NSOM/QD-Based Visualization of GM1 Serving as Platforms for TCR/CD3 Mediated T-Cell Activation

Abstract

Direct molecular imaging of nanoscale relationship between T-cell receptor complexes (TCR/CD3) and gangliosidosis GM1 before and after T-cell activation has not been reported. In this study, we made use of our expertise of near-field scanning optical microscopy(NSOM)/immune-labeling quantum dots- (QD-)based dual-color imaging system to visualize nanoscale profiles for distribution and organization of TCR/CD3, GM1, as well as their nanospatial relationship and their correlation with PKCθ signaling cascade during T-cell activation. Interestingly, after anti-CD3/anti-CD28 Ab co-stimulation, both TCR/CD3 and GM1 were clustered to form nanodomains; moreover, all of TCR/CD3 nanodomains were colocalized with GM1 nanodomains, indicating that the formation of GM1 nanodomains was greatly correlated with TCR/CD3 mediated signaling. Specially, while T-cells were pretreated with PKCθ signaling inhibitor rottlerin to suppress IL-2 cytokine production, no visible TCR/CD3 nanodomains appeared while a lot of GM1 nanodomains were still observed. However, while T-cells are pretreated with PKCα β signaling inhibitor GÖ6976 to suppress calcium-dependent manner, all of TCR/CD3 nanodomains were still colocalized with GM1 nanodomains. These findings possibly support the notion that the formation of GM1 nanodomains indeed serves as platforms for the recruitment of TCR/CD3 nanodomains, and TCR/CD3 nanodomains are required for PKCθ signaling cascades and T-cell activation

Activation of human naïve Th cells increases surface expression of GD3 and induces neoexpression of GD2 that colocalize with TCR clusters

“…In T cells, the lipid rafts can isolate on membrane microdomains the GM1 ganglioside (lipid raft marker), the phosphatidylinositol 4,5-bisphosphate (PIP 2 ), GPI-linked proteins, Src family kinases and GTP binding proteins (Janes et al 2000). Disruption of lipid rafts by depleting cholesterol-using methyl-β-cyclodextrin (MβCD) affects TCR/CD3 clustering and cytokine secretion (Blank et al 2005;Zhu et al 2011;Zhong et al 2013). Our findings point to a potential and specific role of GD2 in TCR clustering or signaling and as a constitutive element of lipid rafts.…”

αβ and γδ T cell receptors: Similar but different

“…However, phosphoantigens induce the clustering and co-localization of the human V𝛾9V𝛿2 TCR and the lipid raft associated lipid GM1, 96 a feature that has been reported for the activated 𝛼𝛽 TCR as well. 97 Thus, activation induced changes in the lipid environment of the 𝛾𝛿 TCR can be assumed as well and might be involved in the regulation of ITAM accessibility.…”

Gene therapy approaches for GM1 gangliosidosis: Focus on animal and cellular studies

“…Following this accumulation of substrate, a variety of disease states with different symptoms occur in the patient. 37,38 According to the latest data collected until 2021, 51,52 Depending on the type of mutation, the symptoms of the disease will also be different, that is, the severity of the mutations is directly related to the age of onset of the disease. 53 For example, one study found that F I G U R E 1 Some important mechanisms in the pathogenesis of GM1 gangliosidosis (The CT scan image of the brain is taken from the study of Nicoli et al 18 ).…”

“…As mentioned earlier, in patients with GM1 gangliosidosis due to a mutation in the glb1 gene, enzyme activity is lost and the enzyme deficiency causes the accumulation of substrate in body tissues and increases urinary metabolites. Following this accumulation of substrate, a variety of disease states with different symptoms occur in the patient 37,38 . According to the latest data collected until 2021, Rha et al have classified the main mechanisms of GM1 disease into five important categories, which include the process of apoptosis through the UPR, neurotransmission, autophagy, neurotrophic factor activity, and neuroinflammation 39 .…”