Firre overexpressing and LincRNA-EPS^−/− mice are more sensitive to LPS endotoxemia than wild-type control mice, whereas mice administered a Mirt2 overexpressing adenovirus vector are more resistant to LPS endotoxemia than mice given an empty vector^31,41,63. Consistent with Neat1’s pro-inflammatory role, inflammation is reduced in Neat1^−/− mouse models of peritonitis and pneumonia relative to wild-type controls⁴³. ILC3s are important for responses to intestinal bacterial infections⁵², and conditional deletion (LncKdm2b^f/fl; Rorc-Cre⁺) of the ILC3 lncRNA, LncKdm2b, causes increased mortality following intestinal infection with Citrobacter rodentium compared with wild-type mice⁵¹. Correlating with Malat1’s tolerogenic phenotype in T cells and DCs^79,88, Malat1^−/− mice clear Leishmania donovani parasites faster than wild-type animals⁷⁹. These studies collectively demonstrate the importance of lncRNAs in outcomes of infection. However, the majority of these studies employed knock-out mouse models to demonstrate function. To translate these discoveries into potential therapies and capitalize on the tissue- and context-specific expression of lncRNAs, future studies will need to develop and test therapeutics directly targeting these lncRNAs or their associated upstream or downstream functional mediators.

LncRNAs Influence Autoimmune and Auto-inflammatory Disorders

Autoimmune and auto-inflammatory disorders may be modulated by lncRNAs. Specifically, the role of lncRNAs in systemic lupus erythematosus (SLE) is increasingly appreciated. SLE is associated with high type-I IFN expression, and in agreement with MALAT1’s negative regulation of type-I IFN induction, PBMCs from patients with SLE express less MALAT1, more IFNG, and have greater amounts of activated IRF3, all of which were reversed following SLE-directed therapy³⁸. Conversely, Lnc-ISIR promotes IRF3 signaling, and its expression is elevated in PBMCs of patients with SLE and correlates with disease severity relative to healthy controls or patients with SLE receiving effective treatment with glucocorticoids and antimalarial drugs³⁷.

In addition to being associated high type-I IFN expression, SLE is associated with atypical memory B cells, and it has a female predominance, suggesting that X-linked genes may contribute to its development⁸⁹. In patients with SLE, maintenance of X-chromosome inactivation (XCI) is aberrant in T and B cells and is associated with a reduction in the repressive chromatin mark H2AK119Ub on the inactivated X chromosome (Xi) in B cells and increased X-linked gene expression in B and T cells^89,90. XCI requires the lncRNA XIST, and genes dependent on XIST for their silencing are overexpressed in PBMCs and atypical memory B cells in female patients with SLE²¹. Furthermore, signaling from TLR7, an X-linked gene, induces atypical memory B cells²¹, and XIST deletion promotes atypical memory B cell development and TLR7 expression. Therefore, aberrant loss of XIST expression has been suggested to underlie atypical memory B cell development that contributes to a degree of predilection for SLE in women²¹. Although, whether this underlies the predilection for other autoimmune diseases in women remains unknown.

The proinflammatory lncRNA NAIL is upregulated in patients with ulcerative colitis (UC), and transgenic mice lacking Nail expression due to a deletion of NF-κB binding sites in the Nail promoter are resistant to dextran sulfate sodium- (DSS) induced colitis relative to wild-type controls⁴⁷. By contrast, deletion of the diurnal anti-inflammatory lncRNA Lnc-UC exacerbates DSS colitis in mice⁴⁸. Similarly, Ptpre-as1^−/− mice are more susceptible to colitis than wild-type control mice³². Another example is that of IFNG-AS1, which promotes expression of the inflammatory cytokine IFNγ^80,91,92. The UC susceptibility locus rs7134599 is associated with IFNG-AS1, and its expression is increased in the colons of patients with UC and in trinitro-benzenesulfonic acid- (TNBS) induced murine colitis compared to healthy human or mock treated mice, respectively⁹³. Interestingly, IgA is important for intestinal mucosal homeostasis, and lncCSR^IgA−/− mice develop intestinal microbial dysbiosis and greater mucosal inflammation compared with wild-type controls²⁶. Importantly, polymorphisms in the lncCSR^IgA locus are enriched in patients with IgA deficiency, suggesting that in certain individuals, mutations at this locus may contribute to IgA deficiency. Although, this will need to be formally tested²⁶. While these studies highlight the diverse ways lncRNAs contribute to intestinal immune homeostasis, future studies will need to carefully examine the interaction between lncRNAs expressed in the intestines and the microbiome^94,95.

Linc-MAF-4 expression was found to be higher in peripheral blood mononuclear cells (PBMCs) from patients with multiple sclerosis (MS) than in healthy controls, and it correlated with MS relapse rates, suggesting that linc-MAF-4 may contribute to the pathogenies of multiple sclerosis⁹⁶. The role of lncRNAs in autoimmune and auto-inflammatory diseases is just beginning to be understood. It will be exciting to learn how partially characterized lncRNAs as well as the thousands of lncRNAs that are yet to be studied might regulate these disorders.

LncRNAs in Alloimmunity

Alloimmune reactions cause acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation (HSCT) and organ rejection after solid organ transplantation. LncRNAs have recently been implicated in the pathology of alloimmunity. LINC00402, when overexpressed in human T cells, enhanced TCR signal transduction, while its depletion in murine and human T cells decreased proliferation in response to an allogeneic stimulus²⁰. Further, LINC00402 expression was decreased in peripheral blood T cells of allogeneic HSC and solid organ transplant recipients suggesting it may be a useful marker of T cell alloimmunity²⁷. Several lncRNAs that support Th1 differentiation and function including Lnc-DC, linc-MAF-4, and IFNG-AS1 are elevated in PBMCs of patients with acute GVHD relative to non-aGVHD controls^97,98. In solid organ murine allograft rejection models, adoptive transfer of Neat1-depleted DCs when compared with control DCs ameliorated myocarditis and cardiac allograft rejection⁴⁵. Conversely, adoptive transfer of Malat1 overexpressing DCs or PVT overexpressing Tregs decreased murine cardiac allograft rejection^84,88. The above studies indicate lncRNAs may be cell-type specific regulators of GVHD and solid organ allo-graft rejection. Both GVHD and graft rejection are difficult to predict and frequently require invasive procedures to diagnose^99,100. Therefore, diagnostic or predictive lncRNA biomarkers may be particularly valuable in these disorders.

LncRNAs in Anti-Tumor Responses

LncRNAs are emerging as important contributors to T cell dysfunction within tumors. NKILA promoted AICD of tumor infiltrating lymphocytes (TIL), while adoptive transfer of NKILA-depleted cytotoxic lymphocytes inhibited breast cancer patient-derived xenograft tumor growth in immunodeficient (NOD.SCID) mice compared to control cytotoxic lymphocytes⁸². TIL apoptosis was increased and overall survival was decreased in breast cancer patients with > 30% of TILs expressing high levels of NKILA (defined as more than 4 gene signals per cell by fluorescence in situ hybridization) compared to those with a lower percentage of high NKILA-expressing TILs⁸². These data suggest that NKILA expression in TILs may regulate anti-tumor T cell responses in certain tumors. Like NKILA, Lnc-EGFR may also shape anti-tumor T cell responses⁷². It is upregulated in Tregs infiltrating human hepatocellular carcinomas (HCC) and enhances Treg differentiation when overexpressed in vitro or in adoptively transferred TILs in an orthotopic HCC model in NOD.SCID mice compared to empty vector control transduced CD4⁺ human T cells⁷². Overexpression of Lnc-EGFR inhibited anti-tumor T cell activity in cytotoxic T lymphocyte killing assays. Further, greater Lnc-EGFR expression in peripheral blood CD4⁺ T cells (cut off using the upper 95^th percentile confidence interval of peripheral blood CD4⁺ T cells in patients with HCC) correlated with increased tumor size (greater than 5 cm)⁷². These data suggest that higher expression of Lnc-EGFR may be an important determinant of Treg differentiation and suppression of anti-tumor T cell responses in patients with HCC. However, this will require independent confirmation, and additional studies will be needed to accurately identify those patients with higher expression.

LncRNAs also contribute to tumor cell evasion of T cell attack by downregulating antigen presenting machinery or by upregulating immune checkpoints. INCR1 is a lncRNA induced by IFNγ from the CD274 locus (encodes the immune checkpoint ligand PD-L1) in human glioblastoma cell lines¹⁰¹. It inhibits in vitro and in vivo T cell-mediated killing of human glioblastoma cells by sequestering the ribonucleoprotein HNRNPH1 thereby blocking it from inhibiting the expression of CD274 and JAK2¹⁰¹. By contrast, EPIC1 was first described as an oncogenic lncRNA in breast cancer that interacts with the oncogene MYC and promotes cell-cycle progression¹⁰². EPIC1 also suppresses breast cancer tumor antigen presentation thereby promoting tumor immune evasion and resistance to checkpoint inhibitor therapy in a murine model¹⁰³. Because EPIC1 can promote both tumor immune evasion when overexpressed in murine breast and colon cancer models and can also promote oncogenesis^102,103, anti-sense oligos to EPIC1 may be potent inhibitors of tumor growth by blocking both the oncogenic and immune evasion activity of EPIC1. By contrast, the lncRNA LIMIT is induced by IFNγ and promotes antigen presentation in human and mouse tumor cells and professional APCs¹⁰⁴. By augmenting antigen presentation, LIMIT enhances anti-tumor and checkpoint inhibitor responses in murine models of melanoma and colorectal carcinoma¹⁰⁴. These studies suggest that targeting lncRNA-mediated tumor immune evasion directly, or indirectly through their associated molecular mechanism(s), may improve cancer outcomes. Future studies will need to test potential lncRNA targets and validate them for off- and on-target side effects.

While some lncRNA genes might contribute to tumor immune evasion, others harbor unrecognized micro-peptides that are a source of targetable tumor-specific antigens called neoantigens. These neoantigens are presented by MHC molecules, recognized by tumor-specific T cells, and are often encoded by specific lncRNA isoforms^105-108. For example, the lncRNA PVT1 cooperates with the oncogene MYC to promote tumorigenesis¹⁰⁹. However, select PVT1 isoforms, expressed prominently in colorectal cancer, encode a micro-peptide that is targeted by patient-derived tumor-infiltrating lymphocytes¹⁰⁷. Hence, lncRNAs might be a major source of targetable tumor-specific antigens for anti-tumor immunotherapy.

Concluding Remarks

LncRNAs are a vast group of genes with diverse molecular mechanisms of action ranging from epigenetic, post-transcriptional, and regulation of signaling molecules. The majority of lncRNA genes are not functionally characterized. Despite this, a growing body of evidence demonstrates that lncRNAs are crucial regulators of innate and adaptive immunity. In addition, emerging studies, despite the limitations inherent to murine models and in vitro culture systems, demonstrate that lncRNAs impact protective and pathologic in vivo immune responses and may offer opportunities for developing more specific diagnostics and therapeutics. In order to realize these opportunities, several important questions need to be addressed (see Outstanding Questions). In addition, more study is needed to fully characterize the function of known immune-related lncRNAs in different tissues and disease contexts. Such characterization will help avoid unintended side effects of any lncRNA selected for therapeutic targeting. Further, the function of the numerous uncharacterized myeloid and lymphoid cell subset-specific lncRNAs need to be determined to better understand the biology of each subset. To facilitate the characterization of these lncRNAs, improved prediction methods of lncRNA function based on sequence and other molecular or biological factors is urgently needed. Moving forward, we expect that the striking tissue-specificity of lncRNAs and emerging technologies to safely target them will lead to improved outcomes for immune-mediated diseases.

Outstanding Questions.

• Can lncRNAs be safely and effectively targeted in vivo? The tissue-specific expression of lncRNAs suggests they may be ideal therapeutic targets using approaches such as ASOs. However, any approach will need to be rigorously tested and targets chosen only after extensive exploration of off- and on-target side effects.

• What is the best way to utilize the tissue- and context-specific expression of lncRNAs as clinical biomarkers? Will they be most useful as lncRNA-only panels or in combination with other existing markers? Future studies will need to extensively validate the specificity and sensitivity of any lncRNA biomarker in multiple cohorts. LncRNA biomarkers will also need to be tested against healthy controls and those with potentially confounding conditions co-experienced in certain patient populations (e.g., infection versus inflammation).

• What is the best way to predict potentially functional immune-regulatory lncRNAs? There are thousands of uncharacterized lncRNA genes. While expression/differential expression relevant to the immune function being studied is one established approach, this typically does not sufficiently narrow most lncRNA gene lists. Other factors commonly used for protein-coding genes such as nucleotide conservation are poor predictors of function. Emerging techniques include k-mer analysis and maintenance of synteny, but these need further detailed validation particularly within the immune system.

Figure 1. Examples of lncRNA molecular mechanisms.

A) LncRNAs in the nucleus often recruit chromatin modifying complexes to gene promoters thereby regulating cis and trans gene expression^{22,59,63,80,81}. B) Some lncRNAs sequester chromatin modifying complexes away from their cis or trans target gene promoters². C) LncRNAs may also form R-loops which recruit chromatin modifying complexes that then regulate cis target gene promoters⁷¹. D) Promoter antisense (PAS) lncRNAs of ligand-induced paused protein-coding genes recruit demethylases that release repressive transcriptional complexes from their paused cis protein-coding genes¹²⁰. E) Enhancer elements within or near lncRNAs recruit transcription factors to promoters of cis and trans target gene promoters². The lncRNA in this situation is dispensable for gene regulation. F) Cytosolic lncRNAs can regulate the translation of mRNAs². G) LncRNAs regulate signal transduction at multiple levels including receptor activation, activity of signaling molecules, and activation and localization of transcription factors^72,75,154. H) Enzyme activity can be regulated by direct lncRNA binding^49,134. I) LncRNA can serve as “sponges” for miRNAs thereby increasing the expression of mRNAs targeted by the “sponged” miRNAs⁴⁵. J) LncRNAs can also harbor occult small open reading frames (sORF) that produce micro-peptides with potentially distinct functions from their parent lncRNA molecules¹²⁹. This figure was created using BioRender (https://biorender.com/).

Highlights.

• Long noncoding RNAs are tissue- and context-specific modulators of immune cell development, differentiation, activation, and function.

• Long noncoding RNA molecular mechanisms are diverse and challenging to predict.

• Long noncoding RNAs impact outcomes of protective and pathogenic immune responses in a variety of diseases.

• Long noncoding RNA genes outnumber protein-coding genes, yet comparatively few long noncoding RNAs have been functionally or mechanistically characterized.

• Further unlocking the role of long noncoding RNAs in immunity may lead to improved outcomes for diseases influenced by protective and pathogenic immune responses.

Glossary

Activation-induced cell death

Programmed cell death of multiply activated T cells mediated by interactions between death receptors and their ligands

Acute graft-versus-host disease

An inflammatory immune reaction that primarily damages the skin, gut, and liver and is driven by donor T cells (graft) responding to host allogeneic antigens

Adaptive immune cells

Include CD4⁺ T cells, CD8⁺ T cells, and B cells.

Alloimmunity

An immune reaction to nonself antigens of the same species

Antisense oligonucleotide-based therapeutics

Oligonucleotide-based therapeutics that work typically by causing target RNA degradation or alternative spicing

Atypical memory B cells

Incompletely defined CD27⁻ CD11c⁺ B cell population that is expanded with certain infectious diseases (malaria, HIV, COVID-19) and certain autoimmune diseases (systemic lupus erythematosus and rheumatoid arthritis)

Autophagy

A regulated conserved lysosomal-dependent degradation process for cellular components

Checkpoint inhibitor

A type of drug that blocks negative regulatory pathways in immune cells, particularly T cells, thereby promoting their activity and in some cases anti-tumor responses

Cis interactions

Occur and produce functional consequences proximal to and on the same chromosome as the lncRNA

Class switch recombination

A biologic process that results in the switching of immunoglobulin type produced by B lymphoid cells

Competing endogenous RNAs

Refers to lncRNAs that regulate the expression of other transcripts by competing for shared micro-RNAs

Danger associated molecular patterns

Molecules released by damaged or dying cells that active innate inflammatory pathways

Group 1 ILCs

Innate lymphoid cells consisting of ILC1 and NK cells

Innate lymphoid cell (ILC)

Innate immune cell counterpart to T cells that contribute to immune responses by secreting cytokines

ILC1

Type 1 innate immune cells are most analogous to CD4⁺ helper type 1 (Th1) cells in terms of their cytokine secretion profile and characteristic transcription factors

ILC3

Type 3 innate immune cells are most analogous to CD4⁺ helper type 17 (Th17) cells in terms of their cytokine secretion profile and characteristic transcription factors. They are important for intestinal mucosal immunity

Immune checkpoints

Negative regulatory pathways in immune cells important for self-tolerance

Inflammasomes

Multi-protein cellular complexes that recognize DAMPS and PAMPS and initiate inflammatory cytokine production

k-mers

In this article it refers to oligonucleotide motifs of a defined (k) length. In general, refers to any oligomer of k length.

Liquid-liquid phase separation

Membraneless macromolecular complexes in cells that condense into a dense phase often resembling liquid droplets

Long noncoding RNA

Non-protein-coding transcript at least 200 nucleotides long

Microbial dysbiosis

A disruption in microbial homeostasis that can include altered numbers and or proportions of typical microbes in a community

Multiple sclerosis

An inflammatory demyelinating neurologic disorder thought to be mediated in part by CD4⁺ type-1 helper T cells

Nuclear paraspeckles

Membraneless, stress-induced protein and RNA nuclear bodies that regulate gene expression by sequestering proteins and RNA

Pathogen-associated molecular patterns

Molecules with conserved motifs associated with infection that activate the innate immune response and are recognized by pattern recognition receptors

Pattern recognition receptors

Receptors that recognize molecular patterns frequently associated with microbial pathogens

Phenylketonuria

Inherited deficiency in the ability to metabolize phenylalanine that can cause neurologic damage if untreated

Professional innate immune cells

Hematopoietic-derived myeloid (neutrophils, eosinophils, basophils, monocytes, macrophages, and dendritic cells) and lymphoid (natural killer cells and ILCs) cells, that respond rapidly to pathogens

Pumilio RNA-binding proteins

Family of RNA binding proteins present in eukaryotes that function as post-transcriptional repressors

R-loop

3-stranded co-transcriptional complex composed of a DNA: RNA hybrid and the associated non-template single-stranded DNA

Systemic lupus erythematosus

A multiorgan autoimmune disease associated with high type-I IFN expression.

Trans interactions

Occur and produce functional consequences via interactions distal to the lncRNA genomic location

Ulcerative colitis

An inflammatory bowel disease (IBD) caused by increased colon inflammation

X-chromosome inactivation

The transcriptional silencing of one X chromosome in somatic female mammalian cells that is dependent on the lncRNA XIST and balances gene dosage between males and females