NFATc1 regulates LAG3+CD8+ T cells in the spleen of mice infected with Plasmodium yoelii NSM

TL;DR

This study shows that LAG3+CD8+ T cells in the spleen of malaria-infected mice become highly active, and their LAG3 expression is regulated by NFATc1.

Contribution

The study identifies NFATc1 as a key regulator of LAG3 expression in CD8+ T cells during malaria infection.

Findings

LAG3 expression is significantly upregulated in splenic CD8+ T cells after Plasmodium infection.

NFATc1 co-expresses with LAG3 and binds to its promoter to regulate expression.

Knockdown of NFATc1 reduces LAG3 expression in CD8+ T cells.

Abstract

Malaria, an infectious disease caused by Plasmodium, is primarily characterized by anemia and splenomegaly. CD8 ⁺ T cells are known to play a key role in anti-malaria immunity. Lymphocyte Activation Gene 3 (LAG3), a critical immune checkpoint molecule, is pivotal in CD8 ⁺ T cell-mediated anti-tumor responses. However, the role of LAG3 ⁺ CD8 ⁺ T cells in anti-malarial immunity and the regulatory factors governing LAG3 expression in CD8 ⁺ T cells remain unclear. In this study, C57BL/6 mice were subcutaneously infected with Plasmodium yoelii NSM. Splenic lymphocytes were isolated and analyzed using flow cytometry (FACs) and single-cell RNA sequencing (scRNA-seq). Results showed a significant upregulation of LAG3 expression in splenic CD8 ⁺ T cells post-infection. These LAG3 ⁺ CD8 ⁺ T cells displayed enhanced activation, responsiveness, proliferative capacity, and cytokine production. Additionally, activated nuclear factor of activated T cells 1 (NFATc1) was found to co-express with LAG3 in splenic CD8 ⁺ T cells from infected mice. Dual-fluorescence reporter gene assays in 293T cells identified NFATc1 as a key transcription factor that binds to the LAG3 promoter sequence. Knockdown of NFATc1 via small interfering RNA (siRNA) reduced LAG3 expression. In conclusion, our findings suggest that splenic LAG3 ⁺ CD8 ⁺ T cells in Plasmodium yoelii NSM-infected C57BL/6 mice display enhanced functionality and imply that NFATc1 could positively regulate LAG3 expression.

The authors observed that in the spleens of mice infected with malaria parasites, the amount of Lag3 molecules on CD8 + T cells (a type of immune cell) increased significantly. What’s more, these CD8 + T cells that had Lag3 molecules were highly active. They could multiply well and were good at releasing many different kinds of cytokines. The authors believe Lag3 is probably a molecule that only starts to appear after T cells are activated.At the same time, the authors also found that NFATc1 might be an important factor that helps trigger the production of Lag3 molecules. They learned this by doing experiments at the cellular and molecular levels, such as single-cell RNA sequencing and flow cytometry (common lab techniques). When they used a method called RNA interference to lower the amount of NFATc1, they found that the production of Lag3 molecules was also reduced. The authors think NFATc1 could become a potential “target gene” meaning in the future, it might be used to stop Lag3 molecules from forming, which could help treat diseases.

Figures