Project description:Epstein-Barr virus (EBV) ubiquitously infects global population and leads to a variety malignancies and autoimmune diseases. It establishes its lytic-latency life cycle relying on the tropism transition between B cell and epithelial cell, during which its critical glycoprotein gB and gHgL act as the fusion apparatus mediating viral recognition and membrane fusion in either cell type. Thus. simultaneous targeting to fusion apparatus would be ideal strategy to construct potent EBV prophylactic vaccine. In this study, we designed chimeric nanoparticle multivalently displaying gB and gHgL with even gp42 upon the nanoparticle surface, inducing significantly robust neutralizing antibody generation in both murine and non-human primate models. As we further identify the comparable immunization potency between chimeric nanoparticle and whole live virion and preponderance of gB in neutralizing antibody elicitation, we used single-B cell sequencing to dissect the B cell response to chimeric nanoparticle immunization in mouse and isolate a gB-specific neutralizing antibody Fab5 targeting novel vulnerable site. These findings offers insight to advanced vaccine design for EBV and other herpesviruses.
Project description:ChIP-seq analysis was performed in a Jurkat cell line to analyze DNA bindings of TAL1-FKBP12 protein using an anti-HA antibody after DMSO or dTAG-13 treatment.
Project description:The effect of CD151 expression onto the kinome of Jurkat T cells was assessed using kinome analysis. CD151 was expressed in Jurkat T cells by retroviral transduction based on a pMSCV vector. Entrez Gene: 977 UniProtKB: P48509 Jurkat T cells were transduced with the MSCV-CD151 vector and successfully transduced cells were selected using puromycin. For the kinome array experiments 3 independent samples of Jurkat cells and three independent samples of J-CD151 cells were collected. To minimize unspecific background signals, lysates from Jurkat and J-CD151 T cells harvested at different growth stages, which were then pooled to provide one sample prior to loading on the Kinexus antibody microarrays.
Project description:We have performed quantitative phosphoproteomic analysis on jurkat cells. Phosphorylation change was compared between jurkat cells incubated on ice and those incubated in 37 degree water bath. And the combination of cold induced and OKT3/4 antibody induced signaling was compared.
Project description:SLAMF6 is a homotypic receptor of the Ig-superfamily associated with progenitor exhausted T cells. In humans, SLAMF6 has three splice isoforms involving its V-domain.We KO isoform 1 of SLAMF6 in a Jurkat cell line and harvest this cell line RNA using Bulk-RNA-Seq after activation in 5 time points. Comparing the results to WT Jurkat cell line, we analyzed the transcriptional landscape and showed that after activation SLAMF6 isoform 3 is the prominent expressed isoform, and that the KO Jurkat cells have a stronger cytotoxic phenotype.
Project description:By performing 10x 3’ scRNA-seq on Jurkat cells post TCR acivation in CRISPR/Cas9 screening, we want to investigate the compatibility of the A/G mixed capture sequence on multiple single cell RNA-seq platforms. By mimicking the adenylated endogenous mRNA, gRNA transcripts could be directly captured by poly(dT) primer during the reverse transcription, and serve as perturbation index in high identification rate.
Project description:Extracellular vesicles (EVs) mediate cell-cell communication including the intercellular transfer of miRNAs, which alter gene expression in target cells. Previously, we have shown that CD47 is also present on EVs and alters their effects on target cells, suggesting that specific surface markers define functionally distinct EVs. This hypothesis will be addressed by comparing total cellular versus EVs miRNA contents between Jurkat and JinB8 (CD47 deficient) T cells. The Jurkat T cell line (E6.1) was purchased from ATCC. The cells were maintained using RPMI 1640 containing 10% FBS, glutamine, penicillin, and streptomycin (Gibco). The Jurkat and JinB8 T cells used for experiments were maintained less than 6 weeks in culture. The cultured Jurkat were washed with IXPBs followed by a second wash with HITES medium (DMEM/F12 supplemented with, bovine serum albumin, hydrocortisone, insulin, transferrin, and trace elements). EVs were isolated from Jurkat T cells were cultured overnight using HITES medium. The conditioned media from Jurkat T cells were collected, and cell debris were removed using centrifugation at 300 g for 5 minutes and 2500 g for 10 minutes respectively. Centrifuged media were concentrated using Ultra-4 centrifugal filters to about 1 ml volume and further centrifuged at 10,000 g for 10 minutes. EVs were isolated using an Exo-Quick kit (SBI). miRNaseazy (Qiagen) kits were used to extract total RNA from Jurkat and JinB8 T cells and their EVs according to the manufacturer's instructions.