Sine oculis homeobox homolog 1 (SIX1) has been supposed to be correlated with the metastasis and poor prognosis of several malignancies. However, the effect of SIX1 on the metastatic phenotype of tumor cells and the underlying mechanisms were still unclear to date. Here we report that SIX1 can promote alpha 5 beta 1-mediated metastatic capability of cervical cancer cells. SIX1 promoted the expression of alpha 5 beta 1 integrin to enhance the adhesion capacity of tumor cells in vitro and tumor cell arrest in circulation in vivo. Moreover, higher expression of SIX1 in tumor cells resulted in the increased production of active MMP-2 and MMP-9, up-regulation of anti-apoptotic genes (BCL-XL and BCL2) and down-regulation of pro-apoptotic genes (BIM and BAX), thus promoting the invasive migration and anoikis-resistance of tumor cells. Importantly, blocking 0601 abrogated the regulatory effect of SIX1 on the expression of these genes, and also abolished the promotional effect of SIX1 on invasive capability of tumor cells. Furthermore, knockdown of alpha 5 could abolish the promoting effect of SIX1 on the development of metastatic lesions in both experimental and spontaneous metastasis model. Therefore, by up-regulating alpha 5 beta 1 expression, SIX1 not only promoted the adhesion capacity, but also augmented ECM-alpha 5 beta 1-mediated regulation of gene expression to enhance the metastatic potential of cervical cancer cells. These results suggest that SIX1/alpha 5 beta 1 might be considered as valuable marker for metastatic potential of cervical cancer cells, or a therapeutic target in cervical cancer treatment. (C) 2014 Elsevier Inc. All rights reserved.