Biomedical subjects

Philip W Ingham

Publications and source records attributed to Philip W Ingham.

14 recordsLinked to original sources

PubMed2006

A transgenic zebrafish model of neutrophilic inflammation.

We have established an in vivo model for genetic analysis of the inflammatory response by generating a transgenic zebrafish line that expresses GFP under the neutrophil-specific myeloperoxidase promoter. We show that inflammation is induced after transection of the tail of zebrafish larvae and that this inflammation subsequently resolves over a similar time course to mammalian systems. Quantitative data can be generated from this model by counting of fluorescent cells or by digital image analysis. In addition, we show that the resolution of experimentally induced inflammation can be inhibited by the addition of a pancaspase inhibitor, zVD.fmk, demonstrating that experimental manipulation of the resolution of inflammation is possible in this model.

Animals↗

PubMed2006

Orchestrating ontogenesis: variations on a theme by sonic hedgehog.

Embryonic development is an emergent process in which increasing complexity is generated by sequential cellular interactions. Recently, it has become clear that such interactions are mediated by just a few families of signalling molecules; but how does this limited repertoire elicit the diversity of form that is characteristic of multicellular organisms? Here we review the various ways in which a member of one such family, the sonic hedgehog (SHH) protein, is deployed during embryonic development. These examples of SHH function provide paradigms for inductive interactions that should help to inform attempts to recapitulate cellular programming and organogenesis in vitro.

Animals↗

PubMed2005

Retinoic acid signaling restricts the cardiac progenitor pool.

Organogenesis begins with specification of a progenitor cell population, the size of which provides a foundation for the organ's final dimensions. Here, we present a new mechanism for regulating the number of progenitor cells by limiting their density within a competent region. We demonstrate that retinoic acid signaling restricts cardiac specification in the zebrafish embryo. Reduction of retinoic acid signaling causes formation of an excess of cardiomyocytes, via fate transformations that increase cardiac progenitor density within a multipotential zone. Thus, retinoic acid signaling creates a balance between cardiac and noncardiac identities, thereby refining the dimensions of the cardiac progenitor pool.

Aldehyde Oxidoreductases↗

PubMed2005

A homologue of the Drosophila kinesin-like protein Costal2 regulates Hedgehog signal transduction in the vertebrate embryo.

Orthologues of nearly all of the core components of the Hedgehog signalling pathway, defined originally through genetic analysis in Drosophila, have now been discovered in vertebrates and shown to have highly conserved functions. The one striking exception to this rule is the kinesin-like protein Costal2, which plays a central role in controlling the activity of the zinc-finger-containing transcriptional regulator, Cubitus interruptus that modulates all Hedgehog-dependent target gene expression, but whose involvement in Hedgehog signalling has not been demonstrated in vertebrates. We report the cloning of a kinesin-related gene from the zebrafish that in structure as well as function, appears to represent the first vertebrate orthologue of costal2. Using a combination of genetic and biochemical analysis, we provide evidence that as in Drosophila, zebrafish Costal2 acts principally as an intracellular repressor of signal transduction, in conjunction with Suppressor of Fused, another protein that negatively regulates signalling in Hedgehog-responsive cells.

Amino Acid Sequence↗

PubMed2004

iguana encodes a novel zinc-finger protein with coiled-coil domains essential for Hedgehog signal transduction in the zebrafish embryo.

Signaling by lipid-modified secreted glycoproteins of the Hedgehog family play fundamental roles during pattern formation in animal development and in humans; dysfunction of Hedgehog pathway components is frequently associated with a variety of congenital abnormalities and cancer. Transcriptional regulation of Hedgehog target genes is mediated by members of the Gli zinc-finger transcription factors. The relative nuclear concentrations of Gli activator (Gli(act)) and repressor (Gli(rep)) forms, together with their nucleocytoplasmic trafficking, appear to be critical determinants for target gene expression. Whereas such stringent controls of Gli activity are critical in ensuring appropriate levels of pathway activation, the mechanisms by which these processes are regulated remain inadequately understood. Here, using genetic analysis, we show that the zebrafish iguana gene product acts downstream of the Smoothened protein to modulate Gli activity in the somites of the developing embryo. Positional cloning reveals that iguana encodes the zebrafish ortholog of Dzip1, a novel zinc-finger/coiled-coil domain protein that we show can shuttle between the cytoplasm and nucleus in a manner correlated with Hedgehog pathway activity.

Animals↗

PubMed2004

Drosophila melanogaster as a model host for Staphylococcus aureus infection.

Staphylococcus aureus is an important pathogen of humans, causing a range of superficial and potentially life-threatening diseases. Infection of the fruit fly Drosophila melanogaster with S. aureus results in systemic infection followed by death. Screening of defined S. aureus mutants for components important in pathogenesis identified perR and pheP, with fly death up to threefold slower after infection with the respective mutants compared to the wild-type. Infection of D. melanogaster with reporter gene fusion strains demonstrated the in vivo expression levels of the accessory gene regulator, agr, alpha-toxin, hla, and a manganese transporter, mntA. The use of the green fluorescent protein as a reporter under the control of the agr promoter (P3) showed S. aureus microcolony formation in vivo. The disease model also allowed the effect of antibiotic treatment on the flies to be determined. D. melanogaster is a genetically tractable model host for high-throughput analysis of S. aureus virulence determinants.

Amino Acid Transport Systems, Neutral↗

PubMed2003

The B-cell maturation factor Blimp-1 specifies vertebrate slow-twitch muscle fiber identity in response to Hedgehog signaling.

Vertebrate skeletal muscles comprise distinct fiber types that differ in their morphology, contractile function, mitochondrial content and metabolic properties. Recent studies identified the transcriptional coactivator PGC-1alpha as a key mediator of the physiological stimuli that modulate fiber-type plasticity in postembryonic development. Although myoblasts become fated to differentiate into distinct kinds of fibers early in development, the identities of regulatory proteins that determine embryonic fiber-type specification are still obscure. Here we show that the gene u-boot (ubo), a mutation in which disrupts the induction of embryonic slow-twitch fibers, encodes the zebrafish homolog of Blimp-1, a SET domain-containing transcription factor that promotes the terminal differentiation of B lymphocytes in mammals. Expression of ubo is induced by Hedgehog (Hh) signaling in prospective slow muscle precursors, and its activity alone is sufficient to direct slow-twitch fiber-specific development by naive myoblasts. Our data provide the first molecular insight into the mechanism by which a specific group of muscle precursors is driven along a distinct pathway of fiber-type differentiation in response to positional cues in the vertebrate embryo.

Animals↗

PubMed2003

Regulation of stem cell maintenance and transit amplifying cell proliferation by tgf-beta signaling in Drosophila spermatogenesis.

The continuous and steady supply of transient cell types such as skin, blood and gut depends crucially on the controlled proliferation of stem cells and their transit amplifying progeny. Although it is thought that signaling to and from support cells might play a key role in these processes, few signals that might mediate this interaction have been identified. During spermatogenesis in Drosophila, the asymmetric division of each germ line stem cell results in its self-renewal and the production of a committed progenitor that undergoes four mitotic divisions before differentiating while remaining in intimate contact with somatic support cells [1]. Previous data have suggested that TGF-beta signaling pathway components punt and schnurri are required in the somatic support cells to restrict germ cell proliferation. Here, by contrast, we show that the maintenance and proliferation of germ line stem cells and their progeny depends upon their ability to transduce the activity of a somatically expressed TGF-beta ligand, the BMP5/8 ortholog Glass Bottom Boat. We further demonstrate that TGF-beta signaling represses the expression of the Bam protein, which is both necessary and sufficient for germ cell differentiation, thereby maintaining germ line stem cells and spermatogonia in their proliferative state.

Animals↗

PubMed2003

Multiple muscle cell identities induced by distinct levels and timing of hedgehog activity in the zebrafish embryo.

BACKGROUND: In the zebrafish embryo, the differentiation of distinct muscle fiber types has been shown to require the activity of members of the Hedgehog (Hh) family of secreted proteins. Evidence from other systems suggests that Hh behaves as a morphogen, inducing cell fates in a concentration-dependent manner. Exactly how Hh signaling contributes to the generation of the correct pattern of cells within the zebrafish myotome, however, has remained obscure. RESULTS: Here, we distinguish four distinct myotomal cell identities in the zebrafish embryo on the basis of their position, morphology, and gene expression patterns. Using morpholino oligonucleotides (MOs) to diminish the activities of the Hh pathway components Patched (Ptc), Fused (Fu), and Suppressor of Fused (Su(fu)), and the teratogen cyclopamine to inhibit the Hh transducer Smoothened (Smo), we show that the appropriate differentiation of each cell type depends upon the levels and range of Hh signaling within the myotome. In addition, by transiently modulating Hh activity by using cyclopamine and a heat-inducible transgene, we demonstrate that the competence of myotomal cells to respond to Hh changes with time. Finally, we show that the Gli1 and Gli2 transcription factors mediate most of the response of myotomal cells to Hh. CONCLUSIONS: Hh signaling acts in a dosage-dependent manner to specify cell fate in the zebrafish myotome. Allocation of the correct number of cells to a specific fate depends upon the range of Hh activity. The timing of exposure to Hh determines the response of cells to the signal.

Animals↗

PubMed2003

Developmental roles and clinical significance of hedgehog signaling.

Cell signaling plays a key role in the development of all multicellular organisms. Numerous studies have established the importance of Hedgehog signaling in a wide variety of regulatory functions during the development of vertebrate and invertebrate organisms. Several reviews have discussed the signaling components in this pathway, their various interactions, and some of the general principles that govern Hedgehog signaling mechanisms. This review focuses on the developing systems themselves, providing a comprehensive survey of the role of Hedgehog signaling in each of these. We also discuss the increasing significance of Hedgehog signaling in the clinical setting.

Animals↗

PubMed2003

The transformation of the model organism: a decade of developmental genetics.

The past decade has seen the development of powerful techniques to dissect the molecular processes that regulate development. New tools have been used to reveal the basis of cell polarity, morphogen gradients and regulation of signaling in developing animals. Cell biology and developmental biology have become closely intertwined, and many genes that had been thought of as regulators of general cell biological (housekeeping) functions have been shown to act as specific developmental regulators. Vertebrate developmental genetics is now flourishing, with forward and reverse genetics in both zebrafish and the mouse providing new dimensions to our understanding of development.

Animals↗

PubMed2003

Hedgehog signaling in the Drosophila eye and head: an analysis of the effects of different patched trans-heterozygotes.

Characterization of different alleles of the Hedgehog receptor patched (ptc) indicates that they can be grouped into several classes. Most mutations result in complete loss of Ptc function. However, missense mutations located within the putative sterol-sensing domain (SSD) or C terminus of ptc encode antimorphic proteins that are unable to repress Smo activity and inhibit wild-type Ptc from doing so, but retain the ability to bind and sequester Hh. Analysis of the eye and head phenotypes of Drosophila melanogaster in various ptc/ptc(tuf1) heteroallelic combinations shows that these two classes of ptc allele can be easily distinguished by their eye phenotype, but not by their head phenotype. Adult eye size is inversely correlated with head vertex size, suggesting an alteration of cell fate within the eye-antennal disc. A balance between excess cell division and cell death in the mutant eye discs may also contribute to final eye size. In addition, contrary to results reported recently, the role of Hh signaling in the Drosophila head vertex appears to be primarily in patterning rather than in proliferation, with Ptc and Smo having opposing effects on formation of medial structures.

Alleles↗

PubMed2002

Hedgehogs tryst with the cell cycle.

Hedgehog proteins play an essential role during pattern formation in animal development and, increasingly, much of our appreciation of their modes of action is emanating from studies of their signalling mechanisms at the cellular level. Recent work has provided insights into how Hedgehog controls the cell cycle in a variety of circumstances. The data suggest that this influence may be direct and operates through interaction of the signalling pathway with cell cycle regulators at multiple points within the cell cycle. These new findings have profound implications in the context of clinical conditions - especially cancers - that arise from de-regulated cell proliferation in response to aberrant Hedgehog signalling activity.

Animals↗

PubMed2002

Notch and Wingless modulate the response of cells to Hedgehog signalling in the Drosophila wing.

During Drosophila wing development, Hedgehog (Hh) signalling is required to pattern the imaginal disc epithelium along the anterior-posterior (AP) axis. The Notch (N) and Wingless (Wg) signalling pathways organise the dorsal-ventral (DV) axis, including patterning along the presumptive wing margin. Here, we describe a functional hierarchy of these signalling pathways that highlights the importance of competing influences of Hh, N, and Wg in establishing gene expression domains. Investigation of the modulation of Hh target gene expression along the DV axis of the wing disc revealed that collier/knot (col/kn), patched (ptc), and decapentaplegic (dpp) are repressed at the DV boundary by N signalling. Attenuation of Hh signalling activity caused by loss of fused function results in a striking down-regulation of col, ptc, and engrailed (en) symmetrically about the DV boundary. We show that this down-regulation depends on activity of the canonical Wg signalling pathway. We propose that modulation of the response of cells to Hh along the future proximodistal (PD) axis is necessary for generation of the correctly patterned three-dimensional adult wing. Our findings suggest a paradigm of repression of the Hh response by N and/or Wnt signalling that may be applicable to signal integration in vertebrate appendages.

Animals↗