Monitoring the CEACAM1 Monomer and Dimer Distribution upon Cell-Substrate Contact

Abstract

The carcinoembryonic antigen-related cellular adhesion molecule 1 (CEACAM1) is an intercellular adhesion molecule that plays a key role in processes related to cell growth, differentiation, and infection in normal and cancerous cells. Expressed on the cell surface, its extracellular domain participates in interactions with CEACAMs on neighbouring cells, as well as interacting with Opa proteins expressed on Neisseria gonorrhoeae and Neisseria meningitides in order to facilitate bacterial entry. While CEACAM1 is known to exist in both monomeric and dimeric states that are heterogeneously distributed at the cell surface, which form participates in the various inter-cellular and bacterial interactions remains a mystery. Resolving this uncertainty is fundamental to understanding the role of the receptor, as the monomer-dimer status of CEACAM1 has been shown to differentially affect its ability to bind certain downstream signalling molecules. We are using live cell fluorescence and homogeneous Forster resonance energy transfer (homo-FRET) microscopy on a combined total internal reflection fluorescence polarization microscopy (TIRFPM) and confocal microscopy platform to investigate the distribution, dynamics, and monomer-dimer equilibrium of EYFP-labeled CEACAM1. Cells expressing the labeled receptor are monitored as they come into contact with CEACAM1-engaging surfaces, such as other cells or glass dishes that have been patterned with Opa+ Neisseria gonorrhoeae. These approaches will allow us to directly examine the initial stages of CEACAM1 association with CEACAM1-interacting proteins and will answer critical questions regarding the nature of these interactions. Such insights are essential for understanding the nature of CEACAM1 signalling, function, and regulation, and ultimately targeting its functions for purposes such as cancer therapy or the treatment of infection.