Two-hybrid selection assay to identify proteins interacting with polymerase II transcription factors and regulators - PubMed (original) (raw)
. 2001 Feb;30(2):296-8, 300, 302.
doi: 10.2144/01302st02.
Affiliations
- PMID: 11233598
- DOI: 10.2144/01302st02
Free article
Two-hybrid selection assay to identify proteins interacting with polymerase II transcription factors and regulators
M Petrascheck et al. Biotechniques. 2001 Feb.
Free article
Abstract
The RNA polymerase III-based two-hybrid system has been developed to detect interactions between proteins such as RNA polymerase II transcription factors and regulators that cannot be studied by the original RNA polymerase II two-hybrid system. This novel method appears to be most useful for a refined analysis of already known protein-protein interactions. However, the application of this system in library screenings has been impaired by the lack of a suitable assay for the selection of the activated pol III reporter gene in yeast. Here, we describe a novel selection assay for the pol III-based two-hybrid system that makes it readily usable for screening expression libraries to search for interacting partners. Our system utilizes a temperature-sensitive (ts) U6 snRNA, which is synthesized by RNA polymerase III from a mutated SNR6 gene in yeast. In this ts strain, interactions between hybrid proteins activate an artificial pol III reporter construct (UASG-SNR6), which controls expression of wild-type U6 snRNA. This wild-type U6 snRNA can suppress the ts phenotype and allow growth at the nonpermissive temperature of 37 degrees C, thus providing a positive selection system for interacting proteins.
Similar articles
- TFIIA is required for in vitro transcription of mammalian U6 genes by RNA polymerase III.
Waldschmidt R, Seifart KH. Waldschmidt R, et al. J Biol Chem. 1992 Aug 15;267(23):16359-64. J Biol Chem. 1992. PMID: 1644819 - Stimulation of Pol III-dependent 5S rRNA and U6 snRNA gene expression by AP-1 transcription factors.
Ahuja R, Kumar V. Ahuja R, et al. FEBS J. 2017 Jul;284(13):2066-2077. doi: 10.1111/febs.14104. Epub 2017 May 29. FEBS J. 2017. PMID: 28488757 - Regulation of snRNA gene expression by the Drosophila melanogaster small nuclear RNA activating protein complex (DmSNAPc).
Hung KH, Stumph WE. Hung KH, et al. Crit Rev Biochem Mol Biol. 2011 Feb;46(1):11-26. doi: 10.3109/10409238.2010.518136. Epub 2010 Oct 6. Crit Rev Biochem Mol Biol. 2011. PMID: 20925482 Review. - A RNA polymerase III-based two-hybrid system to study RNA polymerase II transcriptional regulators.
Marsolier MC, Prioleau MN, Sentenac A. Marsolier MC, et al. J Mol Biol. 1997 May 2;268(2):243-9. doi: 10.1006/jmbi.1997.0979. J Mol Biol. 1997. PMID: 9159467 - Using the yeast two-hybrid system to identify interacting proteins.
Miller J, Stagljar I. Miller J, et al. Methods Mol Biol. 2004;261:247-62. doi: 10.1385/1-59259-762-9:247. Methods Mol Biol. 2004. PMID: 15064463 Review.
Cited by
- Saccharomyces cerevisiae as a Tool to Investigate Plant Potassium and Sodium Transporters.
Locascio A, Andrés-Colás N, Mulet JM, Yenush L. Locascio A, et al. Int J Mol Sci. 2019 Apr 30;20(9):2133. doi: 10.3390/ijms20092133. Int J Mol Sci. 2019. PMID: 31052176 Free PMC article. Review. - Diversity in genetic in vivo methods for protein-protein interaction studies: from the yeast two-hybrid system to the mammalian split-luciferase system.
Stynen B, Tournu H, Tavernier J, Van Dijck P. Stynen B, et al. Microbiol Mol Biol Rev. 2012 Jun;76(2):331-82. doi: 10.1128/MMBR.05021-11. Microbiol Mol Biol Rev. 2012. PMID: 22688816 Free PMC article. Review. - Yeast two-hybrid, a powerful tool for systems biology.
Brückner A, Polge C, Lentze N, Auerbach D, Schlattner U. Brückner A, et al. Int J Mol Sci. 2009 Jun 18;10(6):2763-2788. doi: 10.3390/ijms10062763. Int J Mol Sci. 2009. PMID: 19582228 Free PMC article. Review. - DNA looping induced by a transcriptional enhancer in vivo.
Petrascheck M, Escher D, Mahmoudi T, Verrijzer CP, Schaffner W, Barberis A. Petrascheck M, et al. Nucleic Acids Res. 2005 Jul 7;33(12):3743-50. doi: 10.1093/nar/gki689. Print 2005. Nucleic Acids Res. 2005. PMID: 16002789 Free PMC article. - Organ polarity in Arabidopsis. NOZZLE physically interacts with members of the YABBY family.
Sieber P, Petrascheck M, Barberis A, Schneitz K. Sieber P, et al. Plant Physiol. 2004 Aug;135(4):2172-85. doi: 10.1104/pp.104.040154. Epub 2004 Aug 6. Plant Physiol. 2004. PMID: 15299139 Free PMC article.
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Molecular Biology Databases