Investigating the competing endogenous RNA hypothesis genome-wide and in single cells

The observation that microRNAs (miRNAs), through a titration mechanism can couple interactions of their common targets (competing endogenous RNAs or ceRNAs) has prompted a general "ceRNA hypothesis" that RNAs can regulate each other indirectly through global RNA-miRNA-RNA networks. These ceRNAs are said to "crosstalk" with each other by competing for common miRNAs. Although many individual ceRNAs have been found, fundamental questions about both the magnitude and generality of the crosstalk effect remain. In our study we combine RNA sequencing and single-molecule FISH (smFISH) approaches to both measure the magnitude of the crosstalk effect genome-wide by perturbing three known ceRNAs (Pten, Vapa, Cnot6l) and to identify mechanisms by which the crosstalk effect acts. We identify hundreds of putative ceRNAs and dissect the contributions of individual miRNAs in transmitting crosstalk. We demonstrate that while the crosstalk effect is pervasive, it nevertheless remains bounded by the size of the perturbation. Furthermore, we show that both the number and affinity of shared miRNA binding sites between targets is crucial in determining the magnitude of the crosstalk strength. Using the smFISH data, we examined the single-cell gene expression profiles of pairs of ceRNAs and found that ceRNA gene expression is correlated only in the presence of active miRNAs. Additionally, on inspecting the intra-cellular localization of RNA molecules, we found a miRNA-dependent colocalization of ceRNAs, suggesting a new signature of crosstalk between ceRNAs that extends and modifies the original hypothesis.

Department dc:contributor.department

Massachusetts Institute of Technology. Department of Physics.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2015

Author dc:creator

• Sahay, Apratim

Advisor dc:contributor.advisor

• Alexander van Oudenaarden and Jeff Gore.

• Physics.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/99290

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/99290