Translating the ribosome’s grim role

A large number of long noncoding RNAs (lncRNA) have been found associating with the ribosome, the protein-making machinery in the cytoplasm. What the so-called ‘noncoding’ RNAs are doing on the ribosome, whose main job is to translate RNA into protein, has puzzled the A*STAR researchers who discovered them.

The answer could be macabre, suggest cell biologist, Leah Vardy, at the A*STAR Institute of Medical Biology and bioinformatician, Rory Johnson, at the University of Bern, Switzerland. Translation by the ribosome could lead to the degradation of these lncRNAs, says Vardy. “The ribosome may be acting as a garbage dump or graveyard for lncRNAs at the end of their life.”

In recent years, researchers have found that many RNAs previously labeled as junk, actually play an important role in causing a wide range of diseases, from cancers to metabolic disease and neurodegeneration. Still, many questions remain about their basic biology, with studies covering less than one per cent of the tens of thousands of lncRNAs in our genomes. “LncRNAs are one of the most promising avenues for understanding disease today,” says Johnson.

One question that confounded Vardy and Johnson was why lncRNAs were in the cytoplasm when they were thought to be confined to the nucleus. By applying a technique for studying messenger RNA translation, they discovered that most of the lncRNAs in the cytoplasm were bound to ribosomes. “This was surprising because, by definition, lncRNAs were not thought to be translated,” says Johnson.

To make sense of their results, the researchers tinkered with the translation process to see how it would affect the lncRNAs. The lncRNAs decayed very quickly in normal cells, but when the researchers used a drug to block translation, they decayed at a much slower rate. “Some ribosome-bound lncRNAs become more stable when translation is inhibited,” explains Vardy.

Discovery and quantification of ribosome-associated lncRNAs by polysome profiling and microarray hybridization

long non-coding rna

(A) Outline of the subcellular mapping of K562 lncRNA by polysome profiling and microarray hybridization. Sucrose-gradient ultracentrifugation was used to isolate the indicated fractions of ribosome-associated RNA, quantifications of which are displayed at the upper right. The pooled fractions used in this study are shown below the figure. The total amount of RNA isolated from each fraction is indicated by arrows, from which 0.1 µg was collected and hybridized to custom lncRNA microarrays. Microarrays were normalized using spike-ins: At the lower left is shown a representative example of the linear regression of spike in probe intensity against their starting concentrations. Dashed red line represents the defined detection threshold for this fraction where regression ceases to be linear. Only probes above this threshold were considered detected. (B) Correlation of the sum of the three cytoplasmic fraction concentration estimates and total cytoplasmic concentration estimate, supporting the quantification approach used. (C) Barplot shows for 14 lncRNA examples the relative amount (expressed as a percentage) of transcript molecules estimated to be present in each of the fractions. Sum of percentages of the three fractions has to be 100%, the total of detected molecules in the cytoplasm. Left bars represent quantification by microarrays and right bars by the mean of two quantitative PCR biological replicates. Microarray and PCR experiments represent different biological replicates.

The findings suggest that ribosomes are where some lncRNAs may go to die, a process similar to the well-known mechanism of ‘nonsense mediated decay’, in which ribosomes promote the degradation of malformed messenger RNAs. The researchers found that ribosome-bound lncRNAs look more “mRNA like” than their free-floating counterparts in the cytoplasm. “In some ways, lncRNAs look like malformed mRNAs, and for this reason may enter the nonsense mediated decay pathway,” explains Johnson.

To confirm their hypotheses about the fatal affair between noncoding RNAs and protein-coding ribosomes, Vardy and Johnson plan to study the relationship at an individual level to understand how they are regulated and whether they perform both coding and non-coding functions.

The A*STAR-affiliated researchers contributing to this research are from the Institute of Medical Biology. For more information about the team’s research, please visit the Epidermal Gene Regulation webpage

Carlevaro-Fita J, Rahim A, Guigó R, Vardy LA, Johnson R. (2016) Cytoplasmic long noncoding RNAs are frequently bound to and degraded at ribosomes in human cells. RNA 22(6):867-82. [article]

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