Objective To investigate the effect of c-Jun N-terminal kinase 1 (JNK1) or JNK2 overexpression on the proliferation and apoptosis of SCL-1 human cutaneous squamous cell carcinoma cell lines. Methods JNK1 and JNK2 were separately combined with pHBAD-EF1-MCS-3FLAG-CMV-GFP vector to construct the recombinant adenovirus expression vectors of JNK1 and JNK2. The recombinant vectors were used to infect SCL-1 cells. After the optimization of the infection conditions, the fold changes of over-expression were identified by real-time quantitative PCR (qRT-PCR) and Western blot analysis. CCK-8 assay was performed to determine the proliferative activity of SCL-1 cells. Cell colony forming ability was evaluated by cell colony formation assay. Wound healing assay was used to detect the scratch healing rate of SCL-1 cells. Cell apoptosis was determined by flow cytometry. The qRT-PCR was used to detect the mRNA levels of JNK1, JNK2 and c-Jun. The protein level of phosphorylated c-Jun was tested by Western blot analysis. Results The optimal infection condition of JNK1- and JNK2-overexpressing adenoviral vectors was multiplicity of infection (MOI) of 100. The expression levels of JNK1 and JNK2 in SCL-1 cells significantly increased 48 hours after the infection. Compared with the control group, the over-expression of JNK1 had no significant effect on the proliferation and anti-apoptosis ability of SCL-1 cells. The proliferation and anti-apoptosis ability of Ad-JNK2 was significantly enhanced compared with Ad-JNK1, and the phosphorylated c-Jun protein level was up-regulated. Conclusion JNK2 has the function of enhancing the proliferation and anti-apoptotic ability of SCL-1 human cutaneous squamous cell carcinoma cells.