Regulation of pancreas development by hedgehog signaling

Pancreas organogenesis is regulated by the interaction of distinct signaling pathways that promote or restrict morphogenesis and cell differentiation. Previous work has shown that activin, a TGF(beta+) signaling molecule, permits pancreas development by repressing expression of Sonic hedgehog (Shh), a member of the hedgehog family of signaling molecules that antagonize pancreas development. Here we show that Indian hedgehog (Ihh), another hedgehog family member, and Patched 1 (Ptc1), a receptor and negative regulator of hedgehog activity, are expressed in pancreatic tissue. Targeted inactivation of Ihh in mice allows ectopic branching of ventral pancreatic tissue resulting in an annulus that encircles the duodenum, a phenotype frequently observed in humans suffering from a rare disorder known as annular pancreas. Shh(-)(/)(-) and Shh(-)(/)(-) Ihh(+/)(-) mutants have a threefold increase in pancreas mass, and a fourfold increase in pancreatic endocrine cell numbers. In contrast, mutations in Ptc1 reduce pancreas gene expression and impair glucose homeostasis. Thus, islet cell, pancreatic mass and pancreatic morphogenesis are regulated by hedgehog signaling molecules expressed within and adjacent to the embryonic pancreas. Defects in hedgehog signaling may lead to congenital pancreatic malformations and glucose intolerance.     

Cerebellar proteoglycans regulate sonic hedgehog responses during development

Sonic hedgehog promotes proliferation of developing cerebellar granule cells. As sonic hedgehog is expressed in the cerebellum throughout life it is not clear why proliferation occurs only in the early postnatal period and only in the external granule cell layer. We asked whether heparan sulfate proteoglycans might regulate sonic hedgehog-induced proliferation and thereby contribute to the specialized proliferative environment of the external granule cell layer. We identified a conserved sequence within sonic hedgehog that is essential for binding to heparan sulfate proteoglycans, but not for binding to the receptor patched. Sonic hedgehog interactions with heparan sulfate proteoglycans promote maximal proliferation of postnatal day 6 granule cells. By contrast, proliferation of less mature granule cells is not affected by sonic hedgehog-proteoglycan interactions. The importance of proteoglycans for proliferation increases during development in parallel with increasing expression of the glycosyltransferase genes, exostosin 1 and exostosin 2. These data suggest that heparan sulfate proteoglycans, synthesized by exostosins, may be critical determinants of granule cell proliferation.     

Hedgehog signaling in pancreas development

Hedgehog proteins are secreted molecules that bind to their cell surface receptors to elicit concentration dependent responses essential for numerous tissue patterning and cell differentiation events during embryogenesis. However, during early stages of pancreas organogenesis, hedgehog signaling has been shown to inhibit tissue morphogenesis and cell differentiation. By contrast, recent cell culture studies indicate that an active hedgehog pathway might be required for maintenance of adult endocrine cell functions. This review describes our current understanding of the requirement of hedgehog signaling during pancreas morphogenesis and cell differentiation and discusses how individual hedgehog genes might act at various stages to ensure proper pancreas development and organ function.     

Synthesis and evaluation of novel dimethylpyridazine derivatives as hedgehog signaling pathway inhibitors

We report herein the design and synthesis of a series of structural modified dimethylpyridazine compounds as novel hedgehog signaling pathway inhibitors. The bicyclic phthalazine core and 4-methylamino-piperidine moiety of Taladegib were replaced with dimethylpyridazine and different azacycle building blocks, respectively. The in vitro Gli-luciferase assay results demonstrate that the new scaffold still retained potent inhibitory potency. Piperidin-4-amine moiety was found to be the best linker between pharmacophores dimethylpyridazine and fluorine substituted benzoyl group. Furthermore, the optimization of 1-methyl-1H-pyrazol and 4-fluoro-2-(trifluoromethyl)benzamide by different aliphatic or aromatic rings were also investigated and the SAR were described. Several new derivatives were found to show potent Hh signaling inhibitory activity with nanomolar IC₅₀ values. Among these compounds, compound 11c showed the highest inhibitory potency with an IC₅₀ value of 2.33?nM, which was comparable to the lead compound Taladegib. In vivo efficacy of 11c in a ptch^+/?p53^?/? mouse medulloblastoma allograft model also indicated encouraging results.     

Suppressors of hedgehog signaling

Subversion of signals that physiologically suppress Hedgehog pathway results in aberrant neural progenitor development and medulloblastoma, a malignancy of the cerebellum. The Hedgehog antagonist RENKCTD11 maps to chromosome 17p13.2 and is involved in the withdrawal of the Hedgehog signaling at the granule cell progenitor transition from the outer to the inner external germinal layers, thus promoting growth arrest and differentiation. Deletion of chromosome 17p, the most frequent genetic lesion observed in this tumor, is responsible for the loss of function of RENKCTD11, resulting in upregulated Hedgehog signaling and medulloblastoma. Persistence of signals that limit Hedgehog activity is also associated with malignancy. Hedgehog signaling- induced downregulation of ErbB4 receptor expression is attenuated in medulloblastoma subsets in which the extent of Hedgehog pathway activity is limited, thus favoring the accumulation of ErbB4 with imbalanced alternative splice CYT-1 isoform over the CYT-2. This is responsible for both Neuregulin ligand-induced CYT-1-dependent prosurvival activity and loss of CYT-2-mediated growth arrest.     

Wnt5 signaling in vertebrate pancreas development

Background  

Signaling by the Wnt family of secreted glycoproteins through their receptors, the frizzled (Fz) family of seven-pass transmembrane proteins, is critical for numerous cell fate and tissue polarity decisions during development.     

Unique functions of Sonic hedgehog signaling during external genitalia development.

Coordinated growth and differentiation of external genitalia generates a proximodistally elongated structure suitable for copulation and efficient fertilization. The differentiation of external genitalia incorporates a unique process, i.e. the formation of the urethral plate and the urethral tube. Despite significant progress in molecular embryology, few attempts have been made to elucidate the molecular developmental processes for external genitalia. The sonic hedgehog (Shh) gene and its signaling genes have been found to be dynamically expressed during murine external genitalia development. Functional analysis by organ culture revealed that Shh could regulate mesenchymally expressed genes, patched 1 (Ptch1), bone morphogenetic protein 4 (Bmp4), Hoxd13 and fibroblast growth factor 10 (Fgf10), in the anlage: the genital tubercle (GT). Activities of Shh for both GT outgrowth and differentiation were also demonstrated. Shh(-/-) mice displayed complete GT agenesis, which is compatible with such observations. Furthermore, the regulation of apoptosis during GT formation was revealed for the first time. Increased cell death and reduced cell proliferation of the Shh(-/-) mice GT were shown. A search for alterations of Shh downstream gene expression identified a dramatic shift of Bmp4 gene expression from the mesenchyme to the epithelium of the Shh mutant before GT outgrowth. Regulation of mesenchymal Fgf10 gene expression by the epithelial Shh was indicated during late GT development. These results suggest a dual mode of Shh function, first by the regulation of initiating GT outgrowth, and second, by subsequent GT differentiation.     

Sonic hedgehog signaling from the urethral epithelium controls external genital development

External genital development begins with formation of paired genital swellings, which develop into the genital tubercle. Proximodistal outgrowth and axial patterning of the genital tubercle are coordinated to give rise to the penis or clitoris. The genital tubercle consists of lateral plate mesoderm, surface ectoderm, and endodermal urethral epithelium derived from the urogenital sinus. We have investigated the molecular control of external genital development in the mouse embryo. Previous work has shown that the genital tubercle has polarizing activity, but the precise location of this activity within the tubercle is unknown. We reasoned that if the tubercle itself is patterned by a specialized signaling region, then polarizing activity may be restricted to a subset of cells. Transplantation of urethral epithelium, but not genital mesenchyme, to chick limbs results in mirror-image duplication of the digits. Moreover, when grafted to chick limbs, the urethral plate orchestrates morphogenetic movements normally associated with external genital development. Signaling activity is therefore restricted to urethral plate cells. Before and during normal genital tubercle outgrowth, urethral plate epithelium expresses Sonic hedgehog (Shh). In mice with a targeted deletion of Shh, external genitalia are absent. Genital swellings are initiated, but outgrowth is not maintained. In the absence of Shh signaling, Fgf8, Bmp2, Bmp4, Fgf10, and Wnt5a are downregulated, and apoptosis is enhanced in the genitalia. These results identify the urethral epithelium as a signaling center of the genital tubercle, and demonstrate that Shh from the urethral epithelium is required for outgrowth, patterning, and cell survival in the developing external genitalia.     

Conserved cAMP signaling cascades regulate fungal development and virulence

Two well characterized signal transduction cascades regulating fungal development and virulence are the MAP kinase and cAMP signaling cascades. Here we review the current state of knowledge on cAMP signaling cascades in fungi. While the processes regulated by cAMP signaling in fungi are as diverse as the fungi themselves, the components involved in signal transduction are remarkably conserved. Fungal cAMP signaling cascades are also quite versatile, which is apparent from the differential regulation of similar biological processes. In this review we compare and contrast cAMP signaling pathways that regulate development in the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe, and differentiation and virulence in the human pathogen Cryptococcus neoformans and the plant pathogen Ustilago maydis. We also present examples of interaction between the cAMP and MAP kinase signaling cascades in the regulation of fungal development and virulence.     

Apaf1 apoptotic function critically limits Sonic hedgehog signaling during craniofacial development

Apaf1 is an evolutionarily conserved component of the apoptosome. In mammals, the apoptosome assembles when cytochrome c is released from mitochondria, binding Apaf1 in an ATP-dependent manner and activating caspase 9 to execute apoptosis. Here we identify and characterize a novel mouse mutant, yautja, and find it results from a leucine-to-proline substitution in the winged-helix domain of Apaf1. We show that this allele of Apaf1 is unique, as the yautja mutant Apaf1 protein is stable, yet does not possess apoptotic function in cell culture or in vivo assays. Mutant embryos die perinatally with defects in craniofacial and nervous system development, as well as reduced levels of apoptosis. We further investigated the defects in craniofacial development in the yautja mutation and found altered Sonic hedgehog (Shh) signaling between the prechordal plate and the frontonasal ectoderm, leading to increased mesenchymal proliferation in the face and delayed or absent ossification of the skull base. Taken together, our data highlight the time-sensitive link between Shh signaling and the regulation of apoptosis function in craniofacial development to sculpt the face. We propose that decreased apoptosis in the developing nervous system allows Shh-producing cells to persist and direct a lateral outgrowth of the upper jaw, resulting in the craniofacial defects we see. Finally, the novel yautja Apaf1 allele offers the first in vivo understanding of a stable Apaf1 protein that lacks a function, which should make a useful tool with which to explore the regulation of programmed cell death in mammals.     

Modulators of the hedgehog signaling pathway

Since its discovery by C. Nüsslein-Volhard and E. F. Wieschaus, hedgehog (hh) signaling has come a long way. Today it is regarded as a key regulator in embryogenesis where it governs processes like cell proliferation, differentiation, and tissue patterning. Furthermore, in adults it is involved in the maintenance of stem cells, and in tissue repair and regeneration. But hh signaling has a second—much darker—face: it plays an important role in several types of human cancers where it promotes growth and enables proliferation of tumor stem cells. The etiology of medulloblastoma and basal cell carcinoma is tightly linked to aberrant hh activity, but also cancers of the prostate, the pancreas, the colon, the breasts, rhabdomyosarcoma, and leukemia, are dependent on irregular hh activity. Recent clinical studies have shown that hh signaling can be the basis of an important new class of therapeutic agents with far-reaching implications in oncology. Thus, modulation of hh signaling by means of small molecules has emerged as a valuable tool in combating these hh-dependent cancers. Cyclopamine, a unique natural product with a fascinating history, was the first identified inhibitor of hh signaling and its story is closely linked to the progress in the whole field. In this review we will trace the story of cyclopamine, give an overview on the biological modes of hh signaling both in untransformed and malignant cells, and finally present potent modulators of the hh pathway—many of them already in clinical studies. For more than 30 years now the knowledge on hh signaling has grown steadily—an end to this development is far from being conceivable.