Analyzing Differential Dynamics of CD9 and CD81 with Single Molecule Tracking

Abstract

Tetraspanins CD9 and CD81 are thought to modulate lateral organization of plasma membrane in time and space, hence to have an implication in many cellular processes like adhesion, migration, signaling and fusion. Few years ago, using single molecule tracking (SMT), our group has demonstrated that CD9 proteins can associate to and dissociate from Tetraspanin Enriched Microdomains (TEMs) and that this dynamics can be modulated by cholesterol concentration and cytoskeleton organization. More recently, our collaborators and we have also shown that CD9 is specifically recruited and confined in assembly sites of HIV-1, in correlation with the budding process. Surprisingly, when we investigated the influence of virus egress on CD81, we discovered that, even if this tetraspanin is also trapped into budding sites, its dynamics is different from that of CD9. In order to identify the key regions in CD81 structure that lead to its slow motions as compared to CD9's, we studied the dynamics of chimeric forms of these two tetraspanins using SMT. We also explored the role of the cytoskeleton upon the dynamics of these proteins using specific drugs to disorganize actin or microtubule network. Our results reveal that several domains of the CD81 protein are responsible for its membrane behavior and that both CD9 and CD81 dynamics and partitioning within plasma membrane are largely dependent on the cytoskeleton integrity. Keywords : tetraspanin, CD81, CD9, membrane dynamics, microdomains, cytoskeleton, single molecule tracking.