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Cellular arrays for large‐scale analysis of transcription factor activity
Author(s) -
Bellis Abigail D.,
PeňalverBernabé Beatriz,
Weiss Michael S.,
Yarrington Michael E.,
Barbolina Maria V.,
Pannier Angela K.,
Jeruss Jacqueline S.,
Broadbelt Linda J.,
Shea Lonnie D.
Publication year - 2011
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.22916
Subject(s) - luciferase , transcription factor , computational biology , biology , gene expression , microbiology and biotechnology , biological pathway , signal transduction , reporter gene , chemistry , gene , transfection , biochemistry
Identifying molecular mechanisms or therapeutic targets is typically based on large‐scale cellular analysis that measures the abundance of mRNA or protein; however, abundance does not necessarily correlate with activity. We report a method for direct large‐scale quantification of active pathways that employs a cellular array with parallel gene delivery of constructs that report pathway activity. The reporter constructs encode luciferase, whose expression is influenced by binding of transcription factors (TFs), which are the downstream targets of signaling pathways. Luciferase levels are quantified by bioluminescence imaging (BLI), which allows for rapid, non‐invasive measurements. Activity profiles by BLI of 32 TFs were robust, consistent, and reproducible, and correlated with standard cell lysis techniques. The array identified five TFs with differential activity during phorbol‐12‐myristate‐13‐acetate (PMA)‐induced differentiation of breast cancer cells. A system for rapid, large‐scale, BLI quantification of pathway activity provides an enabling technology for mechanistic studies of cellular responses and processes. Biotechnol. Bioeng. 2011;108: 395–403. © 2010 Wiley Periodicals, Inc.