Epithelial to mesenchymal transition (EMT) is implicated in the progression of primary tumours towards metastasis and is likely caused by a pathological activation of transcription factors regulating EMT in embryonic development. To analyse EMT-causing pathways in tumourigenesis, we identified transcriptional targets of the E-cadherin repressor ZEB1 in invasive human cancer cells. We show that ZEB1 repressed multiple key determinants of epithelial differentiation and cell–cell adhesion, including the cell polarity genes Crumbs3, HUGL2 and Pals1-associated tight junction protein. ZEB1 associated with their endogenous promoters in vivo, and strongly repressed promotor activities in reporter assays. ZEB1 downregulation in undifferentiated cancer cells by RNA interference was sufficient to upregulate expression of these cell polarity genes on the RNA and protein level, to re-establish epithelial features and to impair cell motility in vitro. In human colorectal cancer, ZEB1 expression was limited to the tumour–host interface and was accompanied by loss of intercellular adhesion and tumour cell invasion. In invasive ductal and lobular breast cancer, upregulation of ZEB1 was stringently coupled to cancer cell dedifferentiation. Our data show that ZEB1 represents a key player in pathologic EMTs associated with tumour progression.

Macrophages are critical for granuloma formation, but the cell-intrinsic mechanisms remain unknown. Weichhart and colleagues demonstrate that chronic mTOR activity leads to macrophage-dependent granuloma formation, which may have relevance to sarcoidosis. The aggregation of hypertrophic macrophages constitutes the basis of all granulomatous diseases, such as tuberculosis or sarcoidosis, and is decisive for disease pathogenesis. However, macrophage-intrinsic pathways driving granuloma initiation and maintenance remain elusive. We found that activation of the metabolic checkpoint kinase mTORC1 in macrophages by deletion of the gene encoding tuberous sclerosis 2 (Tsc2) was sufficient to induce hypertrophy and proliferation, resulting in excessive granuloma formation in vivo. TSC2-deficient macrophages formed mTORC1-dependent granulomatous structures in vitro and showed constitutive proliferation that was mediated by the neo-expression of cyclin-dependent kinase 4 (CDK4). Moreover, mTORC1 promoted metabolic reprogramming via CDK4 toward increased glycolysis while simultaneously inhibiting NF-κB signaling and apoptosis. Inhibition of mTORC1 induced apoptosis and completely resolved granulomas in myeloid TSC2-deficient mice. In human sarcoidosis patients, mTORC1 activation, macrophage proliferation and glycolysis were identified as hallmarks that correlated with clinical disease progression. Collectively, TSC2 maintains macrophage quiescence and prevents mTORC1-dependent granulomatous disease with clinical implications for sarcoidosis.

Joseph Gleeson and colleagues report that mutations in TMEM216 cause Joubert, Meckel and related syndromes. They further show that TMEM216 localizes to the base of cilia and that its loss leads to defects in ciliogenesis and centrosome docking. Joubert syndrome (JBTS), related disorders (JSRDs) and Meckel syndrome (MKS) are ciliopathies. We now report that MKS2 and CORS2 (JBTS2) loci are allelic and caused by mutations in TMEM216, which encodes an uncharacterized tetraspan transmembrane protein. Individuals with CORS2 frequently had nephronophthisis and polydactyly, and two affected individuals conformed to the oro-facio-digital type VI phenotype, whereas skeletal dysplasia was common in fetuses affected by MKS. A single G218T mutation (R73L in the protein) was identified in all cases of Ashkenazi Jewish descent (n = 10). TMEM216 localized to the base of primary cilia, and loss of TMEM216 in mutant fibroblasts or after knockdown caused defective ciliogenesis and centrosomal docking, with concomitant hyperactivation of RhoA and Dishevelled. TMEM216 formed a complex with Meckelin, which is encoded by a gene also mutated in JSRDs and MKS. Disruption of tmem216 expression in zebrafish caused gastrulation defects similar to those in other ciliary morphants. These data implicate a new family of proteins in the ciliopathies and further support allelism between ciliopathy disorders.

The Raf kinases play a key role in relaying signals elicited by mitogens or oncogenes. Here, we report that c‐raf‐1^−/− embryos are growth retarded and die at midgestation with anomalies in the placenta and in the fetal liver. Although hepatoblast proliferation does not appear to be impaired, c‐raf‐1^−/− fetal livers are hypocellular and contain numerous apoptotic cells. Similarly, the poor proliferation of Raf‐1^−/− fibroblasts and hematopoietic cells cultivated in vitro is due to an increase in the apoptotic index of these cultures rather than to a cell cycle defect. Furthermore, Raf‐1‐ deficient fibroblasts are more sensitive than wild‐ type cells to specific apoptotic stimuli, such as actinomycin D or Fas activation, but not to tumor necrosis factor‐α. MEK/ERK activation is normal in Raf‐1‐deficient cells and embryos, and is probably mediated by B‐Raf. These results indicate that the essential function of Raf‐1 is to counteract apoptosis rather than to promote proliferation, and that effectors distinct from the MEK/ERK cascade must mediate the anti‐apoptotic function of Raf‐1.

Polarized hepatocytes expressing hyperactive Ha-Ras adopt an invasive and metastatic phenotype in cooperation with transforming growth factor (TGF)-β. This dramatic increase in malignancy is displayed by an epithelial to mesenchymal transition (EMT), which mimics the TGF-β-mediated progression of human hepatocellular carcinomas. In culture, hepatocellular EMT occurs highly synchronously, facilitating the analysis of molecular events underlying the various stages of this process. Here, we show that in response to TGF-β, phosphorylated Smads rapidly translocated into the nucleus and activated transcription of target genes such as E-cadherin repressors of the Snail superfamily, causing loss of cell adhesion. Within the TGF-β superfamily of cytokines, TGF-β1, -β2 and -β3 were specific for the induction of hepatocellular EMT. Expression profiling of EMT kinetics revealed 78 up- and 235 downregulated genes, which preferentially modulate metabolic activities, extracellular matrix composition, transcriptional activities and cell survival. Independent of the genetic background, platelet-derived growth factor (PDGF)-A ligand and both PDGF receptor subunits were highly elevated, together with autocrine secretion of bioactive PDGF. Interference with PDGF signalling by employing hepatocytes expressing the dominant-negative PDGF-α receptor revealed decreased TGF-β-induced migration in vitro and efficient suppression of tumour growth in vivo. In conclusion, these results provide evidence for a crucial role of PDGF in TGF-β-mediated tumour progression of hepatocytes and suggest PDGF as a target for therapeutic intervention in liver cancer.