Differential Allelic Expression among Long Non-Coding RNAs

Long non-coding RNAs (lncRNA) comprise a diverse group of non-protein-coding RNAs >200 bp in length that are involved in various normal cellular processes and disease states, and can affect coding gene expression through mechanisms in cis or in trans. Since the discovery of the first functional lncRNAs transcribed by RNA Polymerase II, H19 and Xist, many others have been identified and noted for their unusual transcriptional pattern, whereby expression from one chromosome homolog is strongly favored over the other, also known as mono-allelic or differential allelic expression. lncRNAs with differential allelic expression have been observed to play critical roles in developmental gene regulation, chromosome structure, and disease. Here, we will focus on known examples of differential allelic expression of lncRNAs and highlight recent research describing functional lncRNAs expressed from both imprinted and random mono-allelic expression domains.

Targeted Double-Stranded cDNA Sequencing-Based Phase Analysis to Identify Compound Heterozygous Mutations and Differential Allelic Expression

There are two combinations of heterozygous mutation, i.e., in trans, which carries mutations on different alleles, and in cis, which carries mutations on the same allele. Because only in trans compound heterozygous mutations have been implicated in autosomal recessive diseases, it is important to distinguish them for clinical diagnosis. However, conventional phase analysis is limited because of the large target size of genomic DNA. Here, we performed a genetic analysis on a patient with Wilson disease, and we detected two heterozygous mutations chr13:51958362;G>GG (NM_000053.4:c.2304dup r.2304dup p.Met769HisfsTer26) and chr13:51964900;C>T (NM_000053.4:c.1841G>A r.1841g>a p.Gly614Asp) in the causative gene ATP7B. The distance between the two mutations was 6.5 kb in genomic DNA but 464 bp in mRNA. Targeted double-stranded cDNA sequencing-based phase analysis was performed using direct adapter ligation library preparation and paired-end sequencing, and we elucidated they are in trans compound heterozygous mutations. Trio analysis showed that the mutation (chr13:51964900;C>T) derived from the father and the other mutation from the mother, validating that the mutations are in trans composition. Furthermore, targeted double-stranded cDNA sequencing-based phase analysis detected the differential allelic expression, suggesting that the mutation (chr13:51958362;G>GG) caused downregulation of expression by nonsense-mediated mRNA decay. Our results indicate that targeted double-stranded cDNA sequencing-based phase analysis is useful for determining compound heterozygous mutations and confers information on allelic expression.