Abstract
Interaction of CD40 with CD154 leads to recruitment of both molecules into lipid rafts, resulting in bi-directional cell activation. The precise mechanism by which CD154 is translocated into lipid rafts and its impact on CD154 signaling remain largely unknown. Our aim is to identify the domain of CD154 facilitating its association to lipid rafts and the impact of such association on signaling events and cytokine production. Thus, we generated Jurkat cell lines expressing truncated CD154 lacking the cytoplasmic domain or chimeric CD154 in which the transmembrane domain was replaced by that of transferrin receptor I, known to be excluded from lipid rafts. Our results show that cell stimulation with soluble CD40 leads to the association of CD154 wild-type and CD154-truncated, but not CD154-chimera, with lipid rafts. This is correlated with failure of CD154-chimera to activate Akt and p38 MAP kinases, known effectors of CD154 signaling. We also found that CD154-chimera lost the ability to promote IL-2 production upon T cell stimulation with anti-CD3/CD28 and soluble CD40. These results demonstrate the implication of the transmembrane domain of CD154 in lipid raft association, and that this association is necessary for CD154-mediated Akt and p38 activation with consequent enhancement of IL-2 production.
Highlights
CD154, known as CD40 ligand (CD40L), is a 33-kDa type II transmembrane protein that belongs to the tumour necrosis factor (TNF) superfamily [1]
In order to investigate whether CD154 translocation into lipid rafts requires an intracellular signaling, and to determine if the cytoplasmic domain of CD154 is involved in such translocation, we used Jurkat E6.1 cells transfected with an empty vector, CD154 wild type (CD154wt), or a truncated form of CD154 lacking its cytoplasmic domain (CD154Dcyto)
We investigated the role of lipid rafts in CD154 signaling
Summary
CD154, known as CD40 ligand (CD40L), is a 33-kDa type II transmembrane protein that belongs to the tumour necrosis factor (TNF) superfamily [1]. CD154 is mainly expressed on activated T cells and platelets and on a wide range of cell types including dendritic cells, mast cells, monocytes, macrophages, fibroblasts, endothelial cells and others [2,3]. CD154 costimulation was shown to enhance IL-2-mediated T cell proliferation and generation of cytotoxic cells [9]. Other biological functions of CD154 stimulation include potentiation of IL-4 production in CD3-CD28 activated T cells [10]. Costimulating T cells via CD3 and CD154 was shown to enhance synthesis of IL-10, TNFa and IFNc, and at later stages to promote apoptosis [11]. CD154 stimulation was shown to induce phosphorylation of MAPKs such as JNK and p38, downstream of Src kinases and Rac-1 activation [13]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
