LNCcation: lncRNA localization and function

Abstract

Subcellular localization of RNAs has gained attention in recent years as a prevalent phenomenon that influences numerous cellular processes. This is also evident for the large and relatively novel class of long noncoding RNAs (lncRNAs). Because lncRNAs are defined as RNA transcripts >200 nucleotides that do not encode protein, they are themselves the functional units, making their subcellular localization critical to their function. The discovery of tens of thousands of lncRNAs and the cumulative evidence involving them in almost every cellular activity render assessment of their subcellular localization essential to fully understanding their biology. In this review, we summarize current knowledge of lncRNA subcellular localization, factors controlling their localization, emerging themes, including the role of lncRNA isoforms and the involvement of lncRNAs in phase separation bodies, and the implications of lncRNA localization on their function and on cellular behavior. We also discuss gaps in the current knowledge as well as opportunities that these provide for novel avenues of investigation.

Figure 1.

Localization of lncRNAs to organelles and macromolecular structures. lncRNAs can function distinctly and have different interaction partners based on localization. The lncRNA PYCARD-AS1 suppresses PYCARD mRNA transcription in the nucleus via promotor binding and G9a, DNMT1 recruitment; in the cytoplasm, PYCARD-AS1 binds PYCARD mRNA, preventing ribosome assembly. While RMRP is essential for processing of preribosomal RNA in the nucleolus, RMRP binds GRSF in the mitochondria to maintain structure and mediate oxidative phosphorylation and mitochondrial DNA replication. The cAMP-dependent up-regulation of LINC00473 directs shuttling to the cytoplasm and localization at the mitochondria-lipid droplet interface, which regulates lipolysis and mitochondrial function. lncRNAs with established roles in the nucleus (TUG1, regulation of transcription) and cytosol (NORAD, PUMILIO sequestering) have been identified in the ER, but ER-specific roles have not been characterized. lncRNAs also associate with ribosomes. While some may code for small peptides, lncRNAs may also be degraded at ribosomes through NMD; GAS5 is an lncRNA that undergoes NMD at ribosomes. SG- and PB-associated lncRNAs have been identified but not functionally characterized. lncRNA recruitment to SGs and PBs may be mediated by interactions with RBPs (RBPs in PBs: IGF2BP1 and HuR; RBPs in SGs: TIA-1 and TIAR). NORAD SG recruitment is regulated by eIF4A, limiting RNA condensation. MALAT1 in nuclear speckles is key for recruitment of splice factors (SRF1, 2, and 3), and NEAT1 is essential for paraspeckle formation, sequestering RNAs and protein, and facilitating miRNA processing via NONO/PSF/microprocessor interactions. MALAT1, TERRA, and H19 were shown to be packaged in exosomes, and the lncRNA LASSIE stabilizes endothelial adherens junctions via association with PECAM-1. Compartments labeled with red text indicate phase-separated structures. Asterisks indicate lncRNAs with uncharacterized functions at the cellular location. RNAs and structures are not drawn to scale and are depicted as they are for artistic purposes.

Figure 2.

Sequence elements and mechanisms that regulate subcellular localization of lncRNAs. Sequence features can promote the nuclear retention or cytoplasmic enrichment of lncRNAs. Examples of nuclear retention elements are indicated by asterisks and include motifs repurposed from transposable elements, known as RIDLs, a pentamer motif in BORG, and a highly structured repeat in FIRRE, which is bound by heterogeneous nuclear ribonucleoprotein U, facilitating chromatin interaction. Loss of the pentamer motif in BORG leads to cytoplasmic export, while the nuclear retention element in XIST sequesters cytoplasmic RNAs to the nucleus. GC-rich RIDLs are associated with lncRNA cytoplasmic enrichment. Inefficient splicing may promote nuclear retention, while inefficiently spliced transcripts may use NXF1 for cytoplasmic export. RNA Pol II pausing is associated with cytoplasmic export of lncRNAs with poor splice site conservation. While m6A modification is a fast-track cytoplasmic export signal in mRNAs, it is unclear how it acts on lncRNAs. circRNAs that are >800 nt are efficiently exported to the cytoplasm through UAP56, whereas shorter ones (<800 nt) are exported through URH49. Questions remain about targeting of lncRNAs to distinct organelles or domains, such as shuttling between the nucleus and mitochondria; RBPs may play a role. The binding of HuR to RMRP in the nucleus facilitates export via CRM1, and association with GSRF influences mitochondrial localization. Stem loop structures in RNAs may facilitate mitochondrial import via PNPASE. lncRNAs associated with ribosomes have longer 5′ UTRs and are deplete of repetitive elements. Asterisks indicate nuclear retention elements. RNAs and structures are not drawn to scale and are depicted as they are for artistic purposes.

Figure 3.

lncRNAs are alternatively spliced or processed into isoforms with potential for distinct subcellular localization and function. (A)GAS5 is an lncRNA that undergoes extensive alternative splicing into multiple isoforms with currently unexplored subcellular localization patterns and functions. (B) The cytoplasmic, human-specific, alternatively spliced variant of FAST, hFAST, is required for stemness, but not the nuclear, mouse-specific, mFast variant. (C) Alternative splicing of PXN-AS1 results in two cytoplasmic isoforms that promote (PXN-AS1-L)or inhibit (PXN-AS1-S) PXN translation. (D) Alternative 3′ end processing of CCAT1 generates a nuclear specific isoform that promotes MYC transcription (CCAT1-L)or a shorter one that is exported to the cytoplasm (CCAT1-S). (E)Due to alternative transcription terminationsites, the NEAT1 lncRNA exists in two isoforms: a shorter, polyadenylated, and abundant NEAT1_1 isoform localizes in nuclear foci distinct from paraspeckles termed microspeckles, and a longer, cell type–specific and paraspeckle-specific, NEAT1_2 isoform. Background genomic and transcript information were obtained through the University of California, Santa Cruz, Genome Browser (GRCh38/hg38 assembly). Specific isoforms discussed in the text are designed in each case with their referenced names and subcellular localization.