The role of the tetraspanin CD81 in the generation and maintenance of the neuronal extracellular matrix

• Applicants: Professor Dr. Thorsten Lang; Professor Silvio-Olivier Rizzoli, Ph.D.
• Subject Area: Molecular Biology and Physiology of Neurons and Glial Cells
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 552580959

Tetraspanins are a family of small membrane proteins involved in a multitude of cellular processes, including the generation of extracellular vesicles (EVs). The functions of tetraspanins and EVs in the brain are not fully understood. We have recently combined forces to investigate the effects of tetraspanins in neurons, and uncovered a major phenotype, relating to the extracellular matrix (ECM), a lattice-like structure, composed of glycosylated proteins, which coats the surfaces of neurons. We found that the tetraspanin CD81 is essential for the maintenance and the dynamics of the ECM. First, during neuron maturation, CD81 mutants strongly reduce transfer of neurocan from glia to neurons, which causes the trapping of neurocan by glia. Second, CD81 mutants blocked the endocytosis and recycling of ECM molecules, and eventually caused the removal of more than 50% of the ECM within the entire culture, within a few days. These effects were caused by a minority of mutant-expressing cells, arguing that the CD81-dependent mechanisms affect entire cell populations, rather than single cells, probably through sharing of EVs between cells. In glia, we further observed that IGSF8, a member of the immunoglobulin superfamily that binds to the neuronal-specific ECM component Tenascin-R and that is an interaction partner of CD81, agglomerates into spots at the cell surface, a behavior we have also characterized for CD81 and IGSF8 in non-neuronal cells. These observations, which argue for a far stronger role of tetraspanins in brain function than ever considered in the past, lead to the hypothesis that tetraspanins (and CD81 in particular) are involved in the regulation of the ECM, both through interactions on cell surfaces and through EVs. We now propose to test this hypothesis, and to determine the cellular pathways behind the tetraspanin-dependent ECM modulation. First, we will study the architecture of CD81/IGSF8-agglomerates and how they recruit ECM molecules in dependency of CD81. Second, we will characterize EV composition and the EV involvement in ECM molecule recruitment. Third, we will study the pathway enabling the ECM donor and receiver cells to communicate. Fourth, we will analyze the organization of CD81, IGSF8 and ECM partners on the surface of cells that received these components. Fifth, we will analyze the relevance of the CD81 mutations on neuronal physiology. Our work will provide unprecedented insight in the function of brain tetraspanins, and should contribute to many branches of neuroscience and cell biology, covering topics from principles of membrane organization to ECM dynamics and EV behavior.

• DFG Programme: Research Grants