Dbl-family guanine nucleotide exchange factors (GEFs) are important activators of Rho GTPases, which are significantly associated with tumorigenesis and metastasis. The catalytic ability of the Dbl-family GEFs to activate Rho GTPases depends on their Dbl-homology domain followed by a pleckstrin-homology domain. In the present study, we showed that C9orf100, a new member of the Dbl-family GEFs with a minimal catalytic unit, may contribute to hepatocellular carcinoma (HCC). Quantitative real-time PCR results demonstrated that C9orf100 was highly and widely upregulated in 42/44 (95.5%, >2‑fold) HCC specimens compared with adjacent non-cancerous livers, and this upregulation was correlated with intrahepatic metastasis and α-fetoprotein levels of HCC. Furthermore, the ectopic expression of C9orf100 promoted cell proliferation and colony formation in Huh-7 and YY-8103 cells, whereas silencing of C9orf100 resulted in the suppression of cell growth in MHCC-97H and PLC/PRF/5 cells. Flow cytometry confirmed this effect on MHCC-97H cell growth and indicated that C9orf100 may function in the G2/M phase. In addition, we showed that C9orf100 is involved in the positive regulation of HCC cell migration by a transwell chamber analysis. Our findings suggest that C9orf100 plays a potential oncogenic role in the development and metastasis of HCC.