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A L Joyner

Publications and source records attributed to A L Joyner.

At least 73 records · Page 4Linked to original sources
PubMed1993

Targeted disruption of the trkB neurotrophin receptor gene results in nervous system lesions and neonatal death.

We have generated mice carrying a germline mutation in the tyrosine kinase catalytic domain of the trkB gene. This mutation eliminates expression of gp145trkB, a protein-tyrosine kinase that serves as the signaling receptor for two members of the nerve growth factor family of neurotrophins, brain-derived neurotrophic factor and neurotrophin-4. Mice homozygous for this mutation, trkBTK(-/-), develop to birth. However, these animals do not display feeding activity, and most die by P1. Neuroanatomical examination of trkBTK (-/-) mice revealed neuronal deficiencies in the central (facial motor nucleus and spinal cord) and peripheral (trigeminal and dorsal root ganglia) nervous systems. These findings illustrate the role of the gp145trkB protein-tyrosine kinase receptor in the ontogeny of the mammalian nervous system.

Animals↗
PubMed1993

Regional assignment of the human homeobox-containing gene EN1 to chromosome 2q13-q21.

The human homeobox-containing genes EN1 and EN2 are closely related to the Drosophila pattern formation gene engrailed (en), which may be important in brain development, as shown by gene expression studies during mouse embryogenesis. Here, we have refined the localization of EN1 to human chromosome 2q13-q21 using a mapping panel of rodent/human cell hybrids containing different regions of chromosome 2 and a lymphoblastoid cell line with an interstitial deletion, del(2)(q21-q23.2). This regional assignment of EN1 increases to 22 the number of currently known genes on human chromosome 2q that have homologs on the proximal region of mouse chromosome 1.

Animals↗
PubMed1993

Dynamic expression of the murine Achaete-Scute homologue Mash-1 in the developing nervous system.

The Drosophila Achaete-Scute Complex genes encode transcriptional regulators belonging to the basic-helix-loop-helix family which control early steps of development of the central and peripheral nervous systems. We have isolated two mouse homologues of Achaete-Scute Complex genes, Mash-1 and Mash-2, by using the conservation of the basic-helix-loop-helix domain in this family. In this article, we analyse the expression of Mash-1 from its onset during neurulation to adult stages by RNA in situ hybridization on whole mounts and sections. As was observed for the rat Mash-1 protein, mouse Mash-1 RNA expression is restricted to cells of the developing central and peripheral nervous systems. We have observed three successive phases in the distribution of Mash-1 transcripts in the developing central nervous system. Initially, between embryonic day 8.5 and 10.5, Mash-1 transcripts are found in restricted domains in the neuroepithelium of the midbrain and ventral forebrain, as well as in the spinal cord. Between embryonic day 10.5 and 12.5, Mash-1 expression pattern changes from a restricted to a widespread one. Mash-1 transcripts are then found at variable levels in the ventricular zone in all regions of the brain. From embryonic day 12.5 to post-natal stages, Mash-1 is also expressed in cells outside of the ventricular zone throughout the brain. In addition, Mash-1 is expressed during development of the olfactory epithelium and neural retina. Overall, its expression pattern suggest that Mash-1 plays a role at early stages of development of specific neural lineages in most regions of the central nervous system and of several lineages in the peripheral nervous system. We have also compared the expression of Mash-1 and mouse Notch because their Drosophila homologues have been shown to interact genetically. The two genes show very similar expression patterns, both spatially and temporally, in the early developing brain and in the retina, suggesting that both genes may participate in the development of the same neural lineages.

Amino Acid Sequence↗
PubMed1993

A mouse model of greig cephalopolysyndactyly syndrome: the extra-toesJ mutation contains an intragenic deletion of the Gli3 gene.

Greig cephalopolysyndactyly syndrome (GCPS) is an autosomal dominant disorder affecting limb and craniofacial development. Recently, the human GLI3 gene has been proposed to be a candidate gene for GCPS. Here we describe the molecular characterization of extra-toes (Xt), which is a mouse model of GCPS. The Xt heterozygotes show craniofacial defects and a polydactyly phenotype similar to GCPS. We show that a deficiency of Gli3 expression in the XtJ mutant is due to a deletion within the 3' end of the gene. Furthermore, structures affected in the mouse mutant and human syndrome were found to correlate with expression domains of Gli3 in mouse. These results strongly suggest that the deficiency of GLI3 function leads to GCPS.

Alleles↗
PubMed1993

Two enhancer regions in the mouse En-2 locus direct expression to the mid/hindbrain region and mandibular myoblasts.

An En-2/lacZ gene fusion containing 9.5 kb of En-2 genomic DNA was capable of directing lacZ expression in an En-2-specific manner both temporally and spatially during embryogenesis and in the adult. lacZ expression was confined in the embryo to cells within the mid/hindbrain and mandibular arch regions and in the adult to cells of the molecular and granular layers of the cerebellum, and within the pons and colliculi regions. Interestingly, in the adult, transgene expression patterns within the cerebellum in two lines appeared to mark distinct anterior-posterior compartments. Analysis of the expression pattern of this transgene, in fetal and adult mice lacking a functional En-2 protein, provided evidence that the En-2 gene in mouse is not autoregulated. Deletion analysis of the En-2 genomic region and the use of a heterologous promoter identified two enhancer-containing regions of 1.5 and 1.0 kb in length, 5' of the transcribed sequences, which independently directed expression in the embryo to either the mid/hindbrain region or mandibular myoblasts, respectively. The 1.5 kb fragment contains the most anterior neural enhancer and the 1.0 kb fragment, the earliest myogenic enhancer thus far characterized. These En-2-specific regulatory regions can now be used in a biochemical analysis to identify proteins important in anterior-posterior patterning of the vertebrate CNS and in the specification of muscle identity as well as in a mutational analysis to direct expression of other developmentally important genes to these regions.

Animals↗
PubMed1993

Culture and transplantation of the mammalian circadian pacemaker.

In transplantation studies using the tau mutation in the golden hamster, it has been demonstrated that suprachiasmatic nucleus (SCN) pacemaker cells and mechanisms of communication with the host brain are retained even after tissue dissociation and maintenance for many weeks in primary cell culture. Brain grafts of cultured SCN cells are capable of restoring overt rhythms of locomotor activity, and preliminary studies where cells from two tau genotypes are combined in a single graft demonstrate that pacemaker cells may communicate with each other to produce coherent rhythms with intermediate periods. The opportunity is presented, therefore, to study pacemaker-pacemaker communication in circadian chimeras produced by SCN transplantation. Immunocytochemical analysis of graft-host interactions requires the positive identification of host versus donor cells. Although grafted blocks of tissue are easily recognized during immunocytochemical analysis, implants of dissociated and cultured cells may be more diffusely located and are not as readily identified. Unless distinct strain- or species-specific markers are available, it is difficult to identify connections that may carry timing information to the host organism. We have taken an anatomical approach that utilizes cell-labeling techniques for hamster tissue along with foreign protein expression in transgenic mice to identify patterns of communication among graft and host cells, focusing specifically on SCN-SCN communication. The data indicate the usefulness of these transgenes as markers in transplantation studies where communication between graft and host is addressed.

Animals↗
PubMed1992

The midbrain-hindbrain phenotype of Wnt-1-/Wnt-1- mice results from stepwise deletion of engrailed-expressing cells by 9.5 days postcoitum.

Mice homozygous for null alleles of the putative signaling molecule Wnt-1 have a reproducible phenotype: loss of the midbrain and adjacent cerebellar component of the metencephalon. By examining embryonic expression of the mouse engrailed (En) genes, from 8.0 to 9.5 days postcoitum, we demonstrate that Wnt-1 primarily regulates midbrain development. The midbrain itself is required for normal development of the metencephalon. Thus, the observed neonatal phenotype is explained by a series of early events, within 48 hr of neural plate induction, that leads to a complete loss of En domains in the anterior central nervous system. Wnt-1 and a related gene, Wnt-3a, are coexpressed from early somite stages in dorsal aspects of the myelencephalon and spinal cord. We suggest that functional redundancy between these two genes accounts for the lack of a caudal central nervous system phenotype.

Animals↗
PubMed1992

The gene trap approach in embryonic stem cells: the potential for genetic screens in mice.

The gene trap approach in embryonic stem cells was developed as a means to screen for genes expressed during early postimplantation development in the mouse. We have validated the approach by showing that lacZ from the integrated vector is activated by splicing to endogenous exons and expressed in embryos in patterns that mimic those of the endogenous genes. These insertions can produce developmental defects in homozygous mice. The results indicate that a large screen of gene trap cell lines on the basis of embryonic lacZ expression is feasible and should provide a new source of genes, mouse mutants and mouse strains that express lacZ in particular domains and lineages. The gene trap approach could be extended to a smaller screen for genes based on mutant phenotypes.

Amino Acid Sequence↗
PubMed1992

Cloning and sequence comparison of the mouse, human, and chicken engrailed genes reveal potential functional domains and regulatory regions.

We have isolated and characterized genomic DNA clones for the human and chicken homologues of the mouse En-1 and En-2 genes and determined the genomic structure and predicted protein sequences of both En genes in all three species. Comparison of these vertebrate En sequences with the Xenopus En-2 [Hemmati-Brivanlou et al., 1991) and invertebrate engrailed-like genes showed that the two previously identified highly conserved regions within the En protein ]reviewed in Joyner and Hanks, 1991] can be divided into five distinct subregions, designated EH1 to EH5. Sequences 5' and 3' to the predicted coding regions of the vertebrate En genes were also analyzed in an attempt to identify cis-acting DNA sequences important for the regulation of En gene expression. Considerable sequence similarity was found between the mouse and human homologues both within the putative 5' and 3' untranslated as well as 5' flanking regions. Between the mouse and Xenopus En-2 genes, shorter stretches of sequence similarity were found within the 3' untranslated region. The 5' untranslated regions of the mouse, chicken and Xenopus En-2 genes, however, showed no similarly conserved stretches. In a preliminary analysis of the expression pattern of the human En genes, En-2 protein and RNA were detected in the embryonic and adult cerebellum respectively and not in other tissues tested. These patterns are analogous to those seen in other vertebrates. Taken together these results further strengthen the suggestion that En gene function and regulation has been conserved throughout vertebrate evolution and, along with the five highly conserved regions within the En protein, raise an interesting question about the presence of conserved genetic pathways.

Amino Acid Sequence↗
PubMed1992

A targeted mutation reveals a role for N-myc in branching morphogenesis in the embryonic mouse lung.

The N-myc proto-oncogene encodes a putative transcription factor that has been postulated to be involved in the control of differentiation in a number of lineages at various stages during mammalian embryogenesis. We have generated a leaky mutation in N-myc by gene targeting in embryonic stem cells. In this allele, the neo(r) gene was inserted into the first intron of N-myc, in such a way that alternative splicing around this insertion could result in the generation of a normal N-myc transcript in addition to a mutant transcript. Mice homozygous for this mutation died immediately after birth owing to an inability to oxygenate their blood. Histological examination revealed a marked underdevelopment in the lung airway epithelium, resulting in a decreased respiratory surface area. Analysis of N-myc expression in wild-type and homozygous mutant embryonic lungs suggests that N-myc is required for the proliferation of the lung epithelium in response to local inductive signals emanating from the lung mesenchyme. Homozygous mutant embryos were slightly smaller than normal and also had a marked reduction in spleen size, whereas other tissues that normally express N-myc appeared to be unaffected by the mutation. Molecular analysis revealed that normal N-myc transcripts were found in tissues from homozygous mutant embryos. Different tissues expressed the normal N-myc transcript at different levels relative to those observed in wild-type embryos, with the lowest levels being observed in the lungs. These results illustrate one way in which gene targeting can be used to generate partial loss-of-function mutations and support the importance of generating a series of alleles at a given locus to elucidate the various different functions of a gene during development.

Animals↗
PubMed1992

A gene trap approach in mouse embryonic stem cells: the lacZ reported is activated by splicing, reflects endogenous gene expression, and is mutagenic in mice.

We have confirmed that the gene trap vector pGT4.5 creates spliced fusion transcripts with endogenous genes and prevents the synthesis of normal transcripts at the site of integration. cDNA was prepared to the lacZ fusion transcript in three ES cell lines to recover endogenous exon sequences upstream of lacZ. Each of the clones detected a unique-sized endogenous transcript, as well as the fusion transcript in the ES cell line from which the clone was derived. Sequence analysis of these clones and larger clones isolated from a random-primed cDNA library showed that the splice acceptor was used properly. For two insertions, the expression patterns of the lacZ reporter and the associated endogenous gene were compared in situ at three embryonic stages and were found to be similar. Three gene trap insertions were transmitted into the germ line, and abnormalities were observed with two of the three insertions in the homozygous state. RNA obtained from mice homozygous for the two mutant gene trap insertions was analyzed for normal endogenous transcripts and negligible amounts were detected, indicating that little splicing around the gene trap insertion occurred. This work demonstrates the capacity of the gene trap vector to generate lacZ fusion transcripts, to accurately report endogenous gene expression, and to mutate the endogenous gene at the site of integration.

Amino Acid Sequence↗
PubMed1991

Subtle cerebellar phenotype in mice homozygous for a targeted deletion of the En-2 homeobox.

The two mouse genes, En-1 and En-2, that are homologs of the Drosophila segmentation gene engrailed, show overlapping spatially restricted patterns of expression in the neural tube during embryogenesis, suggestive of a role in regional specification. Mice homozygous for a targeted mutation that deletes the homeobox were viable and showed no obvious defects in embryonic development. This may be due to functional redundancy of En-2 and the related En-1 gene product during embryogenesis. Consistent with this hypothesis, the mutant mice showed abnormal foliation in the adult cerebellum, where En-2, and not En-1, is normally expressed.

Animals↗
PubMed1991

Gene targeting and gene trap screens using embryonic stem cells: new approaches to mammalian development.

Mouse embryonic stem cell lines offer an attractive route for introducing rare genetic alternations into the gene pool since the cells can be pre-screened in culture and the mutations then transmitted into the germline through chimera production. Two applications of this technique that seem ideally suited for a genetic analysis of development are enhancer and gene trap screens for loci expressed during gastrulation and production of targeted mutations using homologous recombination. These approaches should greatly increase the number of mouse developmental mutants available and help to elucidate the genetic hierarchy controlling embryogenesis.

Animals↗
PubMed1991

Examining pattern formation in mouse, chicken and frog embryos with an En-specific antiserum.

We have raised an antiserum, designated alpha Enhb-1, to a portion of the mouse En-2 protein containing the homeodomain. The antiserum detects both the En-1 and En-2 proteins in mouse, chick and Xenopus embryos by Western blot analysis. Using whole-mount immunohistochemistry, combined in some cases with scanning electron microscopy, we have examined the distribution of the proteins in the early embryos of these species. The major features of expression were similar. The initial production of En protein occurred, just before or during the formation of the first somites, in a band of the anterior neural plate in the prospective mid/hindbrain region. Later in development En-1 protein accumulated in the ventral ectoderm of the developing mouse and chick limb buds, indicating that a dorsal-ventral polarity is present as soon as any limb bud swelling is apparent and that, at least in the mouse, this polarity is established independently of the apical ectodermal ridge. In all three species, alpha Enhb-1 bound to a subset of ventro-lateral differentiating neurons in the spinal cord and hindbrain and their pattern of birth in the mouse reflected the division of the hindbrain into rhombomeres. En-1 protein also accumulated in a lateral stripe of dermatome in the mouse and chick, indicating a dorsal-ventral subdivision of this tissue. The results show that En expression is a good marker for pattern formation in a variety of tissues and will be useful in experimental studies designed to characterize further these processes.

Animals↗
PubMed1989

Mouse embryonic stem cells and reporter constructs to detect developmentally regulated genes.

A strategy was devised for identifying regions of the mouse genome that are transcriptionally active in a temporally and spatially restricted manner during development. The approach is based on the introduction into embryonic stem cells of two types of lacZ reporter constructs that can be activated by flanking mouse genomic sequences. Embryonic stem cells containing the lacZ constructs were used to produce chimaeric mice. Developmental regulation of lacZ expression occurred at a high frequency. Molecular cloning of the flanking endogenous genes and introduction of these potential insertional mutations into the mouse germ line should provide an efficient means of identifying and mutating novel genes important for the control of mammalian development.

Animals↗