Tetraspanins regulate ADAM10-mediated cleavage of TNF-alpha and epidermal growth factor.

Several cytokines and growth factors are released by proteolytic cleavage of a membrane-anchored precursor, through the action of ADAM (a disintegrin and metalloprotease) metalloproteases. The activity of these proteases is regulated through largely unknown mechanisms. In this study we show that Ab engagement of several tetraspanins (CD9, CD81, CD82) increases epidermal growth factor and/or TNF-alpha secretion through a mechanism dependent on ADAM10. The effect of anti-tetraspanin mAb on TNF-alpha release is rapid, not relayed by intercellular signaling, and depends on an intact MEK/Erk1/2 pathway. It is also associated with a concentration of ADAM10 in tetraspanin-containing patches. We also show that a large fraction of ADAM10 associates with several tetraspanins, indicating that ADAM10 is a component of the "tetraspanin web." These data show that tetraspanins regulate the activity of ADAM10 toward several substrates, and illustrate how membrane compartmentalization by tetraspanins can control the function of cell surface proteins such as ectoproteases.

Single-molecule analysis of CD9 dynamics and partitioning reveals multiple modes of interaction in the tetraspanin web.

Tetraspanins regulate cell migration, sperm-egg fusion, and viral infection. Through interactions with one another and other cell surface proteins, tetraspanins form a network of molecular interactions called the tetraspanin web. In this study, we use single-molecule fluorescence microscopy to dissect dynamics and partitioning of the tetraspanin CD9. We show that lateral mobility of CD9 in the plasma membrane is regulated by at least two modes of interaction that each exhibit specific dynamics. The majority of CD9 molecules display Brownian behavior but can be transiently confined to an interaction platform that is in permanent exchange with the rest of the membrane. These platforms, which are enriched in CD9 and its binding partners, are constant in shape and localization. Two CD9 molecules undergoing Brownian trajectories can also codiffuse, revealing extra platform interactions. CD9 mobility and partitioning are both dependent on its palmitoylation and plasma membrane cholesterol. Our data show the high dynamic of interactions in the tetraspanin web and further indicate that the tetraspanin web is distinct from raft microdomains.

Hepatocyte permissiveness to Plasmodium infection is conveyed by a short and structurally conserved region of the CD81 large extracellular domain.

Invasion of hepatocytes by Plasmodium sporozoites is a prerequisite for establishment of a malaria infection, and thus represents an attractive target for anti-malarial interventions. Still, the molecular mechanisms underlying sporozoite invasion are largely unknown. We have previously reported that the tetraspanin CD81, a known receptor for the hepatitis C virus (HCV), is required on hepatocytes for infection by sporozoites of several Plasmodium species. Here we have characterized CD81 molecular determinants required for infection of hepatocytic cells by P. yoelii sporozoites. Using CD9/CD81 chimeras, we have identified in CD81 a 21 amino acid stretch located in a domain structurally conserved in the large extracellular loop of tetraspanins, which is sufficient in an otherwise CD9 background to confer susceptibility to P. yoelii infection. By site-directed mutagenesis, we have demonstrated the key role of a solvent-exposed region around residue D137 within this domain. A mAb that requires this region for optimal binding did not block infection, in contrast to other CD81 mAbs. This study has uncovered a new functionally important region of CD81, independent of HCV E2 envelope protein binding domain, and further suggests that CD81 may not interact directly with a parasite ligand during Plasmodium infection, but instead may regulate the function of a yet unknown partner protein.