Photoaffinity-engineered protein scaffold for systematically exploring native phosphotyrosine signaling complexes in...

Area of Science:

• Biochemistry
• Proteomics
• Cancer Signaling

Background:

• Phosphotyrosine (pTyr)-regulated protein complexes are crucial in cancer signaling.
• Characterizing these transient complexes in tumors is challenging due to limited access and interaction lability.

Purpose of the Study:

• To develop a novel chemical proteomics approach for profiling dynamic pTyr protein complexes in clinical samples.
• To overcome the limitations of traditional methods in detecting pTyr signaling.

Main Methods:

• Engineered Src homology 2 (SH2) domains with chemical probes and genetic mutations.
• Utilized UV light-induced cross-linking, biotin capture, and mass spectrometry.
• Applied the Photo-pTyr-scaffold technique to breast cancer tissue samples.

Main Results:

• Successfully profiled native pTyr protein complexes from breast cancer tissues at a proteome scale.
• Achieved approximately 100-fold higher sensitivity for pTyr signaling proteins compared to immunohistochemistry.
• Identified over 1,000 pTyr proteins, including PDGFRB as a key intercellular signaling regulator.

Conclusions:

• The Photo-pTyr-scaffold approach provides high selectivity and sensitivity for profiling dynamic pTyr signaling.
• Validated PDGFRB as a therapeutic target, demonstrating that its blockade suppresses tumor growth.
• This method offers a versatile tool for analyzing pTyr signaling complexes in clinical settings.