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PubMed1996

Patterns of distal-less gene expression and inductive interactions in the head of the direct developing frog Eleutherodactylus coqui.

The direct developing frog Eleutherodactylus coqui exhibits radical changes in its embryogenesis. A frog-like head forms directly with no appearance of a cement gland or several jaw cartilages characteristic of tadpoles, and limbs appear early in development. The numerous differences in the embryogenesis of E. coqui provide an opportunity to examine developmental causes for the evolutionary shift from biphasic to direct development. We have cloned DNA fragments corresponding to four E. coqui genes related to the Drosophila distal-less gene Dll. While the expression patterns of the distal-less genes are generally conserved, there are some spatiotemporal differences when embryos of E. coqui are compared to those of Xenopus laevis. The changes in gene expression are correlated with the embryonic changes in head structures including craniofacial cartilages and in particular, the cement gland. We have then examined inductive interactions involved in cement gland formation by interspecific transplants and recombinants. E. coqui embryos can generate signaling that culminates in cement gland formation, but E. coqui ectoderm appears to be incapable of a cement gland response. These results show here that inductive interactions in the anterior region of the E. coqui embryo have been modified during the evolution of direct development, and that changes in the competence of the E. coqui ectoderm may be responsible for the loss of certain tadpole-specific structures, such as cement gland.

Amino Acid Sequence↗
PubMed1997

A human homologue (BICD1) of the Drosophila bicaudal-D gene.

We previously isolated a cDNA fragment homologous to the Drosophila Bicaudal-D gene (Bic-D) using a hybridization selection procedure with cosmids derived from the short arm of human chromosome 12. A PCR-mediated cDNA cloning strategy was applied to obtain the coding sequence of the human homologue (BICD1) and to generate a partial mouse (Bicdh1) cDNA. The Drosophila Bicaudal-D gene encodes a coiled coil protein, characterized by five alpha-helix domains and a leucine zipper motif, that forms part of the cytoskeleton and mediates the correct sorting of mRNAs for oocyte- and axis-determining factors during oogenesis. Analysis of the predicted amino acid sequence of the BICD1 cDNA clones indicates that the sequence similarity is essentially limited to the amphipatic helices and the leucine zipper, but the conserved order of these domains suggests a similar function of the protein in mammalians. A database search further indicates the existence of a second human homologue on chromosome arm 9q and a Caenorhabditis elegans homologue. Northern blot analysis indicates that both the human and the murine homologues produce an mRNA species of congruent with9.5 kb expressed in brain, heart, and skeletal muscle and during mouse embryonic development. The conserved structural characteristics of the BICD1 protein and its expression in muscle and especially brain suggest that BICD1 is a component of a cytoskeleton-based mRNA sorting mechanism conserved during evolution.

Adaptor Proteins, Signal Transducing↗
PubMed1998

New eukaryotic semaphorins with close homology to semaphorins of DNA viruses.

Semaphorins were initially described as a family of repulsive guidance molecules in embryonal development. Their basic structure consists of an N-terminal signal sequence, the defining semaphorin domain ofapproximately 500 amino acids, an Ig-like domain,and a variable carboxy-terminus. We recently described a viral semaphorin homologue encoded by the alcelaphine herpesvirus type 1. Less conserved, truncated homologues were also identified in poxviruses. Here we describe new human and murine semaphorin homologues. The respective genes were cloned and sequenced, and they were termed H-Sema-L and M-Sema-L (HGMW-approved symbols SEMAL and Semal, respectively). A multiply spliced mRNA of 3.2 kb is expressed in human placenta, spleen, thymus, and gonadal tissue. H-Sema-L maps to chromosome 15q22.3-q23 and M-Sema-L to the homologous locus 9A3.3-B in the mouse genome. The expression patterns and the presence of related genes in large DNA viruses suggest that this new semaphorin has a relevant function in the immune system.

Adjuvants, Immunologic↗
PubMed1985

Light and electron microscopy study of fusion of facial prominences. A distinctive type of superficial cells at the contact sites.

The contact site between the medial nasal prominence (MNP) and the lateral nasal prominence (LNP) during the period of primary palate formation in the mouse embryo was examined by light and electron microscopy. Throughout this period, a distinctive type of superficial cell was observed at the contact site. These superficial cells had a large nucleus and abundant cytoplasm as well as structural features characteristic of embryonic cells. At earlier stages, these cells were seen at the transitional region between the surface ectoderm and the epithelia of the nasal pit at the end of the isthmus, where initial contact of opposing MNP and LNP took place. At later stages, these superficial cells appeared to bridge the gap between MNP and LNP at the contact sites, which extended to the bottom of the valley formed by MNP and LNP. These cells were also observed on the surface near the contact sites, that is, the presumptive fusion area. These superficial cells displayed well-developed junctional complexes (intermediate and gap junctions, and desmosomes). Many filaments were observed subjacent to the plasma membranes of these superficial cells, some of which were associated with junctional complexes. These observations suggest that this kind of distinctive superficial cell may play critical roles in the contact of MNP and LNP throughout the fusion process.

Animals↗
PubMed1985

Effects of retinoic acid on adult human epidermis in whole skin organ culture.

Adult human skin was cultured in whole-skin organ culture under chemically defined conditions. Retinoic acid was added to the culture at final concentrations of 5 X 10(-7) and 5 X 10(-6) M. Both concentrations elicited cell death in the upper epidermal layers and prevented the terminal differentiation of the cells to mature corneocytes. The inhibition of terminal differentiation was not permanent, as the corneocytes produced later during the culture showed no signs of inhibition. The upper vital cells in epidermis cultured with retinoic acid were very flattened and contained reduced amounts of cytoskeleton components. Fine, granular material not present in normal epidermis was observed in both the intercellular spaces and the intracytoplasmic vesicles of retinoid-treated epidermis. The present results indicate that the response of normal human skin to retinoic-acid treatment involves the same kind of modulation of the epidermal structure previously described in embryonic avian and diseased human skin.

Epidermis↗
PubMed1997

Distribution of glial fibrillary acidic protein-immunopositive structures in the developing brain of the turtle Mauremys leprosa.

This study is a continuation of the description of the glial fibrillary acidic protein (GFAP)-immunopositive structures in the adult turtle brain (Kálmán et al. 1994) and presents a comprehensive description of the development of these structures from the 20th embryonic day (E20) to the adult age. GFAP-immunopositive elements were first detected at E28 and by E34 the GFAP-immunopositivity was apparent throughout the brain, except the cerebellum. The appearance of GFAP seemed to be related to the end of cell migration and the formation of the thickened parts of the brain wall, such as the dorsal ventricular ridge. After hatching the pattern of the GFAP-immunopositivity differed from that in the adult only in minute details, except for the brain tracts in which GFAP-pattern was still changing due to myelination, and the molecular layer of the cerebellum in which a transverse fiber system appeared. The GFAP-positive elements belonged originally to the ependymoglia, but later the distortion due to the morphogenetic processes of branching and division changed the pattern almost beyond recognition. In some cases cell bodies--ependymal and non-ependymal--appeared to be GFAP-positive, but no astrocytes (i.e. stellate cells) were detected. The results are discussed in the light of previous observations on developing mammalian, avian and lizard brains.

Aging↗
PubMed2000

Neurogenin 1 null mutant ears develop fewer, morphologically normal hair cells in smaller sensory epithelia devoid of innervation.

The proneuronal gene neurogenin 1 (ngn1) is essential for development of the inner-ear sensory neurons that are completely absent in ngn1 null mutants. Neither afferent, efferent, nor autonomic nerve fibers were detected in the ears of ngn1 null mutants. We suggest that efferent and autonomic fibers are lost secondarily to the absence of afferents. In this article we show that ngn1 null mutants develop smaller sensory epithelia with morphologically normal hair cells. In particular, the saccule is reduced dramatically and forms only a small recess with few hair cells along a duct connecting the utricle with the cochlea. Hair cells of newborn ngn1 null mutants show no structural abnormalities, suggesting that embryonic development of hair cells is independent of innervation. However, the less regular pattern of dispersal within sensory epithelia may be caused by some effects of afferents or to the stunted growth of the sensory epithelia. Tracing of facial and stato-acoustic nerves in control and ngn1 null mutants showed that only the distal, epibranchial, placode-derived sensory neurons of the geniculate ganglion exist in mutants. Tracing further showed that these geniculate ganglion neurons project exclusively to the solitary tract. In addition to the normal complement of facial branchial and visceral motoneurons, ngnl null mutants have some trigeminal motoneurons and contralateral inner-ear efferents projecting, at least temporarily, through the facial nerve. These data suggest that some neurons in the brainstem (e.g., inner-ear efferents, trigeminal motoneurons) require afferents to grow along and redirect to ectopic cranial nerve roots in the absence of their corresponding sensory roots.

Animals↗
PubMed2004

Collagen types XII and XIV are present in basement membrane zones during human embryonic development.

The collagens constitute a large group of proteins in the extracellular matrix that can be divided into several distinct families. Collagen types XII and XIV belong to a subgroup of non-fibrillar-collagens termed (fibril-associated collagens with interrupted triple-helices) (FACIT) and may be involved in basement membrane regulation providing specific molecular bridges between fibrils and other matrix components. However, the tissue distribution of the two proteins during human embryogenesis is still unclear. As a first step toward the elucidation of their possible cell biological functions, we compared the distribution of the two collagens during human organogenesis at the light microscopical level. We detected specific differences between the expression patterns of the two molecules, which may be related to their respective function within the basement membrane zones during human embryonic development. For example, in the developing intestine, collagen type-XII was present in the basement membrane zones of epithelia and endothelia. However, collagen type-XIV was restricted to the mesothelial basement membrane zones. We conclude that both collagens might well be able to serve different functions during human embryonic development although their structures are highly similar.

Basement Membrane↗
PubMed1990

Vitellogenesis in Drosophila: sequestration of a yolk polypeptide/invertase fusion protein into developing oocytes.

The mechanism of yolk deposition into developing oocytes of Drosophila was investigated by following the fate of a reporter protein fused to a vitellogenin, or yolk polypeptide (YP). Embryos were transformed with a hybrid gene consisting of the promotor and amino terminal 430 codons of the Yp2 gene fused to the cytoplasmic form of the invertase gene from the yeast Saccharomyces cerevisiae. RNA hybridization experiments with established lines of transformed flies showed that the hybrid gene was expressed in female fat bodies and ovaries but not in any male cells. Immunoblotting and endoglycosidase digestion showed that the hybrid protein was secreted from fat body cells via the secretory pathway, transported in hemolymph, and sequestered into developing oocytes. Transfusion experiments with hemolymph and pure invertase showed that sequestration of invertase depended on its attachment to YP. Immunocytochemistry demonstrated that the hybrid protein became localized in yolk granules as oocytes developed. Females homozygous for the fusion gene are generally sterile; their eggs containing the hybrid protein often collapse and their embryos fail to develop, suggesting that the structure of the yolk polypeptides is important for embryonic development. These experiments show that YP2 carries structural information sufficient to direct a reporter protein from fat body cells, through the hemolymph, and into the yolk granules of developing oocytes. This work provides a means of identifying the features of yolk polypeptides that are responsible for their deposition into yolk during oogenesis.

Amino Acid Sequence↗
PubMed1990

Membrane skeleton protein 4.1 in developing Xenopus: expression in postmitotic cells of the retina.

Membrane skeleton protein 4.1 plays a key role in modulating the interactions of spectrin, actin, and integral membrane proteins in erythroid and nonerythroid cells. We have investigated its structure and expression during embryonic development of Xenopus laevis. An analysis of the complete 2758-nucleotide sequence and predicted translation of 801 amino acids (85.5 kDa) of X. laevis oocyte protein 4.1 reveals that, within overlapping regions, oocyte protein 4.1 is 74% identical to a composite amino acid sequence of human erythroid and lymphoid protein 4.1 and has an identity similar to that of amino acid motifs variably expressed in either human erythroid or lymphoid protein 4.1 S1 nuclease protection analysis demonstrates the presence of a single species of protein 4.1 transcript in embryos. Antibodies produced against X. laevis protein 4.1 fusion protein recognize two bands of 180 and 115 kDa on Western blots of X. laevis embryos and retina and, using immunocytochemical techniques, label the developing retina most intensely. In vitro transcription of a cDNA construct fully encoding X. laevis protein 4.1 yields a synthetic mRNA which, when translated in vitro, produces a polypeptide that comigrates on SDS-polyacrylamide gels with the 115-kDa form of embryos and retina. Protein 4.1 is found exclusively in photoreceptors following the terminal mitosis of retinal neurons. When retinal synaptogenesis is complete, protein 4.1 is also expressed in the inner retina. In adult amphibian retinas, protein 4.1 is detected in photoreceptors, bipolar cells, and ganglion cell axons. As these cell types have previously been shown to express spectrin, actin, and ankyrin, it is likely that the membrane skeleton of erythrocytes and retinal cells share functional similarities.

Amino Acid Sequence↗
PubMed1984

Chromatin structure, DNA synthesis and transcription through the lifespan of human embryonic lung fibroblasts.

The presence in terminal embryonic fibroblasts of small molecular weight (MW) DNA independent of bulk DNA could be ascertained by three different techniques performed in parallel. This alteration was not artifactually induced, either by high pH and the detergent used or by the release of cellular enzymes. An increased thermolability of old chromatin was also observed. Cells with altered chromatin synthesized DNA and RNA according to a pattern similar to young type nuclei. Long-term treatment with hydrocortisone significantly increased the cell yield but did not prevent, in the late passages, the occurrence of old-type chromatin; the nucleolar filamentous masses, however, maintained a 'young' pattern. Short-term treatment induced only a moderate reversion in the appearance of chromatin lesions. Direct evidence was obtained of increased gene expression in the presence of hydrocortisone.

Cell Division↗
PubMed1979

Organ culture of adult amphibian heart: a fine structural analysis.

Pieces of hearts from adult newts were cultured up to 2 months. Within 7 days of culture, approximately 37% of the cardiac explants were attached to the substrate and more than 33% of the attached explants and approximately 15% of the unattached explants established pulsation rates ranging from 3 to 67 beats/min. The control and cultured explants were processed at weekly intervals for electron microscopy. The diameter of the control cardiac muscle cells ranged approximately 3-5 micron. The cell surface was provided with microvilli. The intercellular spaces ranged approximately 150-500 A. The intercalated discs lacked the step-like courses observed in the mammalian cardiac muscle. Sarcoplasmic reticulum was scanty. Desmosomal-dense materials were frequently continuous with the Z-bands of both control and cultured cardiac muscle cells. The transverse tubular system and gap junction were absent in newt ventricles. The functional implications of these characterisitics are discussed. At the end of 1 week of culture, the surfaces of the explants were covered by one or more layers of non-muscle cells, and the core of the explants consisted mostly of cardiac muscle cells. In a few cardiac muscle cells the myofibrillar organization was disrupted, resulting in the distribution of scattered patches of myofibrils and free myofilaments in the sarcoplasm. A small number of intact muscle cells contained a considerable number of dense granules in the sarcoplasm. At 15 days in culture, a large number of muscle cells showed structural features reminiscent of embryonic cardiac muscle cells. These cells possessed patches of myofibrils, scattered myofilaments and scanty sarcoplasmic reticulum along with other cellular organelles and inclusions. Several of these altered cardiac muscle cells contained mitotic figures. The cardiac explants maintained the initial beating rate until the end of 2 months of culture, except for the 11% of the explants which stopped beating. By 3-4 weeks in culture, most of the cardiac muscle cells possessed the altered cell morphology mentioned above. The explants after 60 days in culture became more flattened than the earlier explants. The intact cardiac muscle cells were rare, and the cores of the explants were mostly occupied by the altered cardiac muscle cells. It is evident from our studies that the cardiac muscle cells have undergone dedifferentiation in long-term culture, and that this dedifferentiation process has yet had no effect in the maintenance of contractility of the explants. Furthermore, these dedifferentiated cardiac muscle cells are capable of DNA synthesis and mitosis.

Animals↗
PubMed1990

The specificities of Sex combs reduced and Antennapedia are defined by a distinct portion of each protein that includes the homeodomain.

The sequence requirements for distinguishing the functional specificities of two homeodomain proteins, Antennapedia and Sex combs reduced, involved in the specification of segmental identities in Drosophila, have been determined. A series of deletions and hybrid proteins was generated and assayed for their function in vivo after heat shock-induced ectopic expression during development. A distinct portion of each protein, including the residues within and adjacent to both ends of the homeodomain, has been found to almost entirely determine its functional specificity as measured by diagnostic cuticular transformations of embryonic and adult head structures. The remaining sequences contribute to the potency with which the proteins act in different cells and are to a limited extent functionally transferable from one protein to the other.

Amino Acid Sequence↗
PubMed1992

Small eye (Sey): cloning and characterization of the murine homolog of the human aniridia gene.

Phenotypic parallels and genetic evidence from comparative mapping suggest that the murine Small eye (Sey) and human aniridia (AN) disorders are homologous. This report describes the isolation of a murine embryonic cDNA that is structurally homologous to the AN cDNA were recently cloned. The murine cDNA detects a 2.7-kb transcript in the adult mouse eye and cerebellum and in human glioblastomas, suggesting a neuroectodermal involvement in the etiology of Sey/AN. Sequence comparison between the murine and the human cDNAs revealed extensive homology in nucleotide sequence (greater than 92%) and virtual identity at the amino acid level. None of the differing amino acids was located within the paired box and homeobox DNA-binding domains. These results provide evidence for a common molecular basis underlying the two genetic disorders and suggest that the Sey system would be an authentic model for human AN.

Amino Acid Sequence↗
PubMed2004

Caudal duplication syndrome: more evidence for theory of caudal twinning.

Caudal duplication syndrome is a rare entity in which structures derived from the embryonic cloaca and notochord are duplicated to various extents. The term encompasses a spectrum and often is quoted as one type of incomplete separation of monovular twins. The authors present more evidence giving credence to caudal twining as the mechanism behind the syndrome. The authors report successful surgical management of a full-term infant with a constellation of anomalies of caudal duplication syndrome.

Abnormalities, Multiple↗
PubMed2006

Formation of leptofibrils is associated with remodelling of muscle cells and myofibrillogenesis in the border zone of myocardial infarction.

Leptofibrils, or leptomeres, remain the least studied cytoskeletal structures in muscle cells, and their function and mechanism of assembly are still poorly understood. Our ultrastructural study of the surviving cardiac myocytes located in the perinecrotic border zone of the infarcted left ventricle in rats revealed intense formation of leptofibrils and leptofibrillar clusters during 4-15 days following experimental myocardial infarction. In the perinecrotic myocytes, leptofibrils developed predominantly in the subsarcolemmal areas, near disassembled intercalated discs and at the sites of intense myofibrillogenesis in the peripheral zones of the sarcoplasm. We found that the development of these structures occurred before or at the time of assembly of myofibrils. In our material, leptofibrils consisted of longitudinally oriented filamentous bundles inserted in electron dense Z-band-like material and periodically crossed by 3-8 bands of this material with the period of cross-striation of 120-210 nm. The presence of leptofibrils in growing cytoplasmic processes and ruffles developing in the border zone in the areas of lost intercellular contacts indicates their formation de novo during post-infarction period. We observed four major morphological types of localization of these structures: (1) direct contact of one end of leptofibrils with Z bands of nascent, mature or disassembling myofibrils; (2) direct contact with the sarcolemma: (a) multifocal attachment of leptofibrils to the sarcolemma through the lateral surfaces of their minute Z band-like structures; (b) attachment of one or both ends of leptofibrils to the sarcolemma without contacts or in contact with myofibrils; (3) attachment of leptofibrils to subsarcolemmal accumulations of electron dense Z-band material in newly formed fasciae adherentes of the remodeled intercalated disks; (4) clustering and contacts of leptofibrils with one another predominantly at the level of their Z bands. Interestingly, most leptofibrils of all four types were topographically associated with the system of T-tubules, the sarcoplasmic reticulum and subsarcolemmal vesicles. Serial sections through the areas containing leptofibrils indicate their spindle-like or nearly cylindrical shape. Thus, we found that leptofibrils assemble in terminally differentiated cardiac myocytes following destabilization of their differentiated state and partial dedifferentiation induced by myocardial infarction. The results of this study demonstrate that formation of leptofibrils, earlier described mainly in the developing and malignant muscle, is temporally associated with adaptive structural remodelling and the activation of myofibrillogenesis in functionally overloaded cardiac myocytes of adult animals. Our findings suggest that re-expression of some structural characteristics of the embryonic muscle appear to represent one of the mechanisms that underlie adaptive plasticity of the myocardium following injury and under conditions of hyperfunction.

Animals↗