The CEACAM1 transmembrane domain, but not the cytoplasmic domain, directs internalization of human pathogens via membrane microdomains

Abstract

SummarySeveral bacterial pathogens exploit carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) to promote attachment and uptake into eukaryotic host cells. The widely expressed isoform CEACAM1 is involved in cell-cell adhesion, regulation of cell proliferation, insulin homeostasis, and neo-angiogenesis, processes that depend on the cytoplasmic domain of CEACAM1. By analysing the molecular requirements for CEACAM1-mediated internalization of bacteria, we surprisingly find that the CEACAM1 cytoplasmic domain is completely obsolete for bacterial uptake. Accordingly, CEACAM1-4L as well as a CEACAM1 mutant with a complete deletion of the cytoplasmic domain (CEACAM1 DCT) promote equivalent internalization of several human pathogens. CEACAM1-4L-and CEACAM1 DCT-mediated uptake proceeds in the presence of inhibitors of actin microfilament dynamics, which is in contrast to CEACAM3-mediated internalization. Bacteria-engaged CEACAM1-4L and CEACAM1 DCT, but not CEACAM3, localize to a gangliosid GM1-and GPI-anchored protein-containing portion of the plasma membrane. In addition, interference with cholesterol-rich membrane microdomains severely blocks bacterial uptake via CEACAM1-4L and CEACAM1 DCT, but not CEACAM3. Similar to GPIanchored CEACAM6, both CEACAM1-4L as well as CEACAM1 DCT partition into a low-density, Tritoninsoluble membrane fraction upon receptor clustering, whereas CEACAM3 is not detected in this fraction. Bacterial uptake by truncated CEACAM1 or chimeric CEACAM1/CEACAM3 molecules reveals that the transmembrane domain of CEACAM1 is responsible for its association with membrane microdomains. Together, these data argue for a functional role of lipid rafts in CEACAM1-mediated endocytosis that is promoted by the transmembrane domain of the receptor and that might be relevant for CEACAM1 function in physiologic settings.

Caveolin limits membrane microdomain mobility and integrin-mediated uptake of fibronectin-binding pathogens

“…7B). Importantly, mobility of CEACAM3, a receptor that does not localize to membrane microdomains (Muenzner et al, 2008;Schmitter et al, 2007), is not altered in Cav1 -/-cells (Fig. 7C), whereas mobility of GM1, as a second independent marker of membrane microdomains, is also increased in Cav1 -/-cells (supplementary material Fig.…”

“…Total cell-associated bacteria were determined in separate samples by omitting the incubation with gentamicin/lysostaphin. Cell lysis and western blotting were performed as described (Muenzner et al, 2008).…”

Carcinoembryonic antigen (CEA)-related cell adhesion molecules are co-expressed in the human lung and their expression can be modulated in bronchial epithelial cells by non-typable Haemophilus influenzae, Moraxella catarrhalis, TLR3, and type I and II interferons

“…CEACAM1-S can thus not only mediate adhesion as well as internalization of pathogens [27], but can also control the signaling activities of the long CEACAM1 isoforms [28].…”

CEACAM3: An innate immune receptor directed against human-restricted bacterial pathogens

“…In line with the critical role of Rac, actin cytoskeleton dynamics are essential for CEACAM3-initiated bacterial uptake, whereas other CEACAMs appear to be internalized via lipid raft-dependent processes (Muenzner et al, 2008;Schmitter et al, 2007b). Lipid raft association and, therefore, sensitivity to cholesterol depletion, discriminate between bacterial internalization via CEACAM1 or glycosylphosphatidyl-inositol (GPI)-anchored CEACAMs (CEA and CEACAM6) and CEACAM3-mediated uptake (Muenzner et al, 2008).…”

“…Lipid raft association and, therefore, sensitivity to cholesterol depletion, discriminate between bacterial internalization via CEACAM1 or glycosylphosphatidyl-inositol (GPI)-anchored CEACAMs (CEA and CEACAM6) and CEACAM3-mediated uptake (Muenzner et al, 2008). In this regard it is interesting to point out that the opsonin-independent phagocytosis of Opa protein expressing bacteria by human granulocytes is not sensitive to cholesterol depletion (Schmitter et al, 2007b).…”