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Methods. 2019 Apr 15;159-160:157-164. doi: 10.1016/j.ymeth.2019.02.015. Epub 2019 Feb 19.

Imaging-based assays for investigating functions of the RNA polymerase II elongation factor Elongin and the Elongin ubiquitin ligase.

Author information

1
Stowers Institute for Medical Research, 1000 E 50th Street, Kansas City, MO 64110, United States.
2
Stowers Institute for Medical Research, 1000 E 50th Street, Kansas City, MO 64110, United States; Department of Biochemistry & Molecular Biology, University of Kansas Medical Center, Kansas City, KS 66160, United States.
3
Stowers Institute for Medical Research, 1000 E 50th Street, Kansas City, MO 64110, United States; Department of Biochemistry & Molecular Biology, University of Kansas Medical Center, Kansas City, KS 66160, United States. Electronic address: jlc@stowers.org.

Abstract

Elongin A binds to Elongins B and C to form the RNA polymerase II transcription elongation factor Elongin. It also functions as the substrate recognition subunit of a ubiquitin ligase that is formed by binding of Elongin to Cullin protein CUL5 and RING finger protein RBX2 and that targets RNA polymerase II for ubiquitination. In this article, we describe use of acceptor photobleaching fluorescence resonance energy transfer (AP-FRET) and laser microirradiation-based assays to study regulated assembly of the Elongin ubiquitin ligase and its recruitment to regions of localized DNA damage.

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