Background & Aims Pathogenesis of cirrhosis, a disabling outcome of defective liver repair, involves deregulated accumulation of myofibroblasts derived from quiescent hepatic stellate cells (HSC), but the mechanisms that control HSC transdifferentiation are poorly understood. We investigated whether the Hedgehog (Hh) pathway controls HSC fate by regulating metabolism. Methods Microarray, quantitative PCR, and immunoblot analyses were used to identify metabolic genes that were differentially expressed in quiescent vs myofibroblast HSC. Glycolysis and lactate production were disrupted in HSC to determine if metabolism influenced transdifferentiation. Hh signaling and hypoxia-inducible factor (HIF)1α activity were altered to identify factors that alter glycolytic activity. Changes in expression of genes that regulate glycolysis were quantified and localized in biopsy samples from patients with cirrhosis, and liver samples from mice following administration of CCl4 or bile-duct ligation. Mice were given systemic inhibitors of Hh to determine if they affect glycolytic activity of the hepatic stroma; Hh signaling was also conditionally disrupted in myofibroblasts to determine the effects of glycolytic activity. Results Transdifferentiation of cultured, quiescent HSC into myofibroblasts induced glycolysis and caused lactate accumulation. Increased expression of genes that regulate glycolysis required Hh signaling and involved induction of HIF1α. Inhibitors of Hh signaling, HIF1α, glycolysis, or lactate accumulation converted myofibroblasts to quiescent HSC. In diseased livers of animals and patients, numbers of glycolytic stromal cells were associated with the severity of fibrosis. Conditional disruption of Hh signaling in myofibroblasts reduced numbers of glycolytic myofibroblasts and liver fibrosis in mice; similar effects were observed following administration of pharmacologic inhibitors of Hh. Conclusions Hedgehog signaling controls HSC fate by regulating metabolism. These findings might be applied to diagnosis and treatment of cirrhosis.

A widespread decrease of mature microRNAs is often observed in human malignancies giving them potential to act as tumor suppressors. Thus, microRNAs may be potential targets for cancer therapy. The global miRNA deregulation is often the result of defects in the miRNA biogenesis pathway, such as genomic mutation or aberrant expression/localization of enzymes and cofactors responsible of miRNA maturation. Alterations in the miRNA biogenesis machinery impact on the establishment and development of cancer programs. Accumulation of pri-microRNAs and corresponding depletion of mature microRNAs occurs in human cancers compared to normal tissues, strongly indicating an impairment of crucial steps in microRNA biogenesis. In agreement, inhibition of microRNA biogenesis, by depletion of Dicer1 and Drosha, tends to enhance tumorigenesis in vivo. The p53 tumor suppressor gene, TP53, is mutated in half of human tumors resulting in an oncogene with Gain-Of-Function activities. In this review we discuss recent studies that have underlined a role of mutant p53 (mutp53) on the global regulation of miRNA biogenesis in cancer. In particular we describe how a new transcriptionally independent function of mutant p53 in miRNA maturation, through a mechanism by which this oncogene is able to interfere with the Drosha processing machinery, generally inhibits miRNA processing in cancer and consequently impacts on carcinogenesis.

Preservation of the epithelial state involves the stable repression of epithelial-to-mesenchymal transition program, whereas maintenance of the stem compartment requires the inhibition of differentiation processes. A simple and direct molecular minicircuitry between master elements of these biological processes might provide the best device to keep balanced such complex phenomena. In this work, we show that in hepatic stem cell Snail, a transcriptional repressor of the hepatocyte differentiation master gene HNF4a, directly represses the expression of the epithelial microRNAs (miRs)-200c and -34a, which in turn target several stem cell genes. Notably, in differentiated hepatocytes HNF4a, previously identified as a transcriptional repressor of Cellular differentiation implies an orchestrated sequence of events guiding stem cells/precursors toward specialized cell types based on the contemporary and strictly correlated phenomena of loss of stemness and acquisition of histotypic markers and functions. The homeostasis of the stem cell compartment requires mechanisms actively counteracting differentiation; 1 similarly, the maintenance of the differentiated state involves a stable repression of elements capable to induce morphological transition and dedifferentiation. 2 The observation that a number of stem cells are restricted to a specific differentiation fate suggests that elements pivotal for the coordinated execution of the opposite processes could be tissue-specific. Considering that stem cell compartments are rare and give rise to a heterogeneous cellular population capable to reversibly shift among different states, 3 the availability of a stable stem cell line executing specific differentiation programs discloses an unique possibility to investigate mechanisms regulating alternative cellular choices. A simple and direct molecular mini-circuitry of master elements of mutually exclusive biological processes, also able to reciprocally influence their own expression, may provide the theoretically best device to trigger such complex phenomena.We previously characterized a number of stable liver stem cell lines named RLSCs (from resident liver stem cells) that spontaneously acquire an epithelial morphology and differentiate into hepatocytes (named RLSCdH from RLSC-derived hepatocytes). Notably, RLSCs were also proved to recapitulate the hepatocyte post-differentiation patterning defined as 'zonation': their spontaneous differentiation, in fact, generates periportal hepatocytes that may be induced to switch into perivenular hepatocytes by means of the convergence of Wnt signaling on the HNF4a-driven transcription. 4 Furthermore, we identified a simple cross-regulatory circuitry between HNF4a (master regulator of hepatocyte differentiation) and Snail (master regulator of the epithelial-to-mesenchymal transition, EMT), whose expression is mutually exclusive because of their direct reciprocal transcriptional repression. 2,5 These findings, relevant for the comprehension of the EMT and of the reverse process mesenchymal-...

Downregulation of microRNAs (miRNAs) is commonly observed in cancers and promotes tumorigenesis suggesting that miRNAs may function as tumor suppressors. However, the mechanism through which miRNAs are regulated in cancer, and the connection between oncogenes and miRNA biogenesis remain poorly understood. The TP53 tumor-suppressor gene is mutated in half of human cancers resulting in an oncogene with gain-of-function activities. Here we demonstrate that mutant p53 (mutp53) oncoproteins modulate the biogenesis of a subset of miRNAs in cancer cells inhibiting their post-transcriptional maturation. Interestingly, among these miRNAs several are also downregulated in human tumors. By confocal, co-immunoprecipitation and RNA-chromatin immunoprecipitation experiments, we show that endogenous mutp53 binds and sequesters RNA helicases p72/82 from the microprocessor complex, interfering with Drosha-pri-miRNAs association. In agreement with this, the overexpression of p72 leads to an increase of mature miRNAs levels. Moreover, functional experiments demonstrate the oncosuppressive role of mutp53-dependent miRNAs (miR-517a, -519a, -218, -105). Our study highlights a previously undescribed mechanism by which mutp53 interferes with Drosha-p72/82 association leading, at least in part, to miRNA deregulation observed in cancer.