The TRIM4 E3 ubiquitin ligase degrades TPL2 and is modulated by oncogenic KRAS

Area of Science:

• Molecular Biology
• Cancer Biology
• Biochemistry

Background:

• Loss-of-function mutations in TPL2 kinase C-terminus promote oncogenesis by preventing proteasomal degradation.
• The precise degradation mechanism of TPL2 has not been fully elucidated.
• Understanding TPL2 regulation is crucial for developing targeted cancer therapies.

Purpose of the Study:

• To identify the E3 ligase responsible for TPL2 degradation.
• To investigate the role of KRAS mutations in TPL2 stability and oncogenic signaling.
• To elucidate the regulatory pathway involving TPL2, TRIM4, TRIM21, and KRAS.

Main Methods:

• Proximity-dependent biotin identification (BioID) to identify interacting proteins.
• Site-directed mutagenesis to study specific lysine residues (K415, K439) in TPL2.
• Western blotting and immunoprecipitation to assess protein stability and interactions.
• Analysis of signaling pathways including Wnt and GSK3β.

Main Results:

• Tripartite motif-containing 4 (TRIM4) was identified as the E3 ligase that polyubiquitinates and degrades TPL2 at lysines 415 and 439.
• Naturally occurring TPL2 mutants (R442H, E188K) showed impaired TRIM4 binding, leading to increased stability.
• TRIM4 stability is regulated by TRIM21, which is further controlled by KRAS.
• Mutant KRAS promotes RNF185-mediated degradation of TRIM21 and TRIM4, stabilizing TPL2.
• Mutant KRAS-induced TPL2 phosphorylates and degrades GSK3β, stabilizing β-catenin and activating the Wnt pathway.

Conclusions:

• TRIM4 is the E3 ligase targeting TPL2 for degradation, with specific lysine residues mediating this process.
• Mutant KRAS disrupts TPL2 degradation via the TRIM4/TRIM21 axis, leading to oncogenic Wnt pathway activation.
• Targeting TPL2 may represent a therapeutic strategy for KRAS-mutant cancers.