In this study, a modified rat model similar to the classic human evolution of colorectal cancer (CRC) was established. As such, the altered profiles of proteins involved in these processes were further verified in human specimens, so as to determine the potential biomarkers relevant to human CRC development. Protein samples of four specific stages involved in CRC progression ((i) normal mucosa, (ii) adenoma, (iii) carcinoma, and (iv) liver metastasis)) were investigated by 2-DE. One protein spot displayed sequential suppression in the course of colorectal malignant transformation and was identified as transgelin by mass spectrometry. A decrease in its expression in both the epithelium and lamina propria was further confirmed by Western blot and immunohistochemistry analyses. Clinical and pathological parameter analysis revealed that downregulation of transgelin was associated with poor differentiation, and subsequent Dukes Stage and lower survival rate. Paradoxically, its sera level was significantly higher in CRC patients than in healthy donors, and the rise became dramatic, particularly in later Dukes Stages. These results indicate that downregulation of transgelin, in both the epithelium and lamina propria and accompanied with colorectal carcinogenesis, is correlated with worse prognosis. Its elevated serum levels might be the result of pathological hyperplasia of myofibroblasts and smooth muscle cells together with deeper tumor invasion into muscle layers. This altered expression represents interactions between cancer epithelium and stroma, such that transgelin might be a potential marker for CRC genesis and progression.