Abstract
BackgroundSeveral pathogenic bacteria utilize receptors of the CEACAM family to attach to human cells. Binding to different members of this receptor family can result in uptake of the bacteria. Uptake of Neisseria gonorrhoeae, a Gram-negative human pathogen, via CEACAMs found on epithelial cells, such as CEACAM1, CEA or CEACAM6, differs mechanistically from phagocytosis mediated by CEACAM3, a CEACAM family member expressed selectively by human granulocytes.Principal FindingsWe find that CEACAM1- as well as CEACAM3-mediated bacterial internalization are accompanied by a rapid increase in phosphatidylinositol-3,4,5 phosphate (PI(3,4,5)P) at the site of bacterial entry. However, pharmacological inhibition of phosphatidylinositol-3′ kinase (PI3K) selectively affects CEACAM1-mediated uptake of Neisseria gonorrhoeae. Accordingly, overexpression of the PI(3,4,5)P phosphatase SHIP diminishes and expression of a constitutive active PI3K increases CEACAM1-mediated internalization of gonococci, without influencing uptake by CEACAM3. Furthermore, bacterial uptake by GPI-linked members of the CEACAM family (CEA and CEACAM6) and CEACAM1-mediated internalization of N. meningitidis by endothelial cells require PI3K activity. Sensitivity of CEACAM1-mediated uptake toward PI3K inhibition is independent of receptor localization in cholesterol-rich membrane microdomains and does not require the cytoplasmic or the transmembrane domain of CEACAM1. However, PI3K inhibitor sensitivity requires the IgC2-like domains of CEACAM1, which are also present in CEA and CEACAM6, but which are absent from CEACAM3. Accordingly, overexpression of CEACAM1 IgC2 domains blocks CEACAM1-mediated internalization.ConclusionsOur results provide novel mechanistic insight into CEACAM1-mediated endocytosis and suggest that epithelial CEACAMs associate in cis with other membrane receptor(s) via their extracellular domains to trigger bacterial uptake in a PI3K-dependent manner.
Highlights
Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is a widely expressed glycoprotein of the CEACAM family, which in humans comprises 12 genes and a number of pseudogenes [1,2]
Our results provide novel mechanistic insight into CEACAM1-mediated endocytosis and suggest that epithelial CEACAMs associate in cis with other membrane receptor(s) via their extracellular domains to trigger bacterial uptake in a phosphatidylinositol-39 kinase (PI3K)-dependent manner
If PI(3,4,5)P might be involved in CEACAM1-mediated internalization, we cotransfected 293 cells with constructs encoding GFP-tagged PH domain of Bruton’s tyrosine kinase (Btk) together with either mKate-tagged CEACAM1 or mKate-tagged CEACAM3
Summary
Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is a widely expressed glycoprotein of the CEACAM family, which in humans comprises 12 genes and a number of pseudogenes [1,2]. CEACAM1 is involved in a broad spectrum of cellular processes ranging from tissue morphogenesis and apoptosis, to insulin homeostasis, angiogenesis, or regulation of T-cell activity [2] Another member of the CEACAM family is CEACAM3, which is exclusively expressed on granulocytes and harbours a single IgV-like domain followed by a transmembrane helix and a cytoplasmic domain [4]. CEACAM1 and CEACAM3, two additional members of the CEACAM family, namely CEA (the product of the CEACAM5 gene) and CEACAM6, can serve as cellular receptors for a range of gram-negative bacteria [5,6,7,8,9,10,11]. Uptake of Neisseria gonorrhoeae, a Gramnegative human pathogen, via CEACAMs found on epithelial cells, such as CEACAM1, CEA or CEACAM6, differs mechanistically from phagocytosis mediated by CEACAM3, a CEACAM family member expressed selectively by human granulocytes
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