Colorectal cancer (CRC) is the third most common cancer worldwide. Despite the enormous amount of effort in the diagnosis and treatment of CRC, the overall survival rate of patients remains low. The precise molecular and cellular basis underlying CRC has not been completely understood yet. Over time, new genes and molecular pathways involved in the pathogenesis of the disease are being identified. The accurate discovery of these genes and signaling pathways are important and urgent missions for the next generation of anticancer therapy research. Chaperone DnaJ, also known as Hsp40 (heat shock protein 40), has been of particular interest in CRC pathogenesis, as it is involved in the fundamental cell activities for maintaining cellular homeostasis. Evidence shows that protein family members of DnaJ/Hsp40 play both roles, enhancing and reducing the growth of CRC cells. In the present review, we focus on the current knowledge of the molecular mechanisms responsible for DnaJ/Hsp40 in CRC carcinogenesis and biology.