The Notch Intracellular Domain Is Ubiquitinated and Negatively Regulated by the Mammalian Sel-10 Homolog^*

Camilla Öberg, Jinhe Li^§, Adele Pauley^§, Elisabeth Wolf, Mark Gurney^§¶, and Urban Lendahl

From the  Department of Cell and Molecular Biology, Medical Nobel Institute, Karolinska Institute, SE-171 77 Stockholm, Sweden and the ^§ Department of Neurobiology, Pharmacia Corporation, Kalamazoo, Michigan 49001

The Caenorhabditis elegans sel-10 protein is structurally similar to E3 ubiquitin ligases and is a negative regulator of Notch (lin-12) and presenilin signaling. In this report, we characterize the mammalian Sel-10 homolog (mSel-10) and analyze its effects on Notch signaling. We find that mSel-10 localizes to the cell nucleus, and that it physically interacts with the Notch 1 intracellular domain (IC) and reduces Notch 1 IC-mediated activation of the HES 1 promoter. Notch 1 IC is ubiquitinated by mSel-10, and ubiquitination requires the presence of the most carboxyl-terminal region of the Notch IC, including the PEST domain. In the presence of the proteasome inhibitor MG132, the amount of Notch 1 IC and its level of ubiquitination are increased. Interestingly, this accumulation of Notch 1 IC in the presence of MG132 is accompanied by decreased activation of the HES 1 promoter, suggesting that ubiquitinated Notch 1 IC is a less potent transactivator. Finally, we show that mSel-10 itself is ubiquitinated and degraded by the proteasome. In conclusion, these data reveal the importance of ubiquitination and proteasome-mediated degradation for the activity and turnover of Notch ICs, and demonstrate that mSel-10 plays a key role in this process.

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^* This work was supported by grants from Cancerfonden, EU Project QLRT-1999-30934 (to U. L.), and the Foundation for Strategic Research (to C. Ö.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

^¶ Present address: deCode Genetics, 110 Reykjavik, Iceland.

To whom correspondence should be addressed. Tel.: 46-8-7287323; Fax: 46-8-348135.

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Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.

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