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Sci. STKE, 26 February 2002
Vol. 2002, Issue 121, p. tw84
[DOI: 10.1126/stke.2002.121.tw84]


Ubiquitin Ligases Inhibited and Localized by a Pseudosubstrate

The nuclear factor {kappa}B (NF-{kappa}B) pathway is activated upon the phosphorylation-dependent ubiquitination and degradation of the inhibitor of {kappa}B (I{kappa}B). The E3 ligase primarly responsible for this ubiquitination is a member of the Skp1, Cul1, F-box protein (SCF)-type E3s, called SCFβ-TrCP. The F-box subunit of this complex is E3RS, which is localized in the nucleus, whereas I{kappa}B is cytoplasmic. Davis et al. determined that nuclear localization of E3RS was due to interactions with the nuclear protein heterogenous nuclear ribonucleoprotein U (hnRNP U). These interactions occurred through the same WD domain through which E3RS interacted with its substrate I{kappa}B. Indeed, in vitro these interactions were competitive. The entire functional SCFβ-TrCP was capable of interacting with hnRNP U through binding to the F-box subunit; and despite the ability of SCFβ-TrCP to ubiquitinate I{kappa}B, hnRNP U was not ubiquitinated under any conditions. Thus, hnRNP U appears to act as a low-affinity pseudosubstrate that can be displaced by the higher-affinity substrate I{kappa}B. Investigations into the nuclear localization due to this interaction between E3RS and hnRNP U determined that the nuclear localization signal of hnRNP U was essential and that disruption of hnRNP U nuclear localization also disrupted E3RS localization. However, forcing E3RS to the cytoplasm by the attachment of a nuclear export signal did not cause the redistribution of hnRNP U. Mutations that disrupted the hnRNP U and E3RS interaction led to decreased stability of the E3RS complex regardless of whether the proteins were cytoplasmic or nuclear. Thus, the hnRNP U interaction serves three functions: (i) it sequesters the F-box substrate recognition subunit of SCFβ-TrCP in the nucleus, (ii) it inhibits the E3 ligase activity of the complex by acting as a pseudosubstrate, and (iii) it stabilizes the E3RS subunit. How E3RS shuttles in and out of the nucleus to serve as the substrate recognition subunit for SCFβ-TrCP and what processes regulate its movement remain unknown.

M. Davis, A. Hatzubai, J. S. Andersen, E. Ben-Shushan, G. Z. Fisher, A. Yaron, A.Bauskin, F. Mercurio, M. Mann, T. Ben-Neriah, Pseudosubstrate regulation of the SCFβ-TrCP ubiquitin ligase by hnRNP-U. Genes Dev. 16, 439-451 (2002). [Abstract] [Full Text]

Citation: Inhibited and Localized by a Pseudosubstrate. Sci. STKE 2002, tw84 (2002).

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