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The cytoplasmic domains of complement regulatory protein CD46 interact with multiple kinases in macrophages
1Department of Microbiology, University of Washington School of Medicine, Seattle 98195, USA.
Abstract:
Membrane cofactor protein (CD46), which normally protects autologous cells from complement lysis, is the human cell receptor for measles virus (MV). Interaction between MV and CD46 on monocytes can lead to suppression of monocyte activation. We have investigated the interaction between the cytoplasmic sequences of CD46 and kinases in a mouse macrophage cell line. Glutathione-S-transferase (GST) fusion proteins bearing the Cyt1 or Cyt2 alternative cytoplasmic domain of CD46 associate with macrophage kinase activity, which phosphorylates multiple proteins co-purified with the GST fusion proteins. Association with the macrophage kinase activity correlates with tyrosine phosphorylation of the CD46 cytoplasmic domains. Removing the CD46 sequences or introducing a frame-shift mutation abrogates the association with macrophage kinase activity. Renaturation studies reveal multiple kinases with apparent molecular mass of 82, 79, 58, and 50/49 kDa, which associate specifically with both CD46 cytoplasmic domains. Alanine substitutions at a juxtamembrane Tyr-X-X-Leu motif in the Cyt1 domain completely abrogate the association with macrophage kinases and tyrosine phosphorylation of Cyt1; but similar substitutions in the Cyt2 domain only partially reduce the association with kinases and tyrosine phosphorylation of Cyt2. These results reveal a specific interaction between complement regulatory protein CD46 and macrophage kinases. These findings may provide an important clue for understanding immune modulation by MV.
Insights
Membrane cofactor protein (CD46) interacts with macrophage kinases, influencing immune responses. This interaction, crucial for measles virus (MV) entry, may explain MV
Area of Science:
- Immunology
- Virology
- Cell Biology
Background:
- Membrane cofactor protein (CD46) is a complement regulatory protein protecting host cells.
- CD46 serves as the human cell receptor for measles virus (MV).
- MV-CD46 interaction can suppress monocyte activation, indicating immune modulation.
Purpose of the Study:
- To investigate the interaction between CD46 cytoplasmic domains and kinases in macrophages.
- To elucidate the role of CD46 cytoplasmic sequences in kinase association and phosphorylation.
Main Methods:
- Utilized glutathione-S-transferase (GST) fusion proteins of CD46 cytoplasmic domains (Cyt1, Cyt2).
- Co-purification and renaturation studies to identify associated kinases in a mouse macrophage cell line.
- Site-directed mutagenesis (alanine substitutions) to analyze kinase interaction motifs.
Main Results:
- CD46 cytoplasmic domains (Cyt1 and Cyt2) associate with macrophage kinase activity.
- This association leads to tyrosine phosphorylation of CD46 cytoplasmic domains.
- Specific kinases (82, 79, 58, 50/49 kDa) were identified.
- A juxtamembrane motif in Cyt1 is critical for kinase association, while Cyt2's interaction is partially sensitive to mutations.
Conclusions:
- Reveals a specific interaction between CD46 and macrophage kinases.
- Suggests a mechanism for immune modulation by measles virus via CD46 signaling.
- Provides insights into CD46's role in host-pathogen interactions and immune regulation.