Table 3.

Profiling lincRNA biogenesis, coding potential and interactomes

Method	Description	Discovery
Protein-centric
nRIP, nRIP-seq	nRIP followed by PCR or RNA-sequencing Antibody-based complex isolation	Xist, Tsix, RepA70 and HOTAIR interact with PRC2 ( REF. 74); >20% of lncRNAs interact with PRC2 ( REF. 74) Novel lncRNA discovery73
CLIP CLIP-seq, irCLIP	Ultraviolet crosslinking followed by immunoprecipitation In irCLIP, infrared-dye-conjugated adaptors enable reduced input157 Lower background than nRIP158	Firre interacts with hnRNPU to affect nuclear architecture81
RNA-centric
RNA pull-down	An RNA probe is used to isolate RNA with associated proteins	Initial description of HOTAIR-PRC2 interaction12 LincRNA-p21 interacts with hnRNPK to modulate p53 activity159
ChIRP. dChIRP	Biotin-labelled oligonucleotides are tiled across the entire RNA transcriptome; the RNA is isolated using streptavidin and identified by sequencing, western blotting or mass spectrometry100,160 In dChIRP, domain-specific oligonucleotides are used to define the RNA segment involved161	Definition of HOTAIR genome occupancy HOTAIR nucleates Polycomb domains160
CHART	Probes identified using RNase H are used to empirically determine hybridization region	Definition of MALAT1 genome occupancy; enrichment at active genes and for nuclear paraspeckle components162
RAP	Oligonucleotide hybridization using long probes (>60 nucleotides); more stable isolation	Xist depletion at the FIRRE locus, suggesting that FIRRE escapes X chromosome inactivation81 SHARP protein as Xist-specific interactor required for Pol II exclusion from Xi; with HDAC3, required for PRC2 recruitment163

With increasing evidence for long intergenic non-coding RNA (lincRNA)-encoded micropeptides, attention has turned towards estimations of small open reading frame coding potential within lincRNAs by sequence conservation¹⁶⁴ or ribosomal profiling^124,165. Orthogonal approaches to predict effects of non-coding variants using deep convolutional neural networks have been proposed¹⁶⁶. Multiple methods to characterize lincRNA interactomes have been described; several are highlighted here. CHART, capture hybridization of RNA targets; ChIRP, chromatin isolation by RNA purification; CLIP, crosslinking immunoprecipitation; dChIRP, domain-specific ChIRP; HDAC3, histone deacetylase 3; irCLIP, infrared CLIP; lncRNA, long non-coding RNA; nRIP, native RNA precipitation; nRIP-seq, nRIP followed by sequencing; Pol II, RNA polymerase II; RAP, RNA antisense purification; SHARP, enhancer-of-split and hairy-related protein (also known as BHLHE41); Xi, inactive X chromosome.