Chromatin-associated RNAs as facilitators of functional genomic interactions

Abstract

Mammalian genomes are extensively transcribed, which produces a large number of both coding and non-coding transcripts. Various RNAs are physically associated with chromatin, through being either retained in cis at their site of transcription or recruited in trans to other genomic regions. Driven by recent technological innovations for detecting chromatin-associated RNAs, diverse roles are being revealed for these RNAs and associated RNA-binding proteins (RBPs) in gene regulation and genome function. Such functions include locus-specific roles in gene activation and silencing, as well as emerging roles in higher-order genome organization, such as involvement in long-range enhancer-promoter interactions, transcription hubs, heterochromatin, nuclear bodies and phase transitions.

Fig. 1 |. Different modes of RNA-chromatin interactions.

a | Nascent RNAs can form R-loops, which predominately occur at promoters and enhancers (top). Given divergent transcription from many promoters and most enhancers, R-loops may consist of divergent transcripts (bottom) that could jointly contribute to the formation and dynamics of the resulting R-loops^–. b | Nascent RNA is targeted by amplified small RNAs on its site of transcription, a process frequently associated with heterochromatin formation^–. c | Nascent pre-mRNA remains on chromatin for co-transcriptional processing (top) and RNA modification (bottom), which may play important roles in feedback control of transcription^,,,,. d | Trans-acting RNA may form a triplex structure on double-stranded DNA using the Hoogsteen base-pairing rule, a strategy exploited by various long non-coding RNAs^–. e | RNA serves as a scaffold to nucleate the formation of specific protein complexes on chromatin to control gene expression by promoting enhancer-promoter interactions, facilitating chromatin remodelling or directly contributing to specific chromatin modification activities^–,,,. CTCF, CCCTC-binding factor; dsRNA, double-stranded RNA; eRNA, enhancer RNA; Pol II, RNA polymerase II; RBP, RNA-binding protein; RITS, RNA-induced transcriptional silencing complex; siRNA, small interfering RNA.

Fig. 2 |. Strategies for global analysis of RNA-chromatin interactions.

a | One-to-many approach based on RNA capture followed by deep sequencing of associated DNA fragments^–. b | All-to-all strategy through in situ linker ligation to RNA and DNA on fixed nuclei^–. c | A split-pool strategy to barcode isolated nuclear particles, thus enabling the profiling of RNA-RNA, RNA-DNA and DNA-DNA interactions. d | Locus-specific capture either through a hybridized probe or using a nuclease-dead Cas9 (dCas9)-based method, which can be used for de novo identification of RNAs and proteins on specific genomic loci. gRNA, guide RNA; RT, reverse transcription.

Fig. 3 |. Nascent-RNA-decorated transcription hubs and a model for RNA-nested hub formation.

a | Deduced transcription hubs on a human chromosome (chromosome 11) in MDA-MB-231 breast cancer cells based on the RNA-chromatin interactome detected by global RNA interaction with DNA sequencing (GRID-seq). b | ‘RNA cloud’ on a transcription hub, suggesting a potential role of nascent RNA in networking regulatory DNA elements in the hub^,. Also illustrated is the potential local recycling of RNA polymerase II and other transcription factors within the hub. RNP, ribonucleoprotein particle.

Fig. 4 |. Roles of chromatin-associated nascent RNA in transcriptional activation or repression.

a | Formation of an R-loop in the promoter proximal region leads to transcriptional repression through pausing of RNA polymerase II (Pol II) (left). R-Loop-dependent recruitment of transcriptional co-activators may contribute to transcriptional pause release (right). b | Specific sequences from nascent RNA induce translocation of HIV Tat or cellular SRSF2 from the 7SK complex to the nascent RNA (left), thereby relocating the Pol II C-terminal domain (CTD) kinase pTEFb from the inhibitory 7SK complex to the paused Pol II complex to drive transcription pause release (right)^–. c | Two competing models for nascent-RNA-dependent recruitment of Polycomb repressive complex 2 (PRC2) to provide a feedback control mechanism for gene expression. One model proposes that nascent RNA competes with nearby chromatin for PRC2 binding (left)^,,–, whereas the other suggests that PRC2 may use nascent RNA as a stepping stone to catalyse trimethylation of K27 on histone H3 (H3K27me3) (right)^,,. These two models appear to respectively apply to highly active promoters predominately associated with trimethylation of K4 on histone H3 (H3K4me3) and bivalent promoters marked with both H3K4me3 and H3K27me3. ESE, exonic splicing enhancer; SR, splicing factor characterized by repeats of serine (S) and arginine (R) amino acids; TAR, transactivation-responsive region.

Fig. 5 |. RNA-dependent and RNA-independent feedback loops for establishing and locally spreading heterochromatin.

a | In fission yeast, RNA-dependent RNA polymerase (RdRP)-dependent amplification of local repeat-derived RNA occurs and then Dicer-processed small interfering RNAs (siRNAs) target nascent RNA to induce a network of interactions, resulting in the recruitment of SUV39 to deposit H3K9me on chromatin. This inhibitory histone mark next recruits heterochromatin protein 1 (HP1) and, together with SUV39, facilitates H3K9me spreading into adjacent genomic regions^–,,. b | In Drosophila melanogaster, the primary transposon transcript is processed into Piwi-interacting RNAs (piRNAs), which then target nascent RNA to recruit SUV39 and HP1 to establish and spread methylation of K9 on histone H3 (H3K9me)^,,–. Ago3, Argonaute 3; Aub, Aubergine; dsRNA, double-stranded RNA; Pol II, RNA polymerase II; RISC, RNA-induced silencing complex; RITS, RNA-induced transcriptional silencing complex.

Fig. 6 |. RNAs in liquid-liquid phase separation and nuclear body formation.

a | Multivalent interactions in low-complexity domain (LCD)-containing proteins promote the formation of liquid droplets. Their interactions with RNAs may then partition liquid droplets into spatially distinct domains^,,. b | Scheme of dynamic liquid droplets in three different phases in the nucleolus for ribosomal DNA (rDNA) transcription (in the fibrillar centre (FC) domain), ribosomal RNA (rRNA) processing (in the dense fibrillar compartment (DFC) domain) and ribosome assembly (in the granule compartment (GC) domain)^,–. c | Both locally produced and trans-supplied small nuclear RNAs (snRNAs) and histone mRNAs drive the formation of liquid droplets in Cajal bodies^–. d | Formation of nuclear speckles induced by co-transcriptional and post-transcriptional pre-mRNA processing. Paraspeckles are known to form adjacently to nuclear speckles, and a large long non-coding RNA NEAT1 is essential for the formation and maintenance of this nuclear subdomain. The connection between nuclear speckles and paraspeckles might result from a phase separation induced by primary microRNA (pri-miRNA) processing from various introns^,,,. FIB1, fibrillarin; NPM1, nucleoplasmin; NCL, nucleolin; Pol I, RNA polymerase I; Pol II, RNA polymerase II; Pol III, RNA polymerase III; RBP, RNA-binding protein.