A main function of epithelia and endothelia is keeping the composition of the body and its compartments reasonably constant against a variable and potentially harmful environment. Between cells, barriers form against unlimited and uncontrolled passage of substances and pathogens. The tight junction (TJ) is the apical most junctional protein complex in many types of epithelial and endothelial cells. The TJ can be sub-divided into the integral membrane and cytoplasmic proteins. Occludin, tricellulin, marvelD3, and the claudins (of which there are 27 members) are tetra-spanning membrane proteins. The relationship and assembly of TJ membrane proteins remain unclear. How TJs are assemble? How TJ proteins form and preserve the cell-cell interactions? How is their directional oligomerization achieved? How is their expression and assembly changed during pathogenic events? Traditionally, research efforts focused on the barrier and fence functions of TJs; however, there is a new movement in the field, which is to understand how TJ proteins participate in cell proliferation, transformation, and metastasis suppression. Our laboratory is addressing these questions using a bottom-up approach. Our tools are molecular and structural biology, protein engineering, biochemistry, biophysics, microbiology, cell biology, and physiology. Our objective is to understand biomolecular (self-) assembly of TJs, and function of its proteins’ structure and interactions. We aim to apply this knowledge to important problems in human health, biomolecular engineering, and nanotechnology.