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Proc Natl Acad Sci U S A. 2016 Jul 19;113(29):E4238-47. doi: 10.1073/pnas.1603229113. Epub 2016 Jun 29.

Mapping transcription factor interactome networks using HaloTag protein arrays.

Author information

1
Genomic Analysis Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037; Plant Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037; RIKEN Center for Integrative Medical Sciences, Yokohama City, Kanagawa 230-0045, Japan;
2
Genomic Analysis Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037;
3
Plant Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037;
4
Cell and Developmental Biology Section, Division of Biological Sciences, University of California, San Diego, La Jolla, CA 92093;
5
Center for Cancer Systems Biology (CCSB), Dana-Farber Cancer Institute, Boston, MA 02215; Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215; Department of Genetics, Harvard Medical School, Boston, MA 02115;
6
Fondo de Financiamiento de Centros de Investigación en Áreas Prioritarias (FONDAP) Center for Genome Regulation, Millennium Nucleus Center for Plant Systems and Synthetic Biology, Departamento de Genética Molecular y Microbiología, Pontificia Universidad Católica de Chile, 340 Santiago, Chile;
7
Life Technologies, Carlsbad, CA 92008;
8
Plant Systems Biology, School of Life Sciences Weihenstephan (WZW), Technische Universität München (TUM), 85354 Freising, Germany;
9
Plant Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037; Howard Hughes Medical Institute, The Salk Institute for Biological Studies, La Jolla, CA 92037;
10
Personalized Diagnostics, The Biodesign Institute, Arizona State University, Tempe, AZ 85287;
11
Plant Systems Biology, School of Life Sciences Weihenstephan (WZW), Technische Universität München (TUM), 85354 Freising, Germany; Institute for Advanced Study, Technische Universität München (TUM), 85748 Garching, Germany pbraun@wzw.tum.de ecker@salk.edu.
12
Genomic Analysis Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037; Plant Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037; Howard Hughes Medical Institute, The Salk Institute for Biological Studies, La Jolla, CA 92037; pbraun@wzw.tum.de ecker@salk.edu.

Abstract

Protein microarrays enable investigation of diverse biochemical properties for thousands of proteins in a single experiment, an unparalleled capacity. Using a high-density system called HaloTag nucleic acid programmable protein array (HaloTag-NAPPA), we created high-density protein arrays comprising 12,000 Arabidopsis ORFs. We used these arrays to query protein-protein interactions for a set of 38 transcription factors and transcriptional regulators (TFs) that function in diverse plant hormone regulatory pathways. The resulting transcription factor interactome network, TF-NAPPA, contains thousands of novel interactions. Validation in a benchmarked in vitro pull-down assay revealed that a random subset of TF-NAPPA validated at the same rate of 64% as a positive reference set of literature-curated interactions. Moreover, using a bimolecular fluorescence complementation (BiFC) assay, we confirmed in planta several interactions of biological interest and determined the interaction localizations for seven pairs. The application of HaloTag-NAPPA technology to plant hormone signaling pathways allowed the identification of many novel transcription factor-protein interactions and led to the development of a proteome-wide plant hormone TF interactome network.

KEYWORDS:

Arabidopsis thaliana; hormone; interactome; protein arrays; systems biology

PMID:
27357687
PMCID:
PMC4961138
DOI:
10.1073/pnas.1603229113
[Indexed for MEDLINE]
Free PMC Article

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