Papillary thyroid cancer (PTC) is the most common thyroid malignancy, with a strong propensity for cervical lymph node metastasis (CLNM). CLNM is known to increase the risk of locoregional recurrence and decrease the survival in some high-risk groups. Hence, there is a need for a reliable biomarker for the prediction of LNM in thyroid cancer. In the present study, both the mRNA and protein levels of MKL1 were significantly increased in PTC patients with lymph node metastasis (LNM) compared with those without LNM. Further receiver operating characteristic (ROC) analysis showed that MKL1 had a diagnostic value in the differentiation of LNM in PTC (AUC=0.87, P < 0.001). Meanwhile, Kaplan-Meier analysis revealed that high MKL1 expression was associated with a significant worse survival in PTC. Additionally, the biological function of MKL1 in PTC cells was explored. Our study indicated that MKL1 promoted the migration and invasiveness of PTC cells and a requirement for MKL1 in TGF-β/Smad3/MMP2 signal transduction. MKL1 interacted and recruited Smad3 to the promoter of MMP2 to activate MMP2 transcription upon TGF-β, a prominent pro-malignancy cytokine. Moreover, there were significant co-expression correlations between TGF-β, MKL1 and MMP2 in our clinical cohort of PTC specimens. Our results suggest that the detection of MKL1 expression could be used to predict cervical LNM and inform postoperative follow-up in papillary thyroid cancer.