The significance of Th17 cells and interleukin (IL)-17A signaling in host defense and disease development has been demonstrated in various infection and autoimmune models. In addition, the generation of Th17 cells is highly influenced by microbes. However, the specific bacterial components capable of shaping Th17 responses have not been well defined. The goals of this study are to understand how a bacterial toxin, cholera toxin (CT), modulates Th17-dominated response in isolated human CD4+ T cells, and what are the mechanisms associated with this modulation. The CD4+ cells isolated from human peripheral blood are treated with CT. The levels of cytokine production and specific T helper cell responses are determined by ELISA, Luminex assay, and flow cytometry. Along with the decreased production of other pro-inflammatory cytokines (IFN-γ, TNF-α, and IL-2), we found that CT could directly enhance the IL-17A production through a cAMP-dependent pathway. This enhancement is specific for IL-17A but not for IL-17F, IL-22 and CCL20. Interestingly, CT could increase IL-17A production only from Th17-committed cells, such as CCR6+ CD4+ T cells and in-vitro-differentiated Th17 cells. Furthermore, we also demonstrated that this direct effect occurs at a transcriptional level since CT stimulates the reporter activity in Jurkat and primary CD4+ T cells transfected with the IL-17A promoter-reporter construct. This is the first report to show that CT has the capacity to directly shape Th17 responses in the absence of antigen-presenting cells. Our findings highlight the potentials of bacterial toxins in the regulation of human Th17 responses.