Lymph node metastasis is the main factor of poor prognosis in oral squamous cell carcinoma (OSCC) patients. Using proteomic analysis, we found that interferon-stimulated gene 15 (ISG15) was the highest expressed protein related to lymph node metastasis. Abundant ISG15 was also observed in the microarray datasets and immunohistochemical of OSCC patients. In the orthotopic xenograft model, ISG15 knockdown decreased tumor lymphangiogenesis and lymph node metastasis. Similarly, ISG15 knockdown diminished cell migration, invasion and transendothelial migration in OSCC cells. Especially, ISG15-induced cell migration was in an intracellular ISGylation-independent manner and associated with membrane protrusions. Ectopic expression of ISG15 increased Rac1 activity and knockdown of Rac1 impaired ISG15-enhanced migration. Moreover, ISG15 was co-localized and interacted with Rac1 in the region of membrane protrusions by immunofluorescence and proximity ligation asays. Immunoprecipitated ISG15 interacted with Rac1, especially Rac1-GDP form. Our study indicated that ISG15 promotes cell migration and lymph node metastasis via regulation of Rac1 activity and physical interaction with Rac1.
