Molecular Crosstalk Between Adherens Junction Proteins, E-cadherin and Nectin-4.

Cell-cell junctions formed by the association of cell adhesion molecules facilitate physiological events necessary for growth and development of multicellular organisms. Among them, cadherins and nectins organize and assemble to form adherens junction, which thereby mechanically couples interacting cells. A detailed understanding of the crosstalk involving these cell adhesion molecules is fundamental to the study of the various developmental processes. Although, cadherins and nectins can recruit each other in the adherens junction through an interplay of cytoplasmic adaptor molecules, here, we report a direct interaction between N-terminal extracellular domains of E-cadherin and nectin-4 as demonstrated by surface plasmon resonance (SPR) and Atomic Force Microscopy (AFM)-based single molecule force spectroscopy (SMFS). Kinetic studies using SPR demonstrate the binding between the ectodomains of E-cadherin and nectin-4 with a KD of 3.7 ± 0.7 µM and KD of 5.4 ± 0.2 µM (reciprocal experiment). AFM-based SMFS experiments also support interaction between the ectodomains of E-cadherin and nectin-4 with the koff value of 31.48 ± 1.53 s-1 and the lifetime of the complex of 0.036 ± 0.0026 s. We thus propose a cell adhesion mechanism mediated by E-cadherin and nectin-4, which can have functional significance in early embryogenesis as evident from the expression pattern of both the proteins during early development.

Molecular and structural basis of TIGIT: Nectin-4 interaction, a recently discovered pathway crucial for cancer immunotherapy.

TIGIT (T cell immunoglobulin and ITIM domain) is an inhibitory receptor expressed on T and NK cells that interact with cell surface glycoprotein belonging to the nectin and nectin-like family of cell adhesion molecules, particularly nectin-2 and nectin-like 5 (PVR). Nectin-4 has been recently identified as a novel ligand for TIGIT and the interaction among them inhibits NK cell cytotoxicity. In this study, biophysical experiments were conducted to decipher the mechanism of this novel interaction, followed by structure-guided mutagenesis studies to map the nectin-4 binding interface on TIGIT. Using surface plasmon resonance, we deduced that TIGIT recognizes the membrane distal ectodomain of nectin-4 and the interaction is weaker than the well-characterized TIGIT: nectin-2 interaction. Deciphering the molecular basis of this newly identified interaction between TIGIT and nectin-4 will provide us important insight into the manipulation of this inhibitory signaling pathway, especially targeting cancer cells overexpressing nectin-4 that evade the immune surveillance of the body.

High-resolution crystal structure of LpqH, an immunomodulatory surface lipoprotein of Mycobacterium tuberculosis reveals a distinct fold and a conserved cleft on its surface.

Tuberculosis, caused by Mycobacterium tuberculosis, is predominantly a disease of the lungs acquired by inhaling mycobacteria from infected individuals via airborne droplets. In order to facilitate their entry into the alveolar macrophages, mycobacteria have a collection of pathogen-associated molecular patterns (PAMPs) on their surface that are known to detect certain pattern recognition receptors present on the surface of host cells. A major group of these PAMPs includes mycobacterial lipoproteins, of which, the 19 kDa surface antigen LpqH, has been reported to play a critical role in both host-pathogen interactions as well as pleiotropic immune regulation. Despite its crucial involvement in tuberculosis, the detailed structure-function relationship of this protein remains to be explored. Here, we report the high-resolution crystal structure of the non-acylated LpqH (LpqH48-159) at a resolution of 1.26 Å, which adopts a unique fold. Flow cytometry-based experiments show that the protein can bind and induce apoptosis in PMA-activated human monocytic cell line THP-1, indicative of the preservation of functionality of the protein. Furthermore, analysis of conservation of LpqH sequences from Mycobacterium species reveals a patch of conserved residues on the surface which may play a role in its binding partner recognition and hence in host-pathogen interaction.