Pseudosubstrate regulation of the SCF^β-TrCP ubiquitin ligase by hnRNP-U

Abstract

␤-TrCP/E3RS (E3RS) is the F-box protein that functions as the receptor subunit of the SCF␤-TrCP ubiquitin ligase (E3). Surprisingly, although its two recognized substrates, IB␣ and ␤-catenin, are present in the cytoplasm, we have found that E3RS is located predominantly in the nucleus. Here we report the isolation of the major E3RS-associated protein, hnRNP-U, an abundant nuclear phosphoprotein. This protein occupies E3RS in a specific and stoichiometric manner, stabilizes the E3 component, and is likely responsible for its nuclear localization. hnRNP-U binding was abolished by competition with a pIB␣ peptide, or by a specific point mutation in the E3RS WD region, indicating an E3-substrate-type interaction. However, unlike pIB␣, which is targeted by SCF ␤-TrCP for degradation, the E3-bound hnRNP-U is stable and is, therefore, a pseudosubstrate. Consequently, hnRNP-U engages a highly neddylated active SCF ␤-TrCP , which dissociates in the presence of a high-affinity substrate, resulting in ubiquitination of the latter. Our study points to a novel regulatory mechanism, which secures the localization, stability, substrate binding threshold, and efficacy of a specific protein-ubiquitin ligase.

Structural and Transcriptional Analysis of the Self-Incompatibility Locus of Almond: Identification of a Pollen-Expressed F-Box Gene with Haplotype-Specific Polymorphism

“…SFB might function as a component of SCF SFB and ubiquitinate all of the nonself S-RNases for degradation but interact specifically with the self S-RNase to leave it active, leading to the arrest of self pollen tube growth. Recently, it was reported that hnRNP-U functions as a pseudosubstrate that binds to the corresponding F-box protein but is not destined for degradation by the SCF complex (Davis et al, 2002). Based on this example, it is possible that self S-RNase serves as a pseudosubstrate for Figure 6.…”

Non-canonical NF-κB signaling pathway

“…One possibility is that some critical factors mediating constitutive p100 processing are located in the nucleus. For example, a number of ubiquitin ligases, including SCF βTrCP , are predominantly localized in the nucleus [27][28][29][30][31]. Although SCF βTrCP is dispensable for constitutive processing of p100 [15], the possibility for the involvement of another nuclear ubiquitin ligase(s) in this molecular event cannot be excluded.…”

Interaction of Epstein-Barr Virus Latent Membrane Protein 1 with SCFHOS/β-TrCP E3 Ubiquitin Ligase Regulates Extent of NF-κB Activation

“…Because a similar motif is found within the cytoplasmic C-terminal domain of LMP1-95-9 (Fig. 1C), we chose to focus our studies on interaction of LMP1 with HOS, which, unlike ␤-TrCP1, is primarily localized in the cytoplasm (49,50).…”