Advertisement

Assessing Intermolecular RNA:RNA Interactions Within a Ribonucleoprotein Complex Using Heavy Metal Cleavage Mapping

  • Keith T. Gagnon
  • E. Stuart Maxwell
Part of the Methods in Molecular Biology book series (MIMB, volume 1240)

Abstract

Heavy metal cleavage mapping analysis of both assembling and fully mature ribonucleoprotein (RNP) complexes are informative techniques for assessing the intermolecular base pairing between small non-coding RNAs and their interacting target RNAs. Lead cleavage of the RNA in partially or fully assembled RNPs in the absence or presence of the interacting RNA can determine both the accessibility of the base pairing sequence within the RNP itself as well as its interaction with the target RNA. In this chapter, we detail how this technique was used to map the intermolecular RNA:RNA base pairing of a box C/D RNA with its target RNA within the assembling archaeal RNP complex.

Key words

Intermolecular RNA:RNA base pairing Lead cleavage mapping RNP assembly Gel electrophoresis Non-coding RNA 

References

  1. 1.
    Valadkhan S (2010) Role of the snRNAs in spliceosomal active site. RNA Biol 7:345–353PubMedCrossRefGoogle Scholar
  2. 2.
    Bartel D (2004) MiRNAs: genomics, biogenesis, mechanism, and function. Cell 116:281–297PubMedCrossRefGoogle Scholar
  3. 3.
    Terns M, Terns R (2002) Small nucleolar RNAs: versatile trans-acting molecules of ancient evolutionary origin. Gene Expr 10:17–39PubMedGoogle Scholar
  4. 4.
    Gagnon KT, Zhang X, Agris PF, Maxwell ES (2006) Assembly of the archaeal box C/D sRNP can occur via alternative pathways and requires temperature-facilitated sRNA remodeling. J Mol Biol 362(5):1025–1042PubMedCrossRefGoogle Scholar
  5. 5.
    Gagnon KT, Maxwell ES (2011) Electrophoretic mobility shift assay for characterizing rna-protein interaction. In: Nielsen H (ed) Methods in molecular biology, vol 703. Humana, New York, pp 275–291Google Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Department of Biochemistry and Molecular Biology, School of Medicine, Department of Chemistry and BiochemistrySouthern Illinois UniversityCarbondaleUSA
  2. 2.Department of Molecular and Structural BiochemistryNorth Carolina State UniversityRaleighUSA

Personalised recommendations