Re-examination of CD91 function in GRP94 (glycoprotein 96) surface binding, uptake, and peptide cross-presentation

Area of Science:

• Immunology
• Cell Biology
• Molecular Biology

Background:

• Heat shock protein GRP94 (gp96)-peptide complexes are internalized by antigen-presenting cells (APCs) for cross-presentation to CD8+ T cells.
• Conflicting data exists regarding the surface receptors involved in GRP94 recognition and uptake, particularly the role of CD91 (LRP1).

Purpose of the Study:

• To investigate the function of CD91 in GRP94 surface binding, endocytosis, and peptide cross-presentation.
• To elucidate the mechanisms underlying GRP94 uptake and its implications for T cell activation.

Main Methods:

• Utilized mouse embryonic fibroblast (MEF) cell lines with reduced or absent CD91 expression (via RNA interference or genetic disruption).
• Assessed surface binding and endocytosis of GRP94 N-terminal domain (GRP94.NTD) and receptor-associated protein.
• Investigated GRP94.NTD binding after treatment with heparin, sodium chlorate, or heparinase II.
• Examined GRP94.NTD-peptide cross-presentation in CD91-expressing DC2.4 dendritic cells.

Main Results:

• Reduction or loss of CD91 expression abrogated receptor-associated protein binding and uptake but did not affect GRP94.NTD binding or uptake.
• Heparin sulfate proteoglycans were identified as mediators of GRP94.NTD surface binding.
• GRP94.NTD-peptide cross-presentation in dendritic cells was independent of CD91 ligands and primarily occurred via fluid-phase uptake.

Conclusions:

• CD91 is not essential for GRP94 surface binding, endocytosis, or peptide cross-presentation.
• Heparin sulfate proteoglycans play a significant role in GRP94 binding to cell surfaces.
• These findings resolve conflicting data and highlight alternative pathways for GRP94 uptake and cross-presentation.