Cyclin L2 (CCNL2) is a novel member of the cyclin gene family. In a previous study, we demonstrated that CCNL2 expression was upregulated in ventricular septum tissues from patients with ventricular septal defect compared to healthy controls. In the present study, we established a stable CCNL2-overexpressing P19 cell line that can differentiate to myocardial cells when treated with 1% dimethyl sulfoxicle (DMSO). Our data showed that stable CCNL2-overexpressing P19 cells were less differentiated after treatment with 1% DMSO and that expression of myocardial cell differentiation-related genes (such as cardiac actin, GATA4, Mef2C, Nkx2.5, and BNP) were reduced compared to vector-only transfected P19. Moreover, PI 9 cells overexpressing the CCNL2 gene had a reduced growth rate and a remarkably decreased S phase. We also found that these cells underwent apoptosis, as detected by two different apoptosis assays. The antiapoptotic Bcl-2 protein was also downregulated in these cells. In addition, real-time PCR analysis revealed that expression of Wnt and beta-catenin was suppressed and GSK3 beta was induced in the CCNL2-overexpressing P19 cells. These data suggest that overexpression of CCNL2 inhibited proliferation and differentiation of mouse embryonic carcinoma PI 9 cells and induced them to undergo apoptosis, possibly through the Wnt signal transduction pathway. (C) 2009 Elsevier Inc. All rights reserved.