Function of the Tetraspanin CD151–α6β1 Integrin Complex during Cellular Morphogenesis

Upon plating on basement membrane Matrigel, NIH3T3 cells formed an anastomosing network of cord-like structures, inhibitable by anti-α6β1 integrin antibodies. For NIH3T3 cells transfected with human CD151 protein, the formation of a cord-like network was also inhibitable by anti-CD151 antibodies. Furthermore, CD151 and α6β1 were physically associated within NIH3T3 cells. On removal of the short 8-amino acid C-terminal CD151 tail (by deletion or exchange), exogenous CD151 exerted a dominant negative effect, as it almost completely suppressed α6β1-dependent cell network formation and NIH3T3 cell spreading on laminin-1 (an α6β1 ligand). Importantly, mutant CD151 retained α6β1 association and did not alter α6β1-mediated cell adhesion to Matrigel. In conclusion, the CD151–α6β1 integrin complex acts as a functional unit that markedly influences cellular morphogenesis, with the CD151 tail being of particular importance in determining the “outside-in” functions of α6β1-integrin that follow ligand engagement. Also, antibodies to α6β1 and CD151 inhibited formation of endothelial cell cord-like networks, thus pointing to possible relevance of CD151–α6β1 complexes during angiogenesis

Palmitoylation of Tetraspanin Proteins: Modulation of CD151 Lateral Interactions, Subcellular Distribution, and Integrin-dependent Cell Morphology

Here we demonstrate that multiple tetraspanin (transmembrane 4 superfamily) proteins are palmitoylated, in either the Golgi or a post-Golgi compartment. Using CD151 as a model tetraspanin, we identified and mutated intracellular N-terminal and C-terminal cysteine palmitoylation sites. Simultaneous mutations of C11, C15, C242, and C243 (each to serine) eliminated >90% of CD151 palmitoylation. Notably, palmitoylation had minimal influence on the density of tetraspanin protein complexes, did not promote tetraspanin localization into detergent-resistant microdomains, and was not required for CD151-α3β1 integrin association. However, the CD151 tetra mutant showed markedly diminished associations with other cell surface proteins, including other transmembrane 4 superfamily proteins (CD9, CD63). Thus, palmitoylation may be critical for assembly of the large network of cell surface tetraspanin-protein interactions, sometimes called the “tetraspanin web.” Also, compared with wild-type CD151, the tetra mutant was much more diffusely distributed and showed markedly diminished stability during biosynthesis. Finally, expression of the tetra-CD151 mutant profoundly altered α3 integrin-deficient kidney epithelial cells, such that they converted from a dispersed, elongated morphology to an epithelium-like cobblestone clustering. These results point to novel biochemical and biological functions for tetraspanin palmitoylation