TAZ (WWTR1), identified as a 14-3-3 binding protein with a PDZ binding motif, modulates mesenchymal stem cell differentiation. We now show that TAZ plays a critical role in the migration, invasion, and tumorigenesis of breast cancer cells. TAZ is conspicuously expressed in human breast cancer cell lines in which its expression levels generally correlate with the invasiveness of cancer cells. Overexpression of TAZ in lowexpressing MCF10A cells causes morphologic changes characteristic of cell transformation and promotes cell migration and invasion. Conversely, RNA interference-mediated knockdown of TAZ expression in MCF7 and Hs578T cells reduces cell migration and invasion. TAZ knockdown in MCF7 cells also retards anchorage-independent growth in soft agar and tumorigenesis in nude mice. Significantly, TAZ is overexpressed in f20% of breast cancer samples. These results indicate that TAZ plays a role in the migration, invasion, and tumorigenesis of breast cancer cells and thus presents a novel target for the detection and treatment of breast cancer.

The TAZ transcription co-activator promotes cell proliferation and epithelial-mesenchymal transition. TAZ is inhibited by the Hippo tumor suppressor pathway, which promotes TAZ cytoplasmic localization by phosphorylation. We report here that TAZ protein stability is controlled by a phosphodegron recognized by the F-box protein ␤-TrCP and ubiquitylated by the SCF/CRL1 ␤-TrCP E3 ligase. The interaction between TAZ and ␤-TrCP is regulated by the Hippo pathway. Phosphorylation of a phosphodegron in TAZ by LATS primes it for further phosphorylation by CK1⑀ and subsequent binding by ␤-TrCP. Therefore, the Hippo pathway negatively regulates TAZ function by both limiting its nuclear accumulation and promoting its degradation. The phosphodegron-mediated TAZ degradation plays an important role in negatively regulating TAZ biological functions.

The Hippo pathway restricts the activity of transcriptional co-activators TAZ and YAP by phosphorylating them for cytoplasmic sequestration or degradation. In this report, we describe an independent mechanism for the cell to restrict the activity of TAZ and YAP through interaction with angiomotin (Amot) and angiomotin-like 1 (AmotL1). Amot and AmotL1 were robustly co-immunoprecipitated with FLAG-tagged TAZ, and their interaction is dependent on the WW domain of TAZ and the PPXY motif in the N terminus of Amot. Amot and AmotL1 also interact with YAP via the first WW domain of YAP. Overexpression of Amot and AmotL1 caused cytoplasmic retention of TAZ and suppressed its transcriptional outcome such as the expression of CTGF and Cyr61. Hippo refractory TAZ mutant (S89A) is also negatively regulated by Amot and AmotL1. HEK293 cells express the highest level of Amot and AmotL1 among nine cell lines examined, and silencing the expression of endogenous Amot increased the expression of CTGF and Cyr61 either at basal levels or upon overexpression of exogenous S89A. These results reveal a novel mechanism to restrict the activity of TAZ and YAP through physical interaction with Amot and AmotL1.

ObjectiveThe nature of the tumour-infiltrating leucocytes (TILs) is known to impact clinical outcome in carcinomas, including hepatocellular carcinoma (HCC). However, the role of tumour-infiltrating B cells (TIBs) remains controversial. Here, we investigate the impact of TIBs and their interaction with T cells on HCC patient prognosis.DesignTissue samples were obtained from 112 patients with HCC from Singapore, Hong Kong and Zurich and analysed using immunohistochemistry and immunofluorescence. RNA expression of CD19, CD8A, IFNG was analysed using quantitative PCR. The phenotype of freshly isolated TILs was analysed using flow cytometry. A mouse model depleted of mature B cells was used for functional study.ResultsTumour-infiltrating T cells and B cells were observed in close contact with each other and their densities are correlated with superior survival in patients with HCC. Furthermore, the density of TIBs was correlated with an enhanced expression of granzyme B and IFN-γ, as well as with reduced tumour viability defined by low expression of Ki-67, and an enhanced expression of activated caspase-3 on tumour cells. CD27 and CD40 costimulatory molecules and TILs expressing activation marker CD38 in the tumour were also correlated with patient survival. Mice depleted of mature B cells and transplanted with murine hepatoma cells showed reduced tumour control and decreased local T cell activation, further indicating the important role of B cells.ConclusionsThe close proximity of tumour-infiltrating T cells and B cells indicates a functional interaction between them that is linked to an enhanced local immune activation and contributes to better prognosis for patients with HCC.