Orientation-specific signalling by thrombopoietin receptor dimers

Area of Science:

• Biochemistry
• Molecular Biology
• Hematology

Background:

• Thrombopoietin receptor (TPO-R) activation is crucial for megakaryocyte differentiation, platelet formation, and hematopoietic stem cell renewal.
• Ligand binding stabilizes an active TPO-R dimer, but the structural basis for distinct signaling outcomes remains incompletely understood.

Purpose of the Study:

• To investigate how different orientations of TPO-R dimers influence its biological activity and in vivo phenotypes.
• To identify specific dimeric interfaces that mediate distinct signaling pathways and cellular responses.

Main Methods:

• Engineered TPO-R constructs by fusing dimeric coiled coils in all seven possible orientations to the transmembrane (TM)-cytoplasmic domains.
• Utilized cysteine mutagenesis and crosslinking to visualize and confirm specific dimeric orientations.
• Employed functional assays and computational searches to analyze receptor activity and signaling states.

Main Results:

• Identified distinct biological effects and in vivo phenotypes corresponding to different TPO-R dimeric orientations.
• Characterized one orientation representing an inactive dimeric state and another mimicking physiological activation.
• Discovered several other dimeric orientations that induce proliferation and lead to myeloproliferative and myelodysplastic disorders in vivo.

Conclusions:

• The TPO-R can signal through multiple distinct dimeric interfaces, not just a single active conformation.
• Subtle variations in cytokine receptor dimerization represent a novel layer of signaling regulation with implications for disease.
• These findings provide new insights into the activation of signaling pathways by a single receptor and TPO-R-related pathologies.