Project description:We analyzed the total proteome of CD4+ T cells isolated from WT mice, and cultured to perform a CRISPR/CAS9 edition of their genome, in order to introduce an OST sequence at the C-terminus of proteins of interest (LAT or UBASH3A, n=6 biological replicates in each case). Control CD4+ T cells , isolated and cultured in the same way, but not modified by CRISPR/CAS9, were also analyzed (CT, n=6 biological replicates), as well as CD4+ T cells which have undergone a smaller number of expansion cycles than long term CD4+ T cells (WT, n=2 biological replicates). Each sample was analyzed once by single run naoLC-MS, resulting in 20 raw files.
Project description:Identifying Smad4-binding proteins in mouse CD4+ T cells under IL6SB condition. Mouse CD4+ T cells were isolated from mouse spleen and lymph nodes.
Project description:Identifying BTLA interacting proteins in mouse CD4+ effector T cells expressing BTLA at endogenous levels and after stimulation with pervanadate.
Project description:Omnivorous animals, including mice and humans, tend to prefer energy-dense nutrients rich in fat over plant-based diets, especially for short periods of time. The health consequences of this short-term consumption of energy-dense nutrients remain still unclear. We found that every short-term, reiterated switches to feast diets mimicking our social eating behavior, breached the potential buffering effect of the intestinal microbiota and deeply reorganized the immunological architecture of mucosa-associated lymphoid tissues. The first dietary switch was sufficient to induce transient mucosal immune depression and suppress systemic, antigen-specific immunity leading to higher susceptibility to Salmonella Typhimurium and Listeria monocytogenes infections. This was explained by a reduction of CD4+ T cell metabolic fitness and cytokine production due to impaired mTOR activity in response to withdrawal of microbial provision of fiber metabolites. Reintroducing dietary fiber efficiently rewired T cell metabolism and restored both mucosal and systemic CD4+ T cell functions and immunity. Finally, dietary intervention study in human volunteers confirmed the impact of short-term dietary switches on human CD4+ T cell functionality. This work reveals that short-term nutritional changes cause a drastic yet transient depression of both mucosal and systemic immunity, creating windows of opportunities for pathogenic infections.
Project description:CD4+ T cells were isolated from gene-targeted mouse expressing a One-Strep-tag (OST) at the C terminus of the CD5 protein, briefly expanded in vitro and we analyzed the interactome of their OST-tagged CD5 protein. Affinity purification of the OST tagged protein was performed using Streptactin beads, from T cells either non-stimulated, stimulated 30s, 120s or 300s with anti-CD3 and anti-CD4 antibodies, or 300s with pervanadate. Each AP-MS purification is associated with a corresponding control (purification from CD4+ T cells isolated from WT mice, cultured and stimulated in the same conditions). The number of replicate biological experiments was n=3 for all conditions (time-points), and each sample was analyzed once by single-run nanoLC-MS, resulting in 30 raw files.
Project description:We analyzed the total proteome of CD4+ T cells isolated from WT mice, either non stimulated or at 5 different time points of stimulation with anti-CD3 and anti-CD4 antibodies (30s; 120s; 300s; 600s)
Project description:To analyse the influence of pentanoate on the differentiation of naive CD4 T cells under Th17 inducing conditions, murine CD4 T cells were isolated from spleen and lymphnodes of FIR/TIGER mice (IL-10 (GFP) - Foxp3 (RFP) - Reporter mice). The T cells were kept for 3 days under Th17 inducing conditions either with 4 mM pentanoate or without pentanoate (control).