Identification of target mRNAs of regulatory RNA-binding proteins using mRNP immunopurification and microarrays

Nature Protocols

Volumn 2, Issue 8, 2007, Pages 2033-2042

  Sanchez, Mayka ^a,b,c   Galy, Bruno ^b   Hentze, Matthias W ^a,b   Muckenthaler, Martina U ^a,c  

^a Molecular Medicine Partnership Unit   (Germany)

^b EUROPEAN MOLECULAR BIOLOGY LABORATORY   (Germany)

^c UNIVERSITY OF HEIDELBERG   (Germany)

Author keywords

[No Author keywords available]

Indexed keywords

MESSENGER RNA; PROTEIN ANTIBODY; RECOMBINANT PROTEIN; REGULATOR PROTEIN; RIBONUCLEOPROTEIN; RNA BINDING PROTEIN;

ANIMAL CELL; ANIMAL TISSUE; ARTICLE; CONTROLLED STUDY; GENE IDENTIFICATION; GENE TARGETING; IMMUNOPRECIPITATION; METHODOLOGY; MICROARRAY ANALYSIS; NONHUMAN; PRIORITY JOURNAL; PROTEIN MICROARRAY; PROTEIN PURIFICATION; REGULATORY MECHANISM; RNA ANALYSIS; SUPERNATANT; TIME;

DNA, COMPLEMENTARY; DNA-DIRECTED RNA POLYMERASES; IMMUNOPRECIPITATION; OLIGONUCLEOTIDE ARRAY SEQUENCE ANALYSIS; RIBONUCLEOPROTEINS; RNA PROCESSING, POST-TRANSCRIPTIONAL; RNA, MESSENGER; RNA-BINDING PROTEINS; TRANSCRIPTION, GENETIC; VIRAL PROTEINS;

EID: 34548148522     PISSN: 17542189     EISSN: 17502799     Source Type: Journal    
DOI: 10.1038/nprot.2007.293     Document Type: Article

References (17)

1. Moore, M.J. From birth to death: The complex lives of eukaryotic mRNAs. Science 309, 1514-1518 (2005).
2. Sanchez, M., Galy, B., Muckenthaler, M.U. & Hentze, M.W. Iron-regulatory proteins limit hypoxia-inducible factor 2alpha expression in iron deficiency. Nat. Struct. Mol. Biol. 14, 420-426 (2007).
3. Sanchez, M. et al. Iron regulation and the cell cycle: identification of an iron-responsive element in the 3′-untranslated region of human cell division cycle 14A mRNA by a refined microarray-based screening strategy. J. Biol. Chem. 281, 22865-22874 (2006).
4. Muckenthaler, M. et al. Relationships and distinctions in iron-regulatory networks responding to interrelated signals. Blood 101, 3690-3698 (2003).
5. Peritz, T. et al. Immunoprecipitation of mRNA-protein complexes. Nat. Protoc. 1, 577-580 (2006).
6. Tenenbaum, S.A., Lager, P.J., Carson, C.C. & Keene, J.D. Ribonomics: identifying mRNA subsets in mRNP complexes using antibodies to RNA-binding proteins and genomic arrays. Methods 26, 191-198 (2002).
7. Keene, J.D., Komisarow, J.M. & Friedersdorf, M.B. RIP-Chip: The isolation and identification of mRNAs, microRNAs and protein components of ribonucleoprotein complexes from cell extracts. Nat. Protoc. 1, 302-307 (2006).
8. Mili, S. & Steitz, J.A. Evidence for reassociation of RNA-binding proteins after cell lysis: Implications for the interpretation of immunoprecipitation analyses. RNA 10, 1692-1694 (2004).
9. Richter, A. et al. Comparison of fluorescent tag DNA Labeling methods used for expression analysis by DNA microarrays. Biotechniques 33, 620-8 630 (2002).
10. Chomczynski, P. & Sacchi, N. The single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction: twenty-something years on. Nat. Protoc. 1, 581-585 (2006).
11. Gray, N. K. et al. Recombinant iron-regulatory factor functions as an iron-responsive-element-binding protein, a translational repressor and an aconitase. A functional assay for translational repression and direct demonstration of the iron switch. Eur. J. Biochem. 218 657-667 (1993).
12. Galy, B., Ferring, D., Benesova, M., Benes, V. & Hentze, M.W. Targeted mutagenesis of the murine IRP1 and IRP2 genes reveals context-dependent RNA processing differences in vivo. RNA 10, 1019-1025 (2004).
13. Gunshin, H. et al. Iron-dependent regulation of the divalent metal ion transporter. FEBS Lett. 509, 309-316 (2001).
14. Hentze, M.W., Rouault, T.A., Harford, J.B. & Klausner, R.D. Oxidation-reduction and the molecular mechanism of a regulatory RNA-protein interaction. Science 244, 357-359 (1989).
15. Hentze, M.W., Muckenthaler, M.U. & Andrews, N.C. Balancing acts: molecular control of mammalian iron metabolism. Cell 117, 285-297 (2004).
16. Pesole, G. & Liuni, S. Internet resources for the functional analysis of 5′ and 3′ untranslated regions of eukaryotic mRNAs. Trends Genet. 15, 378 (1999).
17. Bengert, P. & Dandekar, T. A software tool-box for analysis of regulatory RNA elements. Nucleic Acids Res. 31, 3441-3445 (2003).