Hedgehog signaling in vertebrate eye development: a growing puzzle

The vertebrate retina has been widely used as a model to study the development of the central nervous system. Its accessibility and relatively simple organization allow analysis of basic mechanisms such as cell proliferation, differentiation and death. For this reason, it could represent an ideal place to solve the puzzle of Hh signaling during neural development. However, the extensive wealth of data, sometimes apparently discordant, has made the retina one of the most complicated models for studying the role of the Hh cascade. Given the complexity of the field, a deep analysis of the data arising from different animal models is essential. In this review, we will compare and discuss all reported roles of Hh signaling in eye development to shed light on its multiple functions.Received 26 September 2003; received after revision 13 November 2003; accepted 19 November 2003     

Hedgehog signaling in vertebrate and invertebrate limb patterning

Invertebrate and vertebrate limbs have very different anatomies and modes of development. Despite these differences, recent studies demonstrate that a significant overlap exists in the signals used to pattern invertebrate and vertebrate limbs. One of these signaling molecules is Hedgehog, a secreted protein that functions to coordinate growth and proliferation along the anterior-posterior axis of developing limbs. Recent studies indicate that the mechanism of action, regulation and function of Hedgehog signaling in Drosophila and vertebrate limb development are often quite similar, yet at other times are distinct. Here we highlight the similarities and differences between the use of Hedgehog signaling in these two systems.     

The roles of endogenous retinoid signaling in organ and appendage regeneration

The ability to regenerate injured or lost body parts has been an age-old ambition of medical science. In contrast to humans, teleost fish and urodele amphibians can regrow almost any part of the body with seeming effortlessness. Retinoic acid is a molecule that has long been associated with these impressive regenerative capacities. The discovery 30 years ago that addition of retinoic acid to regenerating amphibian limbs causes “super-regeneration” initiated investigations into the presumptive roles of retinoic acid in regeneration of appendages and other organs. However, the evidence favoring or dismissing a role for endogenous retinoids in regeneration processes remained sparse and ambiguous. Now, the availability of genetic tools to manipulate and visualize the retinoic acid signaling pathway has opened up new routes to dissect its roles in regeneration. Here, we review the current understanding on endogenous functions of retinoic acid in regeneration and discuss key questions to be addressed in future research.     

Structure and evolution of vertebrate aldehyde oxidases: from gene duplication to gene suppression

Aldehyde oxidases (AOXs) and xanthine dehydrogenases (XDHs) belong to the family of molybdo-flavoenzymes. Although AOXs are not identifiable in fungi, these enzymes are represented in certain protists and the majority of plants and vertebrates. The physiological functions and substrates of AOXs are unknown. Nevertheless, AOXs are major drug metabolizing enzymes, oxidizing a wide range of aromatic aldehydes and heterocyclic compounds of medical/toxicological importance. Using genome sequencing data, we predict the structures of AOX genes and pseudogenes, reconstructing their evolution. Fishes are the most primitive organisms with an AOX gene (AOXα), originating from the duplication of an ancestral XDH. Further evolution of fishes resulted in the duplication of AOXα into AOXβ and successive pseudogenization of AOXα. AOXβ is maintained in amphibians and it is the likely precursors of reptilian, avian, and mammalian AOX1. Amphibian AOXγ is a duplication of AOXβ and the likely ancestor of reptilian and avian AOX2, which, in turn, gave rise to mammalian AOX3L1. Subsequent gene duplications generated the two mammalian genes, AOX3 and AOX4. The evolution of mammalian AOX genes is dominated by pseudogenization and deletion events. Our analysis is relevant from a structural point of view, as it provides information on the residues characterizing the three domains of each mammalian AOX isoenzyme. We cloned the cDNAs encoding the AOX proteins of guinea pig and cynomolgus monkeys, two unique species as to the evolution of this enzyme family. We identify chimeric RNAs from the human AOX3 and AOX3L1 pseudogenes with potential to encode a novel microRNA.     

The roles of collagen genes in skeletal development and morphogenesis

Molecular genetic analyses of osteochondrodysplasias in mice and humans have recently led to the identification of mutations in genes encoding structural proteins growth factor receptors and sulphate transporters. Further analyses of such inherited disorders, using rapid techniques for gene mapping, positional cloning and mutation detection, will undoubtedly uncover other genes that are important for skeletal development. Together with studies of transgenic mice, in which specific genes that are expressed in the skeleton are mutated, these analyses will provide insight into genes that are essential for skeletal morphogenesis.     

Wnt signaling: multiple functions in neural development

Wnt signaling has proven to be essential for neural development at various stages and across species. Wnts are involved in morphogenesis and patterning, and their proliferation-promoting role is a key function in stem cell maintenance and the expansion of progenitor pools. Moreover, Wnt signaling is involved in differentiation processes and lineage decision events during both central and peripheral nervous system development. Additionally, several reports point to a role of Wnt signaling in axon guidance and neurite outgrowth. This article reviews and consolidates the existing evidence for the functions of Wnt signaling in neural development.Received 10 December 2004; received after revision 19 January 2005; accepted 21 January 2005     

ALK1 signaling in development and disease: new paradigms

Activin A receptor like type 1 (ALK1) is a transmembrane serine/threonine receptor kinase in the transforming growth factor-beta receptor family that is expressed on endothelial cells. Defects in ALK1 signaling cause the autosomal dominant vascular disorder, hereditary hemorrhagic telangiectasia (HHT), which is characterized by development of direct connections between arteries and veins, or arteriovenous malformations (AVMs). Although previous studies have implicated ALK1 in various aspects of sprouting angiogenesis, including tip/stalk cell selection, migration, and proliferation, recent work suggests an intriguing role for ALK1 in transducing a flow-based signal that governs directed endothelial cell migration within patent, perfused vessels. In this review, we present an updated view of the mechanism of ALK1 signaling, put forth a unified hypothesis to explain the cellular missteps that lead to AVMs associated with ALK1 deficiency, and discuss emerging roles for ALK1 signaling in diseases beyond HHT.     

Hedgehog signaling in pancreas development and disease

Since its discovery, numerous studies have shown that the Hedgehog (Hh) signaling pathway plays an instrumental role during diverse processes of cell differentiation and organ development. More recently, it has become evident that Hh signaling is not restricted to developmental events, but retains some of its activity during adult life. In mature tissues, Hh signaling has been implicated in the maintenance of stem cell niches in the brain, renewal of the gut epithelium and differentiation of hematopoietic cells. In addition to the basal function in adult tissue, deregulated signaling has been implicated in a variety of cancers, including basal cell carcinoma, glioma and small cell lung cancer. Here, we will focus on the role of Hh signaling in pancreas development and pancreatic diseases, including diabetes mellitus, chronic pancreatitis and pancreatic cancer. Received 5 August 2005; received after revision 4 November 2005; accepted 22 November 2005     

Purinergic signaling in embryonic and stem cell development

Nucleotides are of crucial importance as carriers of energy in all organisms. However, the concept that in addition to their intracellular roles, nucleotides act as extracellular ligands specifically on receptors of the plasma membrane took longer to be accepted. Purinergic signaling exerted by purines and pyrimidines, principally ATP and adenosine, occurs throughout embryologic development in a wide variety of organisms, including amphibians, birds, and mammals. Cellular signaling, mediated by ATP, is present in development at very early stages, e.g., gastrulation of Xenopus and germ layer definition of chick embryo cells. Purinergic receptor expression and functions have been studied in the development of many organs, including the heart, eye, skeletal muscle and the nervous system. In vitro studies with stem cells revealed that purinergic receptors are involved in the processes of proliferation, differentiation, and phenotype determination of differentiated cells. Thus, nucleotides are able to induce various intracellular signaling pathways via crosstalk with other bioactive molecules acting on growth factor and neurotransmitter receptors. Since normal development is disturbed by dysfunction of purinergic signaling in animal models, further studies are needed to elucidate the functions of purinoceptor subtypes in developmental processes.     

Histamine receptor-mediated signaling during development and brain function in adulthood

Histamine might have an important role in brain development. However, most studies have focused on short-term effects of histamine receptor-mediated signaling on brain function in adulthood. Little is known about the potential long-term effects of histamine receptor-mediated signaling during development on brain function in adulthood. We hypothesize that increased postsynaptic histamine receptor-mediated signaling during development has detrimental effects on brain function in adulthood. Our data support this hypothesis. In the developing mouse brain, histamine H3 receptor blockade, which increases histamine release, has detrimental sex-dependent effects on object recognition, spatial learning in the water maze, and pre-pulse inhibition in adulthood. Our data also support the hypothesis that histamine mediates the detrimental long-term sex-dependent effects of methamphetamine exposure early in life on these brain functions in adulthood. Therefore, increased efforts are warranted to carefully evaluate the effects of drugs that directly or indirectly affect histamine receptor-mediated signaling during development on cognitive function later in life.     

The STING pathway and regulation of innate immune signaling in response to DNA pathogens

The innate immune system has evolved a variety of sensing mechanisms to detect and counter microbial invasion. These include the Toll-like receptor (TLR), cytoplasmic, nucleotide binding oligomerization domain (NOD)-like receptor and RIG-I-like helicase (RLH) pathways. However, how the cell detects pathogen-associated DNA to trigger host defense, including the production of interferon, remains to be fully clarified. Understanding these processes could have profound implications into how we understand and treat a variety of microbial-related disease, including viral-associated cancers, as well as autoimmune disorders. Recently, an endoplasmic reticulum-associated molecule referred to as STING (for stimulator of interferon genes) was isolated and shown to be critical for regulating the production of IFN in response to cytoplasmic DNA. Here, we review recent discoveries relating to the detection of foreign DNA, including the importance of the STING and inflammasome pathways and the triggering of innate signaling processes.     

The role of bHLH genes in ear development and evolution: revisiting a 10-year-old hypothesis

In mouse ear development, two bHLH genes, Atoh1 and Neurog1, are essential for hair cell and sensory neuron differentiation. Evolution converted the original simple atonal-dependent neurosensory cell formation program of diploblasts into the derived developmental program of vertebrates that generates two neurosensory cell types, the sensory neuron and the sensory hair cell. This transformation was achieved through gene multiplication in ancestral triploblasts resulting in the expansion of the atonal bHLH gene family. Novel genes of the Neurogenin and NeuroD families are upregulated prior to the expression of Atoh1. Recent data suggest that NeuroD and Neurogenin were lost or their function in neuronal specification reduced in flies, thus changing our perception of the evolution of these genes. This sequence of expression changes was accompanied by modification of the E-box binding sites of these genes to regulate different downstream genes and to form inhibitory loops among each other, thus fine-tuning expression transitions.     

Pepsinogens, progastricsins, and prochymosins: structure, function, evolution, and development

Five types of zymogens of pepsins, gastric digestive proteinases, are known: pepsinogens A, B, and F, progastricsin, and prochymosin. The amino acid and/or nucleotide sequences of more than 50 pepsinogens other than pepsinogen B have been determined to date. Phylogenetic analyses based on these sequences indicate that progastricsin diverged first followed by prochymosin, and that pepsinogens A and F are most closely related. Tertiary structures, clarified by X-ray crystallography, are commonly bilobal with a large active-site cleft between the lobes. Two aspartates in the center of the cleft, Asp32 and Asp215, function as catalytic residues, and thus pepsinogens are classified as aspartic proteinases. Conversion of pepsinogens to pepsins proceeds autocatalytically at acidic pH by two different pathways, a one-step pathway to release the intact activation segment directly, and a stepwise pathway through a pseudopepsin(s). The active-site cleft is large enough to accommodate at least seven residues of a substrate, thus forming S₄ through S₃′ subsites. Hydrophobic and aromatic amino acids are preferred at the P₁ and P₁′ positions. Interactions at additional subsites are important in some cases, for example with cleavage of κ-casein by chymosin. Two potent naturally occurring inhibitors are known: pepstatin, a pentapeptide from Streptomyces, and a unique proteinous inhibitor from Ascaris. Pepsinogen genes comprise nine exons and may be multiple, especially for pepsinogen A. The latter and progastricsin predominate in adult animals, while pepsinogen F and prochymosin are the main forms in the fetus/infant. The switching of gene expression from fetal/infant to adult-type pepsinogens during postnatal development is noteworthy, being regulated by several factors, including steroid hormones. Received 25 May 2001; received after revision 27 August 2001; accepted 30 August 2001     

Role of FGFRL1 and other FGF signaling proteins in early kidney development

The mammalian kidney develops from the ureteric bud and the metanephric mesenchyme. In mice, the ureteric bud invades the metanephric mesenchyme at day E10.5 and begins to branch. The tips of the ureteric bud induce the metanephric mesenchyme to condense and form the cap mesenchyme. Some cells of this cap mesenchyme undergo a mesenchymal-to-epithelial transition and differentiate into renal vesicles, which further develop into nephrons. The developing kidney expresses Fibroblast growth factor (Fgf)1, 7, 8, 9, 10, 12 and 20 and Fgf receptors Fgfr1 and Fgfr2. Fgf7 and Fgf10, mainly secreted by the metanephric mesenchyme, bind to Fgfr2b of the ureteric bud and induce branching. Fgfr1 and Fgfr2c are required for formation of the metanephric mesenchyme, however the two receptors can substitute for one another. Fgf8, secreted by renal vesicles, binds to Fgfr1 and supports survival of cells in the nascent nephrons. Fgf9 and Fgf20, expressed in the metanephric mesenchyme, are necessary to maintain survival of progenitor cells in the cortical region of the kidney. FgfrL1 is a novel member of the Fgfr family that lacks the intracellular tyrosine kinase domain. It is expressed in the ureteric bud and all nephrogenic structures. Targeted deletion of FgfrL1 leads to severe kidney dysgenesis due to the lack of renal vesicles. FgfrL1 is known to interact mainly with Fgf8. It is therefore conceivable that FgfrL1 restricts signaling of Fgf8 to the precise location of the nascent nephrons. It might also promote tight adhesion of cells in the condensed metanephric mesenchyme as required for the mesenchymal-to-epithelial transition.     

Heavy and light roles: myosin in the morphogenesis of the heart

Myosin is an essential component of cardiac muscle, from the onset of cardiogenesis through to the adult heart. Although traditionally known for its role in energy transduction and force development, recent studies suggest that both myosin heavy-chain and myosin light-chain proteins are required for a correctly formed heart. Myosins are structural proteins that are not only expressed from early stages of heart development, but when mutated in humans they may give rise to congenital heart defects. This review will discuss the roles of myosin, specifically with regards to the developing heart. The expression of each myosin protein will be described, and the effects that altering expression has on the heart in embryogenesis in different animal models will be discussed. The human molecular genetics of the myosins will also be reviewed.     

The opportunistic pathogen Pseudomonas aeruginosa activates the DNA double-strand break signaling and repair pathway in infected cells

Highly hazardous DNA double-strand breaks can be induced in eukaryotic cells by a number of agents including pathogenic bacterial strains. We have investigated the genotoxic potential of Pseudomonas aeruginosa, an opportunistic pathogen causing devastating nosocomial infections in cystic fibrosis or immunocompromised patients. Our data revealed that infection of immune or epithelial cells by P. aeruginosa triggered DNA strand breaks and phosphorylation of histone H2AX (γH2AX), a marker of DNA double-strand breaks. Moreover, it induced formation of discrete nuclear repair foci similar to gamma-irradiation-induced foci, and containing γH2AX and 53BP1, an adaptor protein mediating the DNA-damage response pathway. Gene deletion, mutagenesis, and complementation in P. aeruginosa identified ExoS bacterial toxin as the major factor involved in γH2AX induction. Chemical inhibition of several kinases known to phosphorylate H2AX demonstrated that Ataxia Telangiectasia Mutated (ATM) was the principal kinase in P. aeruginosa-induced H2AX phosphorylation. Finally, infection led to ATM kinase activation by an auto-phosphorylation mechanism. Together, these data show for the first time that infection by P. aeruginosa activates the DNA double-strand break repair machinery of the host cells. This novel information sheds new light on the consequences of P. aeruginosa infection in mammalian cells. As pathogenic Escherichia coli or carcinogenic Helicobacter pylori can alter genome integrity through DNA double-strand breaks, leading to chromosomal instability and eventually cancer, our findings highlight possible new routes for further investigations of P. aeruginosa in cancer biology and they identify ATM as a potential target molecule for drug design.