Structural basis for the interaction between the Drosophila RTK Sevenless (dROS1) and the GPCR BOSS.

Area of Science:

• Developmental Biology
• Structural Biology
• Molecular Cell Biology

Background:

• Sevenless (dROS1) is a Drosophila receptor tyrosine kinase (RTK) crucial for R7 photoreceptor differentiation.
• dROS1 activation requires binding to the extracellular region (ECR) of the GPCR BOSS.
• The structural basis of dROS1-BOSS interaction was previously unknown.

Purpose of the Study:

• To elucidate the physical interaction between the dROS1 extracellular region and its ligand BOSS.
• To determine the structural basis for dROS1 activation by BOSS.
• To provide mechanistic insights into ROS1 signaling relevant to human oncogenesis.

Main Methods:

• Cryo-electron microscopy (cryo-EM) to determine the structure of the dROS1 ECR.
• Hydrogen-deuterium exchange mass spectrometry (HDX-MS) to map binding epitopes.
• Site-directed mutagenesis and AlphaFold complex predictions to validate interaction models.

Main Results:

• The dROS1 ECR adopts a folded-over conformation stabilized by disulfide-stapled helical hairpins.
• Specific binding epitopes were identified: beta-strands in dROS1's third Fibronectin type III (FNIII) domain and a C-terminal peptide in BOSS' ECR.
• The interaction involves hydrophobic contacts and beta-strand augmentation.

Conclusions:

• The study reveals the structural mechanism of dROS1-BOSS binding, clarifying a critical step in Drosophila development.
• Findings offer mechanistic insights into the human ROS1 oncogene, a homolog of dROS1.
• This work provides a foundation for understanding RTK-GPCR interactions in signaling pathways.