Quantitative interaction screen of telomeric repeat-containing RNA reveals novel TERRA regulators
- Marion Scheibe1,
- Nausica Arnoult2,
- Dennis Kappei3,
- Frank Buchholz3,
- Anabelle Decottignies2,
- Falk Butter1,4 and
- Matthias Mann1
- ↵* Corresponding author; email: butter{at}biochem.mpg.de
Abstract
Telomeres are actively transcribed into telomeric repeat-containing RNA (TERRA), which has been implicated in the regulation of telomere length and heterochromatin formation. Here, we applied quantitative mass spectrometry (MS)-based proteomics to obtain a high confidence interactome of TERRA. Using SILAC-labeled nuclear cell lysates in an RNA pull-down experiment, and two different salt conditions, we distinguished 115 proteins binding specifically to TERRA out of a large set of background binders. While TERRA binders identified in two previous studies showed little overlap, using quantitative mass spectrometry we obtain good overlap to both studies. To test whether novel candidates found here are involved in TERRA regulation, we performed an esiRNA-based interference analysis for 15 of them. Knock-down of ten proteins significantly affected total cellular levels of TERRA and RNAi of five candidates perturbed TERRA recruitment to telomeres. Notably, depletion of SRRT/Ars2, involved in miRNA processing, up-regulated both total and telomere-bound TERRA. Conversely, knock-down of MORF4L2, a component of the NuA4 histone acetyltransferase complex, reduced TERRA levels both globally and for telomere-bound TERRA. We thus identified new proteins involved in the homeostasis and telomeric abundance of TERRA, extending our knowledge of TERRA regulation.
- Received November 13, 2012.
- Accepted July 30, 2013.
- © 2013, Published by Cold Spring Harbor Laboratory Press
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