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RNA interference machinery influences the nuclear organization of a chromatin insulator

Abstract

RNA interference (RNAi) is a conserved silencing mechanism that can act through alteration of chromatin structure. Chromatin insulators promote higher-order nuclear organization, thereby establishing DNA domains subject to distinct transcriptional controls. We present evidence for a functional relationship between RNAi and the gypsy insulator of D. melanogaster. Insulator activity is decreased when Argonaute genes required for RNAi are mutated, and insulator function is improved when the levels of the Rm62 helicase, involved in double-stranded RNA (dsRNA)-mediated silencing and heterochromatin formation, are reduced. Rm62 interacts physically with the DNA-binding insulator protein CP190 in an RNA-dependent manner. Finally, reduction of Rm62 levels results in marked nuclear reorganization of a compromised insulator. These results suggest that the RNAi machinery acts as a modulator of nuclear architecture capable of effecting global changes in gene expression.

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Acknowledgements

We would like to thank A. Beyer for α-Pep and F. Fuller-Pace for α-p68; J. Birchler, S. Hou, H. Lin, and P. Macdonald for strains and Y. Zheng for fly cages. We are indebted to E. Baxter for assistance with population cages; M. Capelson and members of the Corces laboratory for discussions and J. Birchler, M. Capelson, and C. Karam for comments on the manuscript. E.P.L. is a fellow of The Jane Coffin Childs Memorial Fund for Medical Research. This work was supported by grants from the US National Institutes of Health to V.G.C.

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Competing interests

The authors declare no competing financial interests.

Correspondence to Victor G Corces.

Supplementary information

  1. Supplementary Fig. 1

    Localization of insulator proteins to a site of gypsy insertion in RNAi mutants. (PDF 217 kb)

  2. Supplementary Fig. 2

    Nuclear localization of Rm62 in diploid cells. (PDF 215 kb)

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Figure 1: The insulator protein CP190 interacts with the RNA helicase Rm62 in an RNA-dependent manner.
Figure 2: Mutations in genes required for RNAi affect gypsy insulator function.
Figure 3: Localization of myc-Piwi and insulator proteins on polytene chromosomes.
Figure 4: Localization of insulator proteins on polytene chromosomes of RNAi mutants.
Figure 5: Insulator body nuclear organization is altered in RNAi mutants.
Figure 6: Model for how RNAi affects gypsy insulator chromatin domain formation.