Dietary fiber (DF) is considered an interventional diet beneficial for human health. High DF intake effectively reduces the incidence of three major chronic diseases, type 2 diabetes (T2DM), cardiovascular disease (CVD), and colorectal cancer (CRC). The health benefits of DF are closely related to their physicochemical properties with major positive roles in human digestion and intestinal health. However, mechanisms linking DF with diseases remain unclear. The development of genomics, metabolomics, and immunology, and the powerful combination of animal models and clinical trials, have facilitated a better understanding of the relationships between DF and diseases. Accumulating evidence suggests that the physical existence of DF and DF-microbiota interaction are the key parameters controlling the action mechanisms of DF in chronic diseases. Therefore, this review discusses the potential mechanism of DF modulating T2DM, CVD, and CRC, therefore providing a theoretical basis for more effective use of DF to intervene in chronic diseases.