Unusual Processing Generates SPA LncRNAs that Sequester Multiple RNA Binding Proteins

Abstract

We identify a type of polycistronic transcript-derived long noncoding RNAs (lncRNAs) that are 5' small nucleolar RNA (snoRNA) capped and 3' polyadenylated (SPAs). SPA processing is associated with nascent mRNA 3' processing and kinetic competition between XRN2 trimming and Pol II elongation. Following cleavage/polyadenylation of its upstream gene, the downstream uncapped pre-SPA is trimmed by XRN2 until this exonuclease reaches the co-transcriptionally assembled snoRNP. This snoRNP complex prevents further degradation, generates a snoRNA 5' end, and allows continuous Pol II elongation. The imprinted 15q11-q13 encodes two SPAs that are deleted in Prader-Willi syndrome (PWS) patients. These lncRNAs form a nuclear accumulation that is enriched in RNA binding proteins (RBPs) including TDP43, RBFOX2, and hnRNP M. Generation of a human PWS cellular model by depleting these lncRNAs results in altered patterns of RBPs binding and alternative splicing. Together, these results expand the diversity of lncRNAs and provide additional insights into PWS pathogenesis.

The functions and unique features of long intergenic non-coding RNA

“…An intriguing class of lncRNAs termed 5′ small nucleolar RNA-capped and 3′ polyadenylated (SPAs) were recently implicated in the pathogenesis of syndrome. SPAs affect the binding of several RNA-binding proteins, including TDP43 and RBFOX2, to their RNA interactors, thereby supporting alternative splicing patterns that are consistent with the disease state 64 . Additionally, principal component analysis of differential lncRNA gene expression and differential splicing in autism has revealed a high correlation between altered expression and splicing activity, again suggesting a role for lncRNAs in modulating splicing activity with functional impact 65 .…”

The Output of Protein-Coding Genes Shifts to Circular RNAs When the Pre-mRNA Processing Machinery Is Limiting

“…Once produced, some readthrough transcripts have been proposed to help maintain the nuclear scaffold during osmotic stress (Vilborg et al, 2015). Others serve as precursors to stable RNAs, including Drosophila piRNAs (Chen et al, 2016; Mohn et al, 2014) and human SPA long noncoding RNAs (Wu et al, 2016) that lack their own promoter sequences.…”

“…shRNAs to deplete CPSF3 or CPSF4 from PA1 cells and Western blot procedure were previously described (Wu et al, 2016). Phosphorothioate-modified antisense oligodeoxynucleotides (ASOs) were synthesized at BioSune (Shanghai, China) and introduced to PA1 cell by nucleofection (Lonza) according to the manufacturer’s protocol.…”

Major hnRNP proteins act as general TDP-43 functional modifiers both in Drosophila and human neuronal cells

“…It has been described, in fact, that both TDP-43 and FUS can interact with several lncRNAs and can regulated both their functions and stability (75). In particular, TDP-43 has been shown to interact and regulate important lncRNAs such as gadd7 (76), MALAT1 (77), NEAT1-2 (78), transposable elements general (79) and two SPA lncRNA involved in Prader-Willi syndrome (80). How these hnRNPs may help or hinder TDP-43 in regulating these molecules is something that will need to be determined.…”