Kanduri, C. (2016). Long non-coding RNAs: lessons from genomic imprinting. Biochimica and Biophysica Acta.

Introduction

• In gametogenesis ~1% of protein-coding genes undergo genomic imprinting
  • >150 imprinted genes identified in mouse
  • Typically located in clusters that range from a few kB to 3.0 Mb
• Long non-coding RNAs found in all imprinted clusters
  • Inverse expression pattern relative to protein-coding counterparts
  • Promoters map to differentially methylated regions (DMRs)
  • Deletion of the DMRs often leads to loss of imprinting
  • ICRs: imprinting control regions, 1-3 kb in size, most DMRs are ICRs
• Human genome has more lncRNAs than protein-coding genes
  • Perform various functions in development, differentiation, disease
  • Multiple mechanisms both at transcriptional and post-transcriptional level
  • Most target chromatin modifying complexes e.g. PRC2, SW1/SNF, hnRNPK, G9a
  • Implicated in gene regulation at post-transcriptional level when they are localized to the cytoplasm

Da Sacco et al. (2012). IJMS. Pie chart of the major categories of >16,000 non-coding RNAs in the genome.

Intergenic lncRNAs in Genomic Imprinting

• lncRNAs can be classified into 4 main categories: intergenic, antisense, intronic, and enhancer
  • All except intronic implicated in imprinting and parent-of-origin specific expression
• H19: lncRNA (2.3 kb in length)
  • Maps to a well-investigated cluster on mouse chromosome 7, human chromosome 11
  • only expressed from the maternal allele, silenced on paternal
  • Paternal allele silenced via CpG methylation at the promoter
  • May have lineage-specific roles in the body
    • Deletion of H19 affected Igf2 in mesoderm but not endoderm
    • Deletion also has effects on growth, but is not embryonic lethal
  • Part of IGN (imprinted gene network) that includes 16 genes
  • Potentially controls growth via regulation of the IGN in trans
  • Interacts with MBD1, a methyl CpG-binding protein
    • Complex recruits H3K9 methyltransferase to DMRs of some members of the gene network
    • Establishes H3K9me3 marks to “fine tune” expression from both parental allelles
  • Expressed at high level in embryogenesis, downregulated after birth
    • Exception: remains highly expressed in muscle tissue
    • May promote myogenic differentiation?
  • Also shown to have oncogenic, tumour-suppressive properties
    • Overexpression of H19 is linked to metastasis
• IPW: paternally expressed lncRNA, maps to an imprinting cluster on mouse chromosome 7, human chromosome 15
  • Deletion observed in 70% of cases of Prader-Willi syndrome patients
  • Mouse expression mainly restricted to the brain, but humans seen in all tissues
  • 5′ end contains tandem repeats
  • functional role of IPW cluster has not been investigated, but shown to interact with G9a methyltransferase
    • Targets IG-DMR to modify chromatin structure via H3K9
    • IG-DMR is a master controller of gene expression at the DLK-D103 imprinted cluster
    • First example of a lncRNA shown to promote chrosstalk between two imprinting clusters by altering ICR chromatin
• MEG3: maternally expressed gene 3, imprinted lncRNA, maps to DLK-DI03 locus on human chromosome 14, mouse chromosome 12
  • IG-DMR controls maternal specific expression of MEG3
  • MEG3 expression is a marker of iPSCs with a fully pluripotent state
  • iPSCs without MEG3 expression are not viable to support embryonic development
  • MEG3 may promote interaction of PRC2 with JARID2
  • Complex of PRC2-JARID2 shown to contribute to ESC differentiation
    • MEG3 may therefore underlie the fully pluripotent state

Enhancer RNAs in Genomic Imprinting

• Enhancers are responsible for spatio-temporal gene regulation
  • “landing site” for transcription factors, co-activator complexes
  • Can activate or increase transcription from distal promoters
• Characteristics of enhancers include:
  • DNase I hypersensitivity
  • Post-translational histone modifications (especially H3K4me1/2, H3K27ac)
  • Bidirectional transcription
  • Transcripts generated are low copy number, non-polyadenylated
• Enhancers promote target gene expression via recruitment or stabilization of basic transcription machinery binding
  • Establish higher-order chromatin contacts between enhancer and targets
  • Transcripts from IG-DMR control expression of maternally expressed transcripts at DLK1-Dio3 locus
  • Methylated on the paternal chromosome, unmethylated on the maternal
  • Unmethylated version is critical for expression of multiple lncRNAs including MEG3
    • Therefore acts as putative enhancer with enhancer-specific histone marks, encodes bidirectionally transcribed ncRNAs
• Maternal chromosome bidirectional transcription in ESCs correlates with early replication, inner subnuclear positioning of Dlk1-Dio3 locus
  • IG-DMR transcripts may promote higher order chromatin
  • Enables early replication, subnuclear localization
  • Maintenance of expression of maternally expressed genes