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Mycobacterial lipoarabinomannan recognition requires a receptor that shares components of the endotoxin signaling
R Savedra1, R L Delude, R R Ingalls
1The Maxwell Finland Laboratory for Infectious Diseases, Department of Medicine, Boston City Hospital, MA 02118, USA.
Abstract:
Phagocytic leukocytes respond to a variety of bacterial products including Gram-negative bacterial LPS and mycobacterial lipoarabinomannan (LAM). Anti-CD14 mAbs have been shown to block LPS and LAM activation of myeloid cells, suggesting that CD14 is required for cellular recognition of both ligands. Activation of undifferentiated promonomyelocytic THP-1 cells with LAM or LPS under serum-free conditions was enhanced in the presence of recombinant soluble CD14 (rsCD14). LPS binding protein (LBP), which is present in normal serum, further enhanced the sensitivity of undifferentiated THP-1 cells to both ligands even in the absence of rsCD14. Although CD14-transfected Chinese hamster ovary and human HT1080 fibrosarcoma cell lines can be activated by LPS, neither cell line was activated by LAM. Furthermore, U373 astrocytoma cells, which respond to LPS using sCD14 and LBP, failed to be activated by LAM in the presence of rsCD14 and rLBP. We then tested the effects of lipid IVA and Rhodobacter sphaeroides lipid A, compounds that function as endotoxin inhibitors in human cells by interacting with a molecule thought to be a CD14-dependent LPS signal transducer. Both lipid IVA and R. sphaeroides lipid A inhibited the effects of LPS and LAM in THP-1 cells. Thus, the LPS and LAM receptors share CD14, LBP, and a putative endotoxin antagonist-inhibitable signal transducing component. However, the LAM signaling system appears to require an additional receptor component whose expression is restricted to cells of hemopoietic origin.
Insights
CD14 and LPS binding protein (LBP) are essential for recognizing bacterial lipopolysaccharide (LPS) and lipoarabinomannan (LAM). While both ligands share CD14 and LBP, LAM requires an additional hemopoietic cell-specific receptor.
Area of Science:
- Immunology
- Cell Biology
- Microbiology
Background:
- Phagocytic leukocytes recognize bacterial components like LPS and LAM.
- CD14 is implicated in the cellular response to both LPS and LAM.
- LPS binding protein (LBP) enhances cellular sensitivity to these bacterial ligands.
Purpose of the Study:
- To investigate the shared and distinct mechanisms of LPS and LAM recognition by myeloid cells.
- To identify the roles of CD14 and LBP in mediating cellular responses to LPS and LAM.
- To explore potential differences in signaling pathways for LPS and LAM.
Main Methods:
- Utilizing THP-1 cells (promonocytic) and various cell lines (CHO, HT1080, U373) with and without CD14 transfection.
- Employing recombinant soluble CD14 (rsCD14) and recombinant LBP (rLBP) to study ligand activation.
- Assessing cellular responses to LPS and LAM in the presence of endotoxin inhibitors (Lipid IVA, R. sphaeroides lipid A).
Main Results:
- rsCD14 enhanced THP-1 cell activation by LPS and LAM under serum-free conditions.
- LBP further increased THP-1 cell sensitivity to LPS and LAM, even without rsCD14.
- CD14-transfected cells responded to LPS but not LAM, and U373 cells failed to respond to LAM with rsCD14/rLBP.
- Endotoxin inhibitors blocked both LPS and LAM effects in THP-1 cells.
- LAM signaling appears to require a hemopoietic cell-specific receptor component.
Conclusions:
- CD14 and LBP are crucial for LPS and LAM recognition, indicating shared receptor components.
- A putative signal transducer inhibited by endotoxin antagonists is common to both LPS and LAM signaling.
- Lipoarabinomannan (LAM) signaling necessitates an additional receptor distinct from the LPS pathway, restricted to cells of hemopoietic origin.