Polypyrimidine Tract-Binding Protein 1 (PTBP1) regulates CD4 T cell Activation independent of its role in proliferation

Abstract

Our previous work found that the RNA binding protein polypyrimidine tract-binding protein (PTBP1) is critical for regulating multiple events in T cell activation including changes in proliferation, and expression of activation markers and cytokines. These changes corresponded to the regulation of the ERK1/2 and NF-{kappa}B pathways as well as through changes in steady-state RNA levels. Because proliferation is critical for driving T cell activation, it was unclear whether PTBP1 was required for optimal activation per se or whether changes were secondary to a requirement for initiating/sustaining proliferation. To address this question, the human T cell lymphoma cell line, Jurkat, which recapitulates many of the molecular events of TCR-induced activation, was used to understand how PTBP1 impacts early events in T cell activation with ongoing proliferation. Using two phenotypically distinct Jurkat subclones (D1.1 and B2.7), we first profiled global RNA expression patterns using RNAseq analysis and found marked differences between the two cell lines with the D1.1 line giving a more antigen-experienced phenotype. Reducing PTBP1 by shPTB expression, to 60% WT levels resulted in no significant decrease in proliferation in the two subclones. However, we observed that PTBP1 was required for both optimal expression of activation markers, CD25, CD38, CD69, and CD40L, and signaling through the ERK1/2, P38 and AKT pathways. Importantly, limiting PTBP1 had different effects on the activation signals for each cell line suggesting that the differentiation state of the cell is a critical factor in understanding the role of PTBP1 in T cell activation. This was further reinforced by our finding that PTBP1 regulated distinct groups of genes specific for each line. Together, our findings suggest that PTBP1 regulates specific T cell activation responses independent of its role in proliferation and that the initial phenotype of the T cell plays an essential role in the dependency of the cell on PTBP1 for driving these changes.