pSILAC method coupled with two complementary digestion approaches reveals PRPF39 as a new E7070-dependent DCAF15...

Area of Science:

• Biochemistry
• Molecular Biology
• Cancer Research

Background:

• Targeting specific ubiquitin E3 ligases for protein degradation is a key strategy in cancer therapy.
• The drug E7070 targets DCAF15 E3 ligase, with RBM39 as its only known substrate, and exhibits anticancer activity.
• Elucidating additional substrates of E7070 is crucial for understanding its mechanism of action.

Purpose of the Study:

• To identify novel substrates of the drug E7070.
• To investigate the role of E7070-mediated protein degradation in cancer therapy.
• To explore the potential of PRPF39 as a therapeutic target.

Main Methods:

• Utilized pulsed SILAC (pSILAC) to monitor protein turnover accurately.
• Employed two complementary digestion approaches (LysC-Trypsin and LysN-LysArgiNase) to enhance proteome profiling depth.
• Conducted biological assays to confirm substrate ubiquitination and degradation.

Main Results:

• E7070 treatment altered the turnover rates of 868 proteins.
• Identified pre-mRNA splicing factor 39 (PRPF39) as a potential new substrate due to accelerated turnover.
• Demonstrated that E7070 induces ubiquitination and DCAF15-dependent degradation of PRPF39.

Conclusions:

• PRPF39 is a novel bona fide substrate of E7070, degraded in a DCAF15-dependent manner.
• The degradation of PRPF39 by E7070 may contribute to its anticancer activity.
• This study expands the understanding of E7070's mechanism of action and identifies a new therapeutic target.