Investigating pioneer factor activity and its coordination with chromatin remodelers using integrated synthetic oligo assay

Hengye Chen; Chao Yan; Archana Dhasarathy; Michael Kladde; Lu Bai

doi:10.1016/j.xpro.2023.102279

Investigating pioneer factor activity and its coordination with chromatin remodelers using integrated synthetic oligo assay

STAR Protoc. 2023 Jun 7;4(2):102279. doi: 10.1016/j.xpro.2023.102279. Online ahead of print.

Authors

Hengye Chen¹, Chao Yan², Archana Dhasarathy³, Michael Kladde⁴, Lu Bai⁵

Affiliations

¹ Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA; Center for Eukaryotic Gene Regulation, The Pennsylvania State University, University Park, PA 16802, USA. Electronic address: hengyechen@outlook.com.
² Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA; Center for Eukaryotic Gene Regulation, The Pennsylvania State University, University Park, PA 16802, USA.
³ Department of Biomedical Sciences, University of North Dakota School of Medicine and Health Sciences, Grand Forks, ND 58201, USA.
⁴ Department of Biochemistry and Molecular Biology, College of Medicine, University of Florida, Gainesville, FL 32610, USA; UF Health Cancer Center, University of Florida, Gainesville, FL 32610, USA.
⁵ Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA; Center for Eukaryotic Gene Regulation, The Pennsylvania State University, University Park, PA 16802, USA; Department of Physics, The Pennsylvania State University, University Park, PA 16802, USA. Electronic address: lub15@psu.edu.

Abstract

Chromatin accessibility is regulated by pioneer factors (PFs) and chromatin remodelers (CRs). Here, we present a protocol, based on integrated synthetic oligonucleotide libraries in yeast, to systematically interrogate the nucleosome-displacing activities of PFs and their coordination with CRs. We describe steps for designing oligonucleotide sequences, constructing yeast libraries, measuring nucleosome configurations, and data analyses. This approach potentially can be adapted for use in higher eukaryotes to investigate the activities of many types of chromatin-associated factors. For complete details on the use and execution of this protocol, please refer to Yan et al.,¹ and Chen et al.².

Keywords: Biotechnology and bioengineering; Genetics; Genomics.

Abstract

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