Proteomics reveal cap-dependent translation inhibitors remodel the translation machinery and translatome

Area of Science:

• Molecular Biology
• Proteomics
• Cancer Therapeutics

Background:

• Protein synthesis disruption is a key therapeutic strategy.
• eIF4A-targeting compounds, like zotatifin, are in clinical trials for cancer and COVID-19.
• The full proteomic impact of these inhibitors is not well understood.

Purpose of the Study:

• To investigate the proteomic effects of rocaglates, which target cap-dependent translation initiation.
• To explore the complex cellular mechanisms and translational reprogramming induced by these compounds.

Main Methods:

• Mass spectrometry-based translatome analysis using TMT-pSILAC.
• Analysis of active translation factors using MATRIX (mass spectrometry analysis of active translation factors using ribosome density fractionation and isotopic labeling experiments).

Main Results:

• Rocaglates induce complex translational reprogramming, with numerous upregulated proteins driving novel cytotoxic mechanisms like GEF-H1-mediated RHOA/JNK activation.
• These effects are not fully replicated by eIF4A silencing, suggesting broader translational adaptations.
• Specific translation factors, such as eEF1ε1, are crucial for rocaglate-induced translatome remodeling.

Conclusions:

• The cellular response to rocaglate-based translation inhibitors involves comprehensive remodeling of the translational landscape.
• This study reveals previously unrecognized cytotoxic pathways and highlights the complexity of translational control in response to therapeutic intervention.