Cysteine-rich 61 (Cyr61; CCN1) plays an important role in tumor development and progression in many kinds of human malignancies. Here, we further show the enforced expression of the Cyr61 gene or treatment with recombinant Cyr61 protein enhanced expression of chemokine receptors CXCR1 and CXCR2 in gastric cancer AGS cells. Attenuation of Cyr61 levels in MKN-45 cells by transfecting with antisense Cyr61 significantly reduced the level of CXCR1 and CXCR2. It is suggested that Cyr61 tightly regulates the downstream genes CXCR1 and CXCR2 in gastric cancer cells. Supportively, reverse transcription-PCR and immunohistochemical analysis of human gastric adenocarcinoma showed that there was a high correlation between the expression level of Cyr61 and CXCR1/CXCR2. The up-regulated functionality of CXCR1 andCXCR2 in Cyr61-overexpressing AGS cells could facilitate their chemotactic migration toward interleukin-8, a physiologic ligand of CXCR1 and CXCR2. In addition, the Cyr61-mediated up-regulation of CXCR1/CXCR2 also contributed to transendothelial migration, as well as intravasation in a chick embryo model. Pharmacologic and genetic approaches revealed that phosphoinositide 3-kinase (PI3K)/Akt, but not extracellular signal-regulated kinase 1/2 or p38, signaling pathway is requisite for the up-regulation of CXCR1/CXCR2 mRNA and protein induced by Cyr61. Function-neutralizing antibody to integrin alphavbeta3, but not alpha(2)beta(1), effectively abolished Cyr61-elicited Src activation and the subsequent PI3K/Akt pathway. Antagonists toward integrin alphavbeta3, Src kinase, and PI3K/Akt not only suppressed CXCR1/CXCR2 elevation but also blocked chemotactic migration induced by Cyr61. In conclusion, we suggest that Cyr61 promotes interleukin-8-dependent chemotaxis, transendothelial migration, and intravasation by induction of CXCR1/CXCR2 through integrin alphavbeta3/Src/PI3K/Akt-dependent pathway.