Regulation of Cullin-RING ubiquitin ligase 1 by Spliceosome-associated protein 130 (SAP130)

Lucia Cordero-Espinoza,Thilo Hagen

doi:10.1242/bio.20134374

Abstract

SummaryCullin-RING ubiquitin ligases (CRLs) mediate the ubiquitination of numerous protein substrates and target them for proteasomal degradation. The function of CRLs is under tight regulation by Cullin-binding proteins. It has been reported that the Spliceosome-associated protein 130 (SAP130/SF3b-3) binds to several Cullin proteins, yet it remains unknown whether SAP130 plays any role in regulating the function of CRLs. Here, we report that SAP130 overexpression reduces the binding of adaptor protein Skp1 and substrate receptor Skp2 to Cul1, whereas it has no effect on CAND1 binding to Cul1. Overexpression of SAP130 decreases the degradation rate of p27, a protein substrate of the SCFSkp2 ligase. Interestingly, silencing of SAP130 also inhibits the degradation of p27, suggesting a dual role for SAP130 in the regulation of SCF activity. We hypothesized that the regulatory role of SAP130 could extend to other CRLs; however, overexpression of SAP130 is unable to affect the protein stability of the Cul2 and Cul3 substrates, HIF-1 and NRF-2. SAP130 binds to Cul1, Cul2 and Cul4 with similar affinity, and it binds to Cul3 more strongly. SAP130 localizes in both the nucleus and the cytoplasm. Hence, the inability of SAP130 to regulate Cul2 and Cul3 CRLs cannot be explained by low binding affinity of SAP130 to these cullins or by subcellular sequestration of SAP130. We propose a novel role for SAP130 in the regulation of SCF, whereby SAP130 physically competes with the adaptor protein/F-box protein for Cul1 binding and interferes with the assembly of a functional SCF ligase.

Highlights

Protein degradation is critical for the regulation and maintenance of cellular functions such as cell signaling, transcriptional regulation and cell cycle progression (Petroski and Deshaies, 2005)
It has been reported that the Spliceosome-associated protein 130 (SAP130/SF3b-3) binds to several Cullin proteins, yet it remains unknown whether Spliceosome associated protein 130 (SAP130) plays any role in regulating the function of Cullin-RING ubiquitin ligases (CRLs)
We report that SAP130 overexpression reduces the binding of adaptor protein Skp1 and substrate receptor Skp2 to Cul1, whereas it has no effect on CAND1 binding to Cul1

Summary

Introduction

Protein degradation is critical for the regulation and maintenance of cellular functions such as cell signaling, transcriptional regulation and cell cycle progression (Petroski and Deshaies, 2005). The majority of proteins in the cell are degraded via the ubiquitin-mediated proteolytic pathway (Ciechanover et al, 1984). The E3 ligase plays a pivotal role as it facilitates the transfer of ubiquitin onto a determined protein substrate, providing the substrate specificity of the reaction (Pickart and Cohen, 2004). The substrate recognition module of the CRL assembles at the N-terminus of the cullin protein (Wu et al, 2000). It encompasses a substrate receptor and, with the exception of Cul3-based CRLs, an adaptor protein that links the substrate receptor to the complex (Petroski and Deshaies, 2005)

Methods

Results

Discussion

Conclusion