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Time-resolved phenotypic profiling maps EGFR and c-Met trafficking signatures
1Institute for Comprehensive Medical Sciences, Tokyo Women's Medical University, 8-1, Kawada-cho, Shinjuku-ku, Tokyo, 162-8666, Japan. tanabe.kenji@twmu.ac.jp.
Abstract:
The epidermal growth factor receptor (EGFR) and hepatocyte growth factor receptor (c-Met/MET) are receptor tyrosine kinases (RTKs) whose signalling depends on endocytic routing, and impaired down-regulation can drive tumour progression and therapy resistance, including MET-driven bypass of EGFR inhibition. However, how routing states couple to signalling and lipid programs, and how this differs between receptors under matched conditions, remains unclear. Using a chemically diverse annotated inhibitor library, we performed time-resolved, pathway-focused high-content imaging in A549 cells to compare co-varying phenotypes across matched EGFR and c-Met assay systems. Late receptor persistence defined operational degradation-inhibitor phenotypes that were heterogeneous and dispersed across a fused phenotypic atlas. Despite this heterogeneity, matched degradation-inhibitor sets showed receptor-selective lipid-trafficking coupling: EGFR persistence was associated with reduced phosphatidylinositol 4,5-bisphosphate-linked readouts and signalling biases, whereas c-Met persistence aligned with increased perinuclear phosphatidylinositol 4-phosphate heterogeneity and altered transferrin-recycling readouts. These results provide a comparative imaging framework for mapping RTK trafficking states and nominate phosphoinositide-linked features as candidate readouts of trafficking rewiring in cancer-relevant contexts.
Insights
Investigating receptor tyrosine kinases (RTKs) like EGFR and c-Met reveals how their trafficking impacts cancer. Different RTK persistence states link to distinct lipid signaling, offering new therapeutic targets.
Area of Science:
- Cell Biology
- Molecular Oncology
- Biochemistry
Background:
- Receptor tyrosine kinases (RTKs), including epidermal growth factor receptor (EGFR) and hepatocyte growth factor receptor (c-Met/MET), are crucial for cell signaling.
- Their signaling relies on endocytic routing, and impaired down-regulation contributes to tumor progression and therapy resistance.
- Understanding how RTK trafficking states influence signaling and lipid programs is vital, especially in cancer contexts.
Purpose of the Study:
- To compare the coupling of signaling and lipid programs to routing states between EGFR and c-Met under matched conditions.
- To investigate how late receptor persistence phenotypes relate to specific lipid trafficking and signaling biases.
- To establish a comparative imaging framework for mapping RTK trafficking states in cancer.
Main Methods:
- Utilized a chemically diverse annotated inhibitor library for targeted drug screening.
- Employed time-resolved, pathway-focused high-content imaging in A549 cells.
- Compared co-varying phenotypes across matched EGFR and c-Met assay systems to analyze receptor persistence and associated cellular responses.
Main Results:
- Late receptor persistence phenotypes were heterogeneous and dispersed across a fused phenotypic atlas.
- EGFR persistence correlated with reduced phosphatidylinositol 4,5-bisphosphate readouts and signaling biases.
- c-Met persistence was associated with increased perinuclear phosphatidylinositol 4-phosphate heterogeneity and altered transferrin-recycling readouts, demonstrating receptor-selective lipid-trafficking coupling.
Conclusions:
- A comparative imaging framework was established for mapping RTK trafficking states.
- Phosphoinositide-linked features are nominated as candidate readouts for trafficking rewiring in cancer.
- These findings provide insights into RTK signaling dynamics and potential therapeutic strategies targeting cancer progression and resistance.
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