Emilio Tejera

CD81 regulates cell migration through its association with Rac GTPase

Data presented here provide evidence for a new direct interaction of the GTPase Rac with the C-terminal cytoplasmic domain of tetraspanin CD81. Tetraspanin-enriched, microdomain-dependent compartmentalization is a novel regulatory mechanism of Rac activity turnover, which provides a novel mechanism for regulation of cell motility by tetraspanins.

CD81 Controls Sustained T Cell Activation Signaling and Defines the Maturation Stages of Cognate Immunological Synapses

ABSTRACT In this study, we investigated the dynamics of the molecular interactions of tetraspanin CD81 in T lymphocytes, and we show that CD81 controls the organization of the immune synapse (IS) and T cell activation. Using quantitative microscopy, including fluorescence recovery after photobleaching (FRAP), phasor fluorescence lifetime imaging microscopy-Föster resonance energy transfer (phasorFLIM-FRET), and total internal reflection fluorescence microscopy (TIRFM), we demonstrate that CD81 interacts with ICAM-1 and CD3 during conjugation between T cells and antigen-presenting cells (APCs). CD81 and ICAM-1 exhibit distinct mobilities in central and peripheral areas of early and late T cell-APC contacts. Moreover, CD81–ICAM-1 and CD81-CD3 dynamic interactions increase over the time course of IS formation, as these molecules redistribute throughout the contact area. Therefore, CD81 associations unexpectedly define novel sequential steps of IS maturation. Our results indicate that CD81 controls the temporal progression of the IS and the permanence of CD3 in the membrane contact area, contributing to sustained T cell receptor (TCR)-CD3-mediated signaling. Accordingly, we find that CD81 is required for proper T cell activation, regulating CD3ζ, ZAP-70, LAT, and extracellular signal-regulated kinase (ERK) phosphorylation; CD69 surface expression; and interleukin-2 (IL-2) secretion. Our data demonstrate the important role of CD81 in the molecular organization and dynamics of the IS architecture that sets the signaling threshold in T cell activation.

Tetraspanins CD9 and CD151 at the immune synapse support T-cell integrin signaling

Understanding how the immune response is activated and amplified requires detailed knowledge of the stages in the formation of the immunological synapse (IS) between T lymphocytes and antigen‐presenting cells (APCs). We show that tetraspanins CD9 and CD151 congregate at the T‐cell side of the IS. Silencing of CD9 or CD151 blunts the IL‐2 secretion and expression of the activation marker CD69 by APC‐conjugated T lymphocytes, but does not affect the accumulation of CD3 or actin to the IS, or the translocation of the microtubule‐organizing center toward the T‐B contact area. CD9 or CD151 silencing diminishes the relocalization of α4β1 integrin to the IS and reduces the accumulation of high‐affinity β1 integrins at the cell–cell contact. These changes are accompanied by diminished phosphorylation of the integrin downstream targets FAK and ERK1/2. Our results suggest that CD9 and CD151 support integrin‐mediated signaling at the IS.

Association of syntenin-1 with M-RIP polarizes Rac-1 activation during chemotaxis and immune interactions

In this study, we describe that the PDZ protein syntenin-1 is a crucial element for the generation of signaling asymmetry during the cellular response to polarized extracellular cues. We analyze the role of syntenin-1 in the control of asymmetry in two independent models of T cell polarization – the migratory response to chemoattractants and the establishment of cognate interactions between T cells and antigen-presenting cells (APCs). A combination of mutant, biochemical and siRNA approaches demonstrate that syntenin-1 is vital for the generation of polarized actin structures such as the leading edge and the contact zone with APCs. We found that the mechanism by which syntenin-1 controls actin polymerization relies on its mandatory role for activation of the small GTPase Rac. Syntenin-1 controls Rac through a specific association with the myosin phosphatase Rho interacting protein (M-RIP), which occurs in response to phosphorylation of syntenin-1 by Src at Tyr4. Our data indicate the key role of syntenin-1 in the generation of functional asymmetry in T cells and provide a novel mechanistic link between receptor activation and actin polymerization and accumulation in response to extracellular stimulation.

Lanthanide complexes as imaging agents anchored on nano-sized particles of boehmite

The synthesis of boehmite nanoparticles modified with lanthanides (Eu, Tb and Gd) is described. Their synthesis, characterization and in vitro assays with HeLa cells were performed. The nuclear magnetic relaxation dispersion (NMRD) profiles of the two chelating moieties were studied. Imaging data from laser scanning confocal fluorescence microscopy and flow cytometry revealed that the nanoscaffolds were taken up by the cells, distributed throughout the cytoplasm and showed no toxicity. This platform could represent an alternative to silica-based inert matrices as imaging vehicles.

The Role of Tetraspanins in Cell Migration and Intercellular Adhesion

Tetraspanin-enriched microdomains (TEMs) are specialized platforms in the plasma membrane that include certain adhesion receptors, mainly integrins and receptors of the Ig superfamily. Insertion into TEMs increases the local concentration of these adhesion receptors, facilitating their function as avidity regulators. TEMs also regulate interaction and crosstalk between different receptors at the plasma membrane, as well as their internalization rate. Moreover, certain signaling pathways are regulated by association with tetraspanins. Thus, tetraspanins emerge as critical regulators of biological phenomena involving adhesion to the extracellular matrix or homotypic or heterotypic intercellular interactions. These proteins are implicated in different steps of cancer progression, the regulation of intercellular adhesion between polarized epithelial cells, angiogenesis, antigen presentation and extravasation of leukocytes or tumor cells. In addition, several pathogens hijack these tetraspanin-adhesion platforms to increase their infectivity.

Endothelial adhesion receptors are recruited to adherent leukocytes by inclusion in preformed tetraspanin nanoplatforms

Barreiro et al. 2008. J. Cell Biol. doi:10.1083/jcb.200805076 [OpenUrl][1][Abstract/FREE Full Text][2] [1]: {openurl}?query=rft_id%253Dinfo%253Adoi%252F10.1083%252Fjcb.200805076%26rft_id%253Dinfo%253Apmid%252F18955551%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%

1085 Quantification of specific adhesion to human endothelium of targeted anti-CD9 microbubbles. Results from an “In Vitro” model

Microbubble (µB) based contrast agents (CA) are used in ultrasound molecular imaging to identify specific targets. The aim of this study was to target µB to human endothelial cells (EC) and to evaluate the specificity and efficacy of their adhesion. Methods: Attachment of 5 different CA was assessed in an inverted flow chamber: albumin shell (AS), lipid shell (LS) and a biotinylated shell (BS) that was used not conjugated (naked) or conjugated with 2 monoclonal antibodies: antiCD9, specific for EC receptor CD9, and antiCD19, as a control since CD19 is not expressed by EC. Three types of culture dishes were prepared: Fibronectin (FN), as a control for non-specific chemical adhesion, and monolayers of EC isolated from umbilical cords, resting (rEC) and TNFactivated (aEC). Infusion of CA was recorded with video- microscopy. Wall shear stress was increased (0.5-5 dyn/cm 2 ) at 10 intervals of 1 minute. To rinse unspecific attachments, a washing period of 3 min with buffer up to 3