Interaction of C1q with the receptor calreticulin requires a conformational change in C1q
A Steinø1, C S Jørgensen, I Laursen
1Department of Research and Development, Statens Serum Institut, Copenhagen, Denmark.
Insights
Calreticulin binds to C1q when C1q is altered, such as when bound to immune complexes. This suggests calreticulin acts as a cell surface receptor for modified C1q, aiding in clearing immune complexes and apoptotic cells.
Area of Science:
- Immunology
- Molecular Biology
- Biochemistry
Background:
- Calreticulin is a key chaperone protein involved in protein folding and quality control within the endoplasmic reticulum.
- C1q is the initiating molecule of the classical complement pathway, crucial for innate and adaptive immunity.
- The interaction between C1q and calreticulin is not fully understood, particularly in non-ER environments.
Purpose of the Study:
- To investigate the conditions under which calreticulin interacts with C1q.
- To elucidate the binding characteristics and molecular sites involved in the C1q-calreticulin interaction.
- To determine the functional implications of calreticulin binding to altered C1q conformations.
Main Methods:
- Surface immobilization of C1q and subsequent binding assays with calreticulin.
- Varying solution conditions including protein concentrations, salt concentrations, pH, and denaturing agents.
- Inhibition studies using serum amyloid P component and ovalbumin fragments.
- Characterization of binding kinetics and identification of involved protein domains.
Main Results:
- No direct interaction between C1q and calreticulin in solution at equal concentrations.
- Specific and rapid binding of calreticulin to immobilized, heat-treated, or immunoglobulin (Ig)-bound C1q.
- A two-step binding process: initial salt-sensitive phase involving C1q's collagen-like domain, followed by stable, salt-insensitive binding to calreticulin's peptide-binding site.
- Binding was inhibited by serum amyloid P component and modified ovalbumin, with reciprocal inhibition observed.
Conclusions:
- Calreticulin acts as a receptor for altered C1q conformations, particularly when C1q is bound to immune complexes.
- The interaction involves stable binding to calreticulin's peptide-binding site and initial interaction with C1q's collagen-like domain.
- Calreticulin's chaperone and scavenger functions may extend to the cell surface, facilitating the clearance of immune complexes and apoptotic cells.
Abstract:
The interaction between C1q and the chaperone calreticulin was studied under various conditions. When both proteins were present in equal amounts in solution, no interaction could be demonstrated. However, C1q immobilized on a hydrophobic surface, exposed to heat-treatment or bound to immunoglobulins (Igs) showed a strong, rapid and specific binding of calreticulin. The interaction appeared to be a two-step process, and the initial phase of interaction was sensitive to high concentrations of salt but not to a physiological salt concentration. The following strong binding was insensitive to salt and extremes of pH but sensitive to strongly denaturing agents (urea and guanidine). The sensitivity to salt during the initial phase of interaction was practically identical to that observed when calreticulin was bound to type V collagen. Binding between C1q and calreticulin could be inhibited by serum amyloid P component and by proteinase K-digested ovalbumin, and the binding of calreticulin to proteinase K-digested ovalbumin was shown to be inhibited by C1q. The data indicate that C1q binds stably to the peptide-binding site of calreticulin and that the initial binding of calreticulin to C1q involves the collagen-like domain of the C1q molecule. In conclusion, our results suggest calreticulin as a potential receptor for an altered conformation of C1q as occurs during binding to Igs. Thus, the chaperone and protein-scavenging function of calreticulin may extend from the endoplasmic reticulum to the topologically equivalent cell surface, where it may contribute to the elimination of immune complexes and apoptotic cells.
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