Cross-Talk between CD14 and Complement Receptor 3 Promotes Phagocytosis of Mycobacteria: Regulation by Phosphatidylinositol 3-Kinase and Cytohesin-1

Abstract

The glycosylphosphatidyl anchored molecule CD14 to the monocyte membrane plays a prominent role in innate immunity, and the paradigms for CD14 selective signaling are beginning to be elucidated. In this study, transfected human monocytic cell line THP-1 and Chinese hamster ovary (CHO) fibroblastic cells were used to examine phagocytosis of Mycobacterium bovis bacillus Calmette-Guérin (BCG). Flow cytometry was combined with molecular and biochemical approaches to demonstrate a dual mechanism for BCG internalization involving either CD14 alone or a CD14-regulated complement receptor (CR)3-dependent pathway. Phagocytosis by CD14-positive THP-1 cells was attenuated by phosphatidylinositol-3 inhibitors LY294002 and wortmannin and experiments using transfected CHO cells showed substantial accumulation of phosphatidylinositol-3,4,5-trisphosphate at the BCG attachment site in CHO cells expressing CD14 and TLR2 suggesting that bacteria bind to CD14 and use TLR2 to initiate a PI3K signaling pathway. Additional experiments using blocking Abs showed that anti-TLR2 Abs inhibit phagocytosis of BCG by THP-1 cells. Furthermore, knockdown of cytohesin-1, a PI3K-regulated adaptor molecule for β2 integrin activation, specifically abrogated CD14-regulated CR3 ingestion of BCG consistent with the observation of physical association between CR3 and cytohesin-1 in cells stimulated with mycobacterial surface components. These findings reveal that mycobacteria promote their uptake through a process of “inside-out” signaling involving CD14, TLR2, PI3K, and cytohesin-1. This converts low avidity CR3 into an active receptor leading to increased bacterial internalization.

Subversion of Innate Immunity by Periodontopathic Bacteria via Exploitation of Complement Receptor-3

“…Our initial report that P. gingivalis fimbriae stimulate TLR2 inside-out signaling for CR3 activation was published concomitantly with a study by an independent group which showed that mycobacterial lipoarabinomannan also activates this proadhesive pathway (Sendide et al, 2005). Strikingly, mycobacteria exploit the TLR2/ CR3 pathway for promoting their entry into monocytes/macrophages (Sendide et al, 2005) where they can parasitize (Ernst, 1998).…”

“…Strikingly, mycobacteria exploit the TLR2/ CR3 pathway for promoting their entry into monocytes/macrophages (Sendide et al, 2005) where they can parasitize (Ernst, 1998). The potential for CR3 exploitation by certain pathogens may, at least partly, be related to the notion that CR3 is not linked to vigorous microbicidal mechanisms, in contrast to most phagocytic receptors (Lowell, 2006;Wright and Silverstein, 1983;Yamamoto and Johnston, 1984;Caron and Hall, 1998;Rosenberger and Finlay, 2003).…”

AKAP13, a RhoA GTPase-specific Guanine Exchange Factor, Is a Novel Regulator of TLR2 Signaling

“…Further, the GEF for ARF-GTPase, cytohesin-1, is involved in CD14/TLR2-mediated phagocytosis of mycobacteria by human macrophages. Knockdown of cytohesin-1 resulted in a marked attenuation of phagocytosis of mycobacteria (22).…”

Engagement of CD14 Mediates the Inflammatory Potential of Monosodium Urate Crystals

“…However, we have not yet determined whether experimental addition of sCD14, or chimerism devised by delivery of congenic CD14-expressing phagocytes, can modulate MSU crystal-induced inflammation of air pouches in the CD14-deficient mouse. Last, MSU crystals engage the integrin CD11b/CD18, and engagement of CD11b/CD18 and phagocyte activation by certain bacterial products are mediated by complex formation of CD14, TLR2, and CD11b/CD18 (28). Hence, it is conceivable that the role of CD14 in MSU crystal-induced inflammation is primarily via macromolecular complex formation with TLRs and integrins that are directly responsible for modulating cellular responsiveness to MSU crystals.…”

“…CD14 lacks both transmembrane and intracellular domains. However, the association of CD14 with other plasma membrane molecules, including formation of cell surface complexes of CD14 with TLR2, TLR4 (22,23), and certain integrins and FcR (28,29) provide the means for CD14 to modulate signal transduction.…”