Aim To study the role of mutations of Mfn2 protein kinase A phosphorylation site in the proliferation of vascular smooth muscel cell(VSMC) and related signaling pathways. Methods Two novel mutations were constructed,Adv-Mfn2-αPKA and Adv-Mfn2-asnPKA,then VSMC were infected with them.The effect of mutations on the proliferation of VSMC was explored by WST-1 assay.The cell cycle was determined using flow cytometry.Western blotting were used to detect the expression of Mfn2 and p-ERK1/2. Results The expression of Mfn2 protein has no significant difference in Adv-Mfn2,Adv-Mfn2-alaPKA and Adv-Mfn2-asnPKA groups.Mfn2-alaPKA and Mfn2 both showed stronger inhibitory effect on VSMC proliferation(P<0.01).Most of the cells in these two groups were blocked in the stage of G0/G1(P<0.01),and the expression levels of p-ERK1/2 also decrease(P<0.01).Mfn2-alaPKA is superior to Mfn2 in attenuating the proliferation of VSMCs.The effect of Mfn2-asnPKA is similar to the control. Conclusion Mfn2-alaPKA has more stronger inhibitory effect on the proliferation of VSMC than Mfn2,while Mfn2-asnPKA has no effect.PKA phosphorylation site plays an important role in regulating the function of Mfn2 gene.