Rôle fonctionnel de l'interaction du CD154 avec le CD40 associé au CD20

CD154 is a 33-39 KDa type II transmembrane glycoprotein, belonging to the TNF (Tumor Necrosis Factor) superfamily. This molecule is expressed in a trimeric, inducible and transient form on the surface of several cell types, mainly on the surface of activated T cells and platelets. CD154 also exists in a soluble form with a molecular weight of 18 KDa (sCD154). This form comes in part from an enzymatic cleavage of the membrane bound CD154 on activated T cells and remains a biological active trimer. A close correlation exists between the increase of sCD154 in the plasma and the progression of the pathology in various autoimmune or cardiovascular diseases.  CD40 is the principal receptor of CD154. The interaction of CD154 with CD40 induces a bidirectional signal which is crucial for T-B cells interaction involving cell proliferation, cytokine production, B-cell differentiation, isotypic immunoglobulin switching and apoptosis. On the other hand, CD40 is physically associated with CD20 on the surface of B cells. Until now, CD20 has no ligand and its biological role is little known. However, this molecule has a very important apoptotic function using monoclonal antibodies such as Rituximab. The latter has shown a very important efficacy in the treatment of cancer and different autoimmune diseases. In addition to CD40, three other receptors of CD154 have been identified: αIIbβ3, α5β1 and αMβ2, belonging to the integrin family. CD154 interacts with these four receptors via different residues and induces different biological responses. However, we have demonstrated that only CD40, in its soluble or membrane form, is capable of inducing CD154 cleavage from the surface of T cells. This cleavage is mediated by the activation of ADAM-10 and ADAM-17 by the PKC pathway. The work illustrated in this thesis consists in studying the functional role of the association of CD40 with CD20 and the effect of this association on the responses induced by the CD154/CD40 dyad; and in determining the impact of the bidirectional signals, induced by this dyad, in the regulation of these responses. The results generated here show that the association of CD40 with CD20 amplifies cell death induced via CD40 or via CD20, and that signals from CD40 are involved in this amplification. This association also up-regulated the cleavage of CD154. However, the deletion of the cytoplasmic domain of CD40 affects this cleavage, showing that the signals from CD40 are also involved in this response. In addition, the deletion of the cytoplasmic domains of both CD40 and CD154, abolished completely the cleavage of CD154. Since sCD154 plays a role in the progression of several diseases, we have generated a non-cleavable form of CD154 that has shown greater cytotoxicity than the cleavable form, due to the retention of CD154 at the cell membrane inducing more pronounced signals. This form also showed more pronounced efficacy in others B cell responses such as proliferation and immunoglobulin production. This work will allow us to broaden our knowledge on the role of the CD154/CD40 couple in the regulation of many cellular responses, as well as that of the association of CD40 with CD20 in the amplification of these responses in order to optimize the outcome of these molecules in the treatment of several autoimmune diseases, cardiovascular and even in cancer.

Keyphrases: CD154, CD40, CD20, Association, Cleavage, Bidirectional signaling, Cellular responses