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Analyzing the Interaction of RBPJ with Mitotic Chromatin and Its Impact on Transcription Reactivation upon Mitotic Exit

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Notch Signaling Research

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2472))

Abstract

The sequence-specific transcription factor RBPJ, also known as CSL (CBF1, Su(H), Lag1), is an evolutionarily conserved protein that mediates Notch signaling to guide cell fates. When cells enter mitosis, DNA is condensed and most transcription factors dissociate from chromatin; however, a few, select transcription factors, termed bookmarking factors, remain associated. These mitotic chromatin-bound factors are believed to play important roles in maintaining cell fates through cell division. RBPJ is one such factor that remains mitotic chromatin associated and therefore could function as a bookmarking factor. Here, we describe how to obtain highly purified mitotic cells from the mouse embryonal carcinoma cell line F9, perform chromatin immunoprecipitation with mitotic cells, and measure the first run of RNA synthesis upon mitotic exit. These methods serve as basis to understand the roles of mitotic bookmarking by RBPJ in propagating Notch signals through cell division.

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Acknowledgments

This work was supported by American Heart Association 17GRNT33400020 to H.Y.F., and Cancer Center Support Grant P30CA118100. Support for microscopy imaging was provided by the University of New Mexico Cancer Center Fluorescence Microscopy Shared Resource, funded by NCI 2P30 CA118100 and NIGMS 5P50 GM085273.

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Author notes

  1. Kostiantyn Dreval

    Present address: Department of Molecular Biology and Biochemistry, Simon Fraser University, Burnaby, BC, Canada

Authors and Affiliations

  1. The Program in Cellular and Molecular Oncology, University of New Mexico Comprehensive Cancer Center, Albuquerque, NM, USA

    Kostiantyn Dreval, Robert J. Lake & Hua-Ying Fan

  2. Department of Internal Medicine, Division of Molecular Medicine, University of New Mexico Health Science Center, Albuquerque, NM, USA

    Kostiantyn Dreval, Robert J. Lake & Hua-Ying Fan

Authors
  1. Kostiantyn Dreval
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  2. Robert J. Lake
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  3. Hua-Ying Fan
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Corresponding author

Correspondence to Hua-Ying Fan .

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Editors and Affiliations

  1. Department of Biology, Georgia Southern University, Statesboro, GA, USA

    Dongyu Jia

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Dreval, K., Lake, R.J., Fan, HY. (2022). Analyzing the Interaction of RBPJ with Mitotic Chromatin and Its Impact on Transcription Reactivation upon Mitotic Exit. In: Jia, D. (eds) Notch Signaling Research. Methods in Molecular Biology, vol 2472. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2201-8_9

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  • DOI: https://doi.org/10.1007/978-1-0716-2201-8_9

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2200-1

  • Online ISBN: 978-1-0716-2201-8

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