Cytoplasmic long noncoding RNAs are frequently bound to and degraded at ribosomes in human cells.

Joana Carlevaro-Fita,Roderic Guigó,Leah A Vardy,Anisa Rahim,Rory Johnson

doi:10.1261/rna.053561.115

Abstract

Recent footprinting studies have made the surprising observation that long noncoding RNAs (lncRNAs) physically interact with ribosomes. However, these findings remain controversial, and the overall proportion of cytoplasmic lncRNAs involved is unknown. Here we make a global, absolute estimate of the cytoplasmic and ribosome-associated population of stringently filtered lncRNAs in a human cell line using polysome profiling coupled to spike-in normalized microarray analysis. Fifty-four percent of expressed lncRNAs are detected in the cytoplasm. The majority of these (70%) have >50% of their cytoplasmic copies associated with polysomal fractions. These interactions are lost upon disruption of ribosomes by puromycin. Polysomal lncRNAs are distinguished by a number of 5′ mRNA-like features, including capping and 5′UTR length. On the other hand, nonpolysomal “free cytoplasmic” lncRNAs have more conserved promoters and a wider range of expression across cell types. Exons of polysomal lncRNAs are depleted of endogenous retroviral insertions, suggesting a role for repetitive elements in lncRNA localization. Finally, we show that blocking of ribosomal elongation results in stabilization of many associated lncRNAs. Together these findings suggest that the ribosome is the default destination for the majority of cytoplasmic long noncoding RNAs and may play a role in their degradation.

Highlights

The past decade has witnessed the discovery of tens of thousands of long non-protein-coding RNAs in our genome, with profound implications for our understanding of molecular genetics, disease, and evolution
We show evidence that long non-protein-coding RNAs (lncRNAs) can be divided into classes based on ribosomal association, and these classes are distinguished by a variety of features, most notably transposable element insertions and mRNA-like features at the 5′ end
Consistent with previous studies (Zhang et al 2012; Wong et al 2016), extracts were divided into three pools: “heavy polysomal,” corresponding to high molecular weight complexes cofractioning with greater than six ribosomes; “light polysomal,” cofractioning with two to six ribosomes; and low-molecular weight complexes corresponding to nontranslated, cytoplasmic RNAs (Fig. 1A)

Summary

Introduction

The past decade has witnessed the discovery of tens of thousands of long non-protein-coding RNAs (lncRNAs) in our genome, with profound implications for our understanding of molecular genetics, disease, and evolution. Focus is shifting to understanding the function of these molecules, which is likely to be intimately linked to localization within the cell. Following the first compelling discoveries of chromatin regulatory lncRNAs such as XIST (Brown et al 1991) and AIR (Wutz et al 1997; Lyle et al 2000), a paradigm was established for lncRNAs as nuclear-restricted, epigenetic regulatory molecules (Khalil et al 2009). Growing evidence points to lncRNAs having diverse roles outside of Article published online ahead of print. Available online through the RNA Open Access option

Methods

Results

Conclusion