The ubiquitin ligase CUL4A has been implicated in tumorigenesis but its contributions to progression and metastasis have not been evaluated. Here we show that CUL4A is elevated in breast cancer as well as ovarian, gastric and colorectal tumors where its expression level correlates positively with distant metastasis. CUL4A overexpression in normal or malignant human mammary epithelial cells increased their neoplastic properties in vitro and in vivo, markedly increasing epithelial-mesenchymal transition (EMT) and the metastatic capacity of malignant cells. In contrast, silencing CUL4A in aggressive breast cancer cells inhibited these processes. Mechanistically, we found CUL4A modulated histone H3K4me3 at the promoter of the EMT regulatory gene ZEB1 in a manner associated with its transcription. ZEB1 silencing blocked CUL4A-driven proliferation, EMT, tumorigenesis and metastasis. Further, in human breast cancers ZEB1 expression correlated positively with CUL4A expression and distant metastasis. Taken together, our findings reveal a pivotal role of CUL4A in regulating the metastatic behavior of breast cancer cells.

cell cycle ͉ proliferation ͉ Wnt ͉ corepressor T he process of skin carcinogenesis involves a series of transitional events, ranging from hyperplasia, dysplasia, and papilloma to invasive squamous cell carcinoma. The current model indicates that the progression of benign lesions to malignancy depends on the cell type targeted by these mutations (1, 2). Indeed, recent evidence has indicated that the bulge region of the hair follicles contains selfrenewing, slow-cycling stem cells, which give rise to transient amplifying cells and several differentiated cell lineages in the interfollicular epidermis (3, 4). The progression of skin carcinogenesis can be modified by mutations involving Tgf␤1, Smad3, ubiquitin ligase Fbxw7/Cdc4, or Pten (5-9), suggesting that a genetic network synergizes with activated Ras mutation to promote the initiation or malignant transformation of skin tumors (10). Another important regulator of skin carcinogenesis is cyclin D1. Overexpression of cyclin D1 increases the propensity of skin carcinogenesis, whereas loss of cyclin D1 reduces tumor formation (11,12). One mechanism to control cyclin D1 expression is through transcriptional regulation via the Wnt/␤-catenin signaling pathway (13,14), which consists of a core set of highly evolutionarily conserved proteins that have wide-ranging effects on gene expression affecting proliferation, migration, pluripotency, morphogenesis, and tumorigenesis (15-17). In the epidermis, targeted deletion of ␤-catenin during embryogenesis results in severe abnormalities in cell fate determination and maintenance of hair follicle formation (18). In contrast, mutations that activate ␤-catenin function lead to skin tumorigenesis in both humans and mice (19, 20).Homeodomain interacting protein kinase (HIPK) contains three distinct members that regulate apoptosis, cell growth, and proliferation. HIPK2 has been identified as a transcriptional corepressor for homeodomain proteins NK3 and Brn3a (21,22). Previous results have indicated that Brn3a and HIPK2 regulate a delicate balance of gene expression that controls programmed cell death in sensory and dopamine neurons (22)(23)(24). Both HIPK1 and HIPK2 can interact with human p53 and, under DNA damage conditions, activate p53 by promoting phosphorylation and acetylation (25)(26)(27). Contrary to the predicted function, HIPK1-deficient mice show a higher resistance to the development of DMBA-induced skin tumors (27). Recent evidence, however, has demonstrated a functional redundancy between HIPK1 and HIPK2 during development, which could compensate for the loss of HIPK1 (28). Indeed, HIPK2 has a wide range of functions, including promoting the degradation of CtBP in response to UV-induced DNA damage (29) and regulating cell proliferation through the Wnt signaling pathway (30). Furthermore, HIPK2 can interact with several proteins containing the high-mobility group I (HMGI), a domain highly conserved in transcription factors in the lymphoid enhancer-binding factor 1/T cell factor (LEF1-TCF) family (31).In this work, we ...