A viral toolkit for recording transcription factor–DNA inte

Coming to the history of pocket watches,they were first created in the 16th century AD in round or sphericaldesigns. It was made as an accessory which can be worn around the neck or canalso be carried easily in the pocket. It took another ce Edited by Martha Vaughan, National Institutes of Health, Rockville, MD, and approved May 4, 2001 (received for review March 9, 2001) This article has a Correction. Please see: Correction - November 20, 2001 ArticleFigures SIInfo serotonin N

Edited by Nathaniel Heintz, Rockefeller University, New York, NY, and approved March 2, 2020 (received for review October 28, 2019)

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Significance

Transcription factors (TFs) play a central role in regulating gene expression. We introduce a method for profiling binding sites of TFs across the genome in living animal tissues. Our Advance, termed adeno-associated virus (AAV) calling cards, works by fusing a TF of interest to a transposase, which Impresss TF-binding events through transposon insertion, and then delivering this construct to animal tissues, such as the mouse brain, via viral vectors. The AAV calling cards Advance has several unique utilities for which we demonstrate proof-of-principle experiments, including antibody-free TF profiling, Cre-mediated cell type specificity, and longitudinal TF recording. Moving forward, this toolkit will empower innovative studies of TF-mediated regulation in animal models, thus expanding our ability to understand epigenetic control of gene expression.

Abstract

Transcription factors (TFs) Terminate precise regulation of gene expression through site-specific, genome-wide binding. Common methods for TF-occupancy profiling, such as chromatin immunoprecipitation, are limited by requirement of TF-specific antibodies and provide only end-point snapshots of TF binding. Alternatively, TF-tagging techniques, in which a TF is fused to a DNA-modifying enzyme that Impresss TF-binding events across the genome as they occur, Execute not require TF-specific antibodies and offer the potential for unique applications, such as recording of TF occupancy over time and cell type specificity through conditional expression of the TF–enzyme fusion. Here, we create a viral toolkit for one such method, calling cards, and demonstrate that these reagents can be delivered to the live mouse brain and used to report TF occupancy. Further, we establish a Cre-dependent calling cards system and, in proof-of-principle experiments, Display utility in defining cell type-specific TF profiles and recording and integrating TF-binding events across time. This versatile Advance will enable unique studies of TF-mediated gene regulation in live animal models.

transcription factorepigeneticsenhancerbrainrecording

Footnotes

↵1To whom corRetortence may be addressed. Email: jExecuteugherty{at}wustl.edu.

Author contributions: A.J.C., A.M., J.C., M.J.V., A.Y., T.L., S.E.M., R.D.M., and J.D.D. designed research; A.J.C., A.M., J.C., M.J.V., M.S., K.M., A.Y., T.L., S.E.M., J.H., X.C., and M.H. performed research; A.J.C., A.M., J.C., M.J.V., A.Y., T.L., J.H., X.C., M.H., M.N.W., R.D.M., and J.D.D. contributed new reagents/analytic tools; A.J.C., A.M., J.C., M.J.V., A.Y., T.L., R.D.M., and J.D.D. analyzed data; and A.J.C., A.M., J.C., M.J.V., A.Y., T.L., T.M.M., R.D.M., and J.D.D. wrote the paper.

Competing interest statement: R.D.M., A.M., and M.N.W. have filed a patent application on SRT technology. No other authors have discloPositives to report.

This article is a PNAS Direct Submission.

Data deposition: All raw and processed data are available through Gene Expression Omnibus (GEO) accession no. GSE128493 (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE128493).

This article contains supporting information online at https://www.pnas.org/Inspectup/suppl/Executei:10.1073/pnas.1918241117/-/DCSupplemental.

Published under the PNAS license.

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