Restoration of LAT signaling through the expression of a novel Adjunctive LAT-Activating CAR (ALA-CART) enhances the antigen-sensitivity and persistence of CAR T cells
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA991436 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR25133426_1.fastq.gz | Fastqsanger.gz | |||
| SRR25133426_2.fastq.gz | Fastqsanger.gz | |||
| SRR25133427_1.fastq.gz | Fastqsanger.gz | |||
| SRR25133427_2.fastq.gz | Fastqsanger.gz | |||
| SRR25133428_1.fastq.gz | Fastqsanger.gz |
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Project description:T cell antigen receptor (TCR) signaling depends upon the kinases Lck and Zap70. Lck phosphorylates the TCR, facilitating Zap70 recruitment to the stimulated TCR. Lck also phosphorylates Zap70, relieving its auto-inhibition and activating its catalytic domain. Zap70 then phosphorylates the critical adaptors LAT and SLP76 which serve to nucleate key effector molecules required for downstream responses. However, mechanisms facilitating the interaction of Zap70 with its substrates have not been described. We report an evolutionarily conserved proline-rich motif in LAT is important for Zap70-induced phosphorylation of LAT and downstream signaling. This LAT proline-rich motif associated with the Lck SH3 domain, thereby facilitating Zap70-mediated phosphorylation of LAT and downstream functions. Our results suggest Lck orchestrates multiple steps in TCR signaling including the newly described facilitation of the interaction of Zap70 with its substrate LAT. This previously unrecognized feature of TCR proximal signaling may contribute to the development of more immunomodulatory therapies.
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Project description:Cocaine and amphetamine-regulated transcript (CART) is expressed in pancreatic islet cells and neuronal elements. We have previously established insulinotropic actions of CART in human and rodent islets. The receptor for CART in the pancreatic beta cells is unidentified. We used RNA sequencing of Cartpt knock down (KD) INS-1 832/13 cells and identified GPR162 as the most Cartpt-regulated receptor. We therefore tested if GPR162 mediates the effects of CART in beta cells. Binding of CART to GPR162 was established using proximity ligation assay, radioactive binding and co-immunoprecipitation, and KD of Gpr162 mRNA caused reduced binding. Gpr162 KD cells had blunted CARTp-induced exocytosis, and reduced CARTp-induced insulin secretion. Furthermore, we identified a hitherto undescribed GPR162-dependent role of CART as a regulator of cytoskeletal arrangement. Thus, our findings provide mechanistic insight into the effect of CART on insulin secretion and show that GPR162 is the CART receptor in beta cells.
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