Phenotypic and transcriptional profile correlates with functional plasticity of antigen-specific CD4+ T cells

The role of CD4⁺ T cells in the control of infectious pathogens is highly complex with a myriad of functions but how these T cells acquire differential functional potentiality remains poorly defined. Here we show that human cytomegalovirus (CMV)-specific CD4⁺ T cells directed towards different viral antigens expressed predominantly TNF-α alone or TNF-α and IFN-γ. TNF-α⁺ and IFN-γ⁺ CD4⁺ T cells expressed significantly higher levels of T-box transcription factors T-bet with graded loss of Eomesodermin (Eomes) expression (T-bet^Hi Eomes^Hi/Lo) when compared with TNF-α⁺ CD4⁺ T cells expressing lower levels of both T-bet and Eomes (T-bet^- Eomes^-). Furthermore, TNF-α⁺ and IFN-γ⁺ CD4⁺ T cells expressed significantly higher levels of perforin and interleukin (IL)-2 and displayed a terminally differentiated phenotype (CCR7^- CD27 ^- CD45RA^- CD57⁺ CD62L^-). In contrast, TNF-α⁺ alone CMV-specific CD4⁺ T cells were predominantly early-memory phenotype with a proportion of these cells displaying T memory stem-cell phenotype (CD95⁺ CD45RA⁺ CCR7⁺ CD27⁺). In vitro stimulation of CMV-specific CD4⁺ T cells with viral antigen in the presence of IL-12 was sufficient to dramatically change the transcriptional and functional profile of TNF-α⁺ CD4⁺ T cells, whereas TNF-α⁺ and IFN-γ⁺ CD4⁺ T cells remained unaltered. These findings illustrate an intrinsic link between cytokine expression, transcriptional regulation and cellular differentiation, and their impact on functional plasticity of virus-specific CD4⁺ T cells.