Journal of Biological Chemistry
Volume 286, Issue 33, 19 August 2011, Pages 28844-28857
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Gene Regulation
Hierarchical Phosphorylation within the Ankyrin Repeat Domain Defines a Phosphoregulatory Loop That Regulates Notch Transcriptional Activity*

https://doi.org/10.1074/jbc.M111.243600Get rights and content
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The Notch signal transduction pathway mediates important cellular functions through direct cell-to-cell contact. Deregulation of Notch activity can lead to an altered cell proliferation and has been linked to many human cancers. Casein kinase 2 (CK2), a ubiquitous kinase, regulates several cellular processes by phosphorylating proteins involved in signal transduction, gene expression, and protein synthesis. In this report we identify NotchICD as a novel target of phosphorylation by CK2. Using mapping and mutational studies, we identified serine 1901, located in the ankyrin domain of Notch, as the target amino acid. Interestingly, phosphorylation of serine 1901 by CK2 appears to generate a second phosphorylation site at threonine 1898. Furthermore, threonine 1898 phosphorylation only occurs when Notch forms a complex with Mastermind and CSL. Phosphorylation of both threonine 1898 and serine 1901 resulted in decreased binding of the Notch-Mastermind-CSL ternary complex to DNA and consequently lower transcriptional activity. These data indicate that the phosphorylation of serine 1901 and threonine 1898 negatively regulates Notch function by dissociating the complex from DNA. This study identifies a new component involved in regulation of NotchICD transcriptional activity, reinforcing the notion that a precise and tight regulation is required for this essential signaling pathway.

Notch Pathway
Protein-DNA Interaction
Protein Kinases
Protein Phosphorylation
Signal Transduction
Transcription Regulation
Hierarchical Phosphorylation
Transcriptional Activation Complex

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*

This work was supported, in whole or in part, by National Institutes of Health Grants ROI CA 83736 and 125044-04 (NCI; to A. J. C.). This work was also supported by the Samuel Waxman Foundation for Cancer Research (to A. J. C.).

1

These authors contributed equally to this work.

2

Present address: Dept. of Cancer Biology, 233 S. 10th St., Philadelphia, PA 19107.