The HARPER Lab

Department of Cell Biology

Targets and Mechanisms of CRLs

SCF targets and regulatory kinases we have worked on over the years.
Structures of phosphodegrons bound to 2 F-box proteins (collaboration with Nikola Pavletich)
Systematic analysis of substrates for the FBXL family of CRLs

The Uniquitin-Proteasome Pathway (UPP) controls the abundance and activity of a large number of proteins in the cell. Through the work of this and other labs, the Cullin-RING ubiquitin ligase system has emerged as a major system controlling the stability and regulated proteolysis of hundreds of cellular proteins in key signaling and disease systems. CRLs are composed of an E3 ligase module containing a neddylated cullin (CUL1-7) and a RING finger protein (RBX1 or 2) and a substrate adaptor module which interacts with substrate. For example, CUL1’s adaptor module is composed of SKP1 and a member of the F-box family of proteins, of which 68 exist in mammals. Thus, this super-family of E3s which is referred to as the SCF (SKP1-CUL1-F-box protein) encodes a large variety of E3s involved in a plethora of signaling pathways, ranging from Wnt, NFkB, Hedgehog, and numerous circuits that control cell division. Our research efforts on the CRL system, including: 1) identification of new substrates for CRLs, with a focus on CUL1 and CUL4-based CRLs linked with disease, 2) architecture of the CRL system in mammalian cells analyzed using quantitative protoemics, 3) mechanisms of SCF regulation.

Recently, we have developed methods for systematic identification of CRL substrates [referred to as PAC (Parallel Adaptor Capture) Proteomics] and have applied this approach to a class of F-box proteins - The leucine rich repeat FBXL class. This led to the identification of numerous candidte substrates for each of the 20 FBXL proteins analyzed. We examined a candidate substrate of FBXL17 in detail. This substrate - BACH1 - is a transcriptional repressor of NRF2 dependent genes. We found that FBXL17 promotes turnover of BACH2 in order to "license" NRF2 promoters for activation by NRF2. The use of this approach has the potential to identify substrates broadly across all CRL type ubiquitin ligases. See: Tan et al, Molecular Cell, 2013.