Introduction: T follicular helper (Tfh) cells are a specialized subset of T helper cells necessary for germinal center reaction, affinity maturation, and the differentiation of germinal center B cells to antibody-producing plasma and memory B cells. The differentiation of Tfh cells is a multistage, multifactorial process involving a variety of cytokines, surface molecules and transcription factors. While to date early signaling events for fate committed differentiation of Tfh cells are sufficiently studied, mechanisms allowing Tfh cells to maintain their commitment/programming are still unclear.
Methods: The regulation of IL-21 expression by STAT1 in Tfh cells was assessed by FACS staining, quantitative Real-Time PCR, Chromatin Immunoprecipitation (ChIP) assay, luciferase reporter assay and retroviral transduction. For these assays FACS-sorted CD4+CD44hiCXCR5hiPD1hi (Tfh) cells were utilized from wild-type and STAT1 deficient mice subjected to Ova immunization. Immunoblot analysis was performed to explore the mechanism whereby STAT1 along with STAT3 contributes to IL-21 production in Tfh cells.
Results: Here we report that STAT1 deficiency in CD4+ T cells leads to diminished number of Tfh cells at day 7 and 14 after antigen immunization, while their numbers were comparable between STAT1-sufficient and deficient mice at day 3, suggesting the role of STAT1 in Tfh lineage maturation and maintenance. Interestingly STAT1 deficiency in Tfh cells leads to significant drop in IL-21 expression, while the expression of Tfh-specific genes such as Bcl6, CXCR5 and c-Maf was not affected. Functionally, STAT1 in cooperation with STAT3 triggers IL-21 production in Tfh cells by directly binding to and activation of the IL-21 gene locus. In addition, STAT1/STAT3 cooperation influences the expression of transcriptional factor Batf as well as determines Batf-driven IL-21 expression in Tfh cells.
Conclusions: Our results thus indicate STAT1 dependent mechanism for IL-21 expression and Tfh lineage maturation and maintenance.