Fbx4 is a specific substrate recognition component of SCF ubiquitin ligases that catalyzes the ubiquitination and subsequent degradation of cyclin D1 and Trxl. Two isoforms of human Fbx4 protein, the full length Fbx4 alpha and the C-terminal truncated Fbx4 beta have been identified, but their functions remain elusive. In this study, we demonstrated that the mRNA level of Fbx4 was significantly lower in hepatocellular carcinoma tissues than that in the corresponding non-tumor tissues. More importantly, we identified three novel splicing variants of Fbx4: Fbx4 gamma (missing 168-245nt of exonl), Fbx4 delta (missing exon6) and a N-terminal reading frame shift variant (missing exon2). Using cloning sequencing and RT-PCR, we demonstrated these novel splice variants are much more abundant in human cancer tissues and cell lines than that in normal tissues. When expressed in Sk-Hepl and NIH3T3 cell lines, Fbx4 beta, Fbx4 gamma and Fbx4 delta could promote cell proliferation and migration in vitro. Concordantly, these isoforms could disrupt cyclin D1 degradation and therefore increase cyclin D1 expression. Moreover, unlike the full-length isoform Fbx4 alpha, that mainly exists in cytoplasm, Fbx4 beta, Fbx4 gamma, and Fbx4 delta locate in both cytoplasm and nucleus. Since cyclin D1 degradation takes place in cytoplasm, the nuclear distribution of these Fbx4 isoforms may not be involved in the down-regulation of cytoplasmic cyclin Dl. These results define the impact of alternative splicing on Fbx4 function, and suggest that the attenuated cyclin D1 degradation by these novel Fbx4 isoforms provides a new insight for aberrant cyclin D1 expression in human cancers. (C) 2014 The Authors. Published by Elsevier Inc.