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Biomedical subjects

D G Wilkinson

Publications and source records attributed to D G Wilkinson.

At least 73 records · Page 4Linked to original sources

The psychogeriatrician's view: management of chronic disability in the community.

Psychiatric services for elderly people are based on principles which are relevant to the management of chronic disorders such as Parkinson's disease. Psychological and biological factors are closely intertwined in dementia, as they are in PD. Although medical diagnosis is important the emphasis is on continuing support. The aim is to provide services within the community where they are needed, and to acknowledge the importance of carers and family for assessment and management.

Activities of Daily Living↗

Cloning and developmental expression of Sna, a murine homologue of the Drosophila snail gene.

The genetic analysis of dorsoventral patterning in Drosophila has identified a zinc-finger gene, snail, that is required for mesoderm formation. The cloning and nuclease protection analysis of a Xenopus homologue of this gene has suggested a possible role in the mesoderm of vertebrates. Here, we describe the cloning of a murine homologue of snail, Sna, and in situ hybridisation studies of its developmental expression. Sequence analysis reveals substantial conservation of the second to fifth zinc fingers, but not of the first zinc finger in the Sna gene. Expression occurs in the ectoplacental cone, parietal endoderm, embryonic and extraembryonic mesoderm, in neural crest and in condensing precartilage. Based on the timing and spatial restriction of expression in embryonic mesoderm, we suggest that Sna might be required for the early development of this tissue, as is the case for its Drosophila counterpart. In addition, we propose that Sna might have an analogous role in the development of neural crest. The expression in condensing precartilage indicates that this gene also has a later function in chondrogenesis.

Amino Acid Sequence↗

A receptor protein tyrosine kinase implicated in the segmental patterning of the hindbrain and mesoderm.

Pattern formation in the hindbrain and paraxial mesoderm of vertebrates occurs by the formation of a series of repeated segments. These processes of segmentation appear different at the morphological level, since hindbrain segments, the rhombomeres, form by the subdivision of the neural epithelium into compartments, whereas the mesodermal somites form by the sequential aggregation of mesenchymal cells into epithelial balls. Previous studies have implicated genes encoding transcription factors in the development of hindbrain segments, but nothing is known of genes involved in the formation of somites. Cellular interactions and signal transduction must be an important aspect of hindbrain segmentation, so we have screened for tyrosine kinases expressed in rhombomere-restricted patterns in the developing mouse embryo. We have identified a receptor protein tyrosine kinase, Sek, that has high relative levels of expression in rhombomeres 3 and 5. This alternating pattern is established coincidentally, both spatially and temporally, with the expression of Krox-20, a zinc-finger gene expressed prior to the morphological formation of rhombomeres. In addition, Sek expression occurs in several other developing tissues, including a dynamic regulation in the developing forebrain, spinal cord, early mesoderm and anterior presomitic mesoderm (segmental plate). The latter expression occurs in two stripes that correlate with, and presage, the formation of somites. Sek expression initially occurs throughout the presumptive somite, then becomes restricted anteriorly, and finally is down-regulated as the definitive somite is formed. These data suggest that despite the morphological differences in the segmentation of the hindbrain and mesoderm, Sek is involved in the segmental patterning of both of these tissues.

Animals↗

An Eph-related receptor protein tyrosine kinase gene segmentally expressed in the developing mouse hindbrain.

In search of genes possibly involved in the regulation of hindbrain segmentation, we have isolated mouse cDNA clones corresponding to putative protein kinase genes by polymerase chain reaction amplification of cDNA from 9.5-day-old embryo hindbrains. In situ hybridization analysis revealed that one of these genes, Sek, was expressed in an alternating segment-restricted pattern in the developing hindbrain. Isolation and analysis of Sek cDNAs covering the entire coding sequence indicated that Sek encoded a putative receptor protein tyrosine kinase, belonging to the Eph family. These data are consistent with a role of the Sek gene product in a signal transduction process involved in pattern formation in the hindbrain.

Amino Acid Sequence↗

Conserved segmental expression of Krox-20 in the vertebrate hindbrain and its relationship to lineage restriction.

The zinc-finger gene Krox-20 is expressed in two alternating segments, rhombomeres (r) 3 and 5, in the developing mouse hindbrain. This expression pattern is established prior to rhombomere formation in the mouse, but it is not known how the timing of expression relates to cellular events of segmentation, such as lineage restriction. We have cloned Krox-20 sequences from Xenopus and the chick and shown that its alternating expression pattern is conserved in these systems, suggesting that its role in hindbrain development is conserved. Analysis of the early stages of Krox-20 expression in the chick show that both domains of expression precede the restriction of cell lineage to specific rhombomeres, consistent with a role of this gene in early events of hindbrain segmentation. The finding that expression is not coincident with lineage restriction indicates that early expression may not reflect an irreversible commitment of cells to r3 and r5 and/or may be mosaic.

Amino Acid Sequence↗

Expression pattern of the mouse T gene and its role in mesoderm formation.

Formation of mesoderm is a crucial event in vertebrate development, establishing many of the important features of the body. Recent studies have implicated molecules that are similar to growth factors in mesoderm formation in Xenopus, but other gene products involved in this process have yet to be identified. Genetic evidence indicates that in the mouse the T gene (Brachyury) has a role in the formation and organization of mesoderm. Mice homozygous for mutant alleles of the T gene do not generate enough mesoderm, and show severe disruption in morphogenesis of mesoderm-derived structures, in particular the notochord. The cloning of the T gene has now allowed us to examine its expression pattern. We report that T-gene expression occurs in both early stage mesoderm and its epithelial progenitor, and then becomes restricted to the notochord. This expression pattern correlates with the tissues affected in the T-gene mutant, and indicates that the T gene has a direct role in the early events of mesoderm formation and in the morphogenesis of the notochord.

Animals↗

Molecular approaches to the segmentation of the hindbrain.

Recent studies have shown that the rhombomeric bulges of the developing vertebrate hindbrain reflect segmental mechanisms that generate pattern in this region of the CNS. Although little is known of the genetic basis of this segmentation, in situ hybridization studies have provided circumstantial evidence that certain 'zinc-finger' and homeobox genes have roles in the development of segments in the early mouse hindbrain. We discuss the implications of these findings for the function of these genes in hindbrain development.

Animals↗

Segmental expression of Hox-2 homoeobox-containing genes in the developing mouse hindbrain.

The vertebrate hindbrain develops in a segmental pattern, with distinctive groups of neurons originating from different segments. We report here that members of the Hox-2 cluster of murine homoeobox genes are expressed in segment-specific patterns in the developing hindbrain, with successive genes having boundaries at two-segment intervals. These data indicate that Hox genes specify segment phenotype, a role analogous to that of their Drosophila homologues.

Animals↗

Segment-specific expression of a zinc-finger gene in the developing nervous system of the mouse.

The process of segmentation, in which repeated homologous structures are generated along the anterior-posterior axis of the embryo is a widespread mechanism in animal development. In vertebrates, segmentation is most apparent in the somites and the peripheral nervous system, but the existence of repetitive bulges, termed neuromeres, in the early neural epithelium of vertebrates suggests that the CNS may also be segmented. Consistent with this, cranial ganglia and certain neurons are associated with specific hindbrain neuromeres. Here, we report that Krox-20, a zinc-finger gene, is expressed in two alternate neuromeres in the mouse early hindbrain. This pattern subsequently decays and Krox-20 is transiently expressed in specific hindbrain nuclei. In addition, Krox-20 is expressed in early neural crest cells, and then in the neural crest-derived boundary caps, glial components of the cranial and spinal ganglia. The demonstration that neuromeres are domains of gene expression provides molecular evidence for the segmentation of the CNS.

Animals↗

Expression pattern of the FGF-related proto-oncogene int-2 suggests multiple roles in fetal development.

The FGF-related proto-oncogene int-2 is implicated in mouse embryogenesis, since it is expressed in specific tissues during gastrulation and neurulation (Wilkinson et. al. 1988). Here, we describe the expression of this gene during subsequent fetal development, int-2 transcripts are restricted to Purkinje cells in the cerebellum and to regions of the developing retina containing early-stage differentiating cells. This high level expression is not detected in the mature cerebellum or retina. In addition, int-2 RNA is detected in the mesenchyme of the developing teeth and in sensory regions of the inner ear. This complex and dynamic pattern suggests multiple roles of this proto-oncogene during fetal development of the mouse.

Animals↗

Tissue-specific expression of c-jun and junB during organogenesis in the mouse.

c-jun and junB are cellular genes related to the viral oncogene v-jun and encode members of the AP-1 transcription factor gene family. These genes have been implicated in the control of the G0/G1 transition in fibroblasts. Here, we have investigated the potential roles of c-jun and junB during fetal growth and organogenesis in the mouse by in situ hybridization analysis of their expression patterns. c-jun expression is detected throughout organogenesis, and transcripts are detected in many tissues, although in restricted cell populations within developing cartilage, gut and the central nervous system (CNS). In cartilage, c-jun expression is associated with rapidly proliferating perichondrial cells, but occurs in postmitotic motor neurones in the CNS. junB expression is initiated between 14.5 and 17.5 days of development, and is restricted to differentiating epidermal cells and endodermal gut epithelium. These data suggest that c-jun and junB have distinct, tissue-specific roles in cell proliferation and differentiation during fetal development.

Animals↗

Expression of the FGF-related proto-oncogene int-2 during gastrulation and neurulation in the mouse.

The proto-oncogene int-2 has been implicated in the formation of mouse mammary-tumour-virus-induced mammary tumours. Analysis of the predicted coding sequence indicates that int-2 is a member of the fibroblast growth factor family. Previous studies using Northern blot analysis suggested that normal expression of int-2 may be confined to extra-embryonic endoderm lineages of embryonic stages of mouse development. We have used in situ hybridization and Northern blot analysis to examine directly int-2 expression in embryo stem cells and in the developing embryo from early gastrulation to midsomite stages. Complex patterns of accumulation of int-2 RNA were observed in embryonic and extra-embryonic tissues. The data suggest multiple roles for int-2 in development which may include migration of early mesoderm cells and induction of the otocyst.

Animals↗

Expression of the proto-oncogene int-1 is restricted to specific neural cells in the developing mouse embryo.

We have used in situ hybridization and computer-aided reconstruction to study the spatial distribution of expression of the mammary tumor proto-oncogene int-1 during mouse embryogenesis. int-1 RNA accumulation is restricted to specific regions of the neural plate and its derivatives between 9 and 14.5 days of development. int-1 RNA accumulates throughout the neural plate at the anterior head folds of the 9 day embryo but only at its lateral tips in more posterior regions. Following neural tube closure, int-1 expression is restricted to specific regions of the dorsal wall of the brain ventricles and spinal cord, the ventral wall of the midbrain and the diencephalon, and the lateral walls of the neuroepithelium at the midbrain-hindbrain junction. These data suggest that int-1 has a role in the early stages of central nervous system development in the mouse embryo.

Animals↗

Metallothionein genes MTa and MTb expressed under distinct quantitative and tissue-specific regulation in sea urchin embryos.

Sea urchin embryo metallothionein (MT) mRNAs MTa and MTb have distinct cDNA sequences and are transcripts of different genes of a multigene family. These MT mRNAs differ in size and in their 3'-untranslated sequences. They encode proteins that are unusual among MT isotypes in that the relative positions of their cysteine residues are partially out of register, suggesting potential differences in function. In pluteus larvae MTa mRNA is expressed abundantly and exclusively in the ectoderm, while MTb mRNA, which is restricted to the endomesoderm at a low endogenous level, can be induced to a high level by heavy metal ions (M2+). MT mRNA is present in the maternal reservoir of the egg and is predominantly (greater than 95%) MTa mRNA. Endogenous expression in the embryo, which is at a much higher level than in the egg, requires M2+ for gene transcription, is developmentally regulated, and is greater than 90% MTa mRNA. When induced by added M2+, however, MTa and MTb mRNAs accumulate to almost equal levels. The differences in the ratios of MTa/MTb expressed endogenously and inductively are not attributable to differences in the stabilities of these MT mRNAs, which were observed under conditions of M2+ depletion, or in their inducibilities, which were observed at moderate to high M2+ levels. We found, instead, that the MTa gene responds to M2+ at a lower threshold level than MTb, so that at very low M2+ concentrations the ratio of induced MTa/MTb mRNA is high and equivalent to the endogenous ratio. Thus, endogenous expression of the MTa gene is selectively enhanced in the ectoderm by determinants that are responsive at low M2+ threshold concentrations.

Animals↗

A molecular analysis of mouse development from 8 to 10 days post coitum detects changes only in embryonic globin expression.

The pattern of protein synthesis in 8-, 9- and 10-day post coitum (p.c.) mouse embryos was examined by 2-D gel electrophoresis of [35S]methionine-labelled proteins. Of the 600-800 polypeptides detected only one, a 14 X 10(3) Mr (14K) protein, was found to accumulate over this period. To isolate cDNA clones that potentially encode this protein, 32P-labelled cDNA was synthesized from 9 and 10 days p.c. embryo poly(A) +RNA, and used for the differential screening of an 8.5-day p.c. mouse embryo cDNA library cloned in lambda gt10. Six clones that hybridized strongly to the 10-day probe were purified and their inserts subcloned into plasmid vectors. Cross hybridization and restriction mapping of these inserts indicate that they fall into four distinct groups. Each of these hybridize with transcripts of approximately 600 nucleotides, which accumulate in the embryo from 9 to 10 days p.c. Expression was barely detectable in adult tissues and restricted to liver and spleen. Expression of one of these clones, 10.1, was examined by in situ hybridization of 35S-labelled RNA probes to 8.5-12.5 day p.c. embryo sections. Strong hybridization was observed in yolk sac blood islands, fetal liver and embryonic erythrocytes, suggesting that 10.1A encodes an erythrocyte-specific protein. DNA sequence analysis indicates that the four classes of cDNA were derived from transcripts of the alpha 1, zeta, beta h1 and epsilon globin genes. Labelling of 10-day p.c. erythrocyte proteins with [35S]methionine, followed by 2-D gel electrophoresis, clearly demonstrates that the most abundant polypeptide migrates to the same position as the 14K protein which accumulates from 8 to 10 days p.c. Thus the only abundant transcripts and corresponding proteins that change over a period of profound morphogenetic change correspond to globins of the newly established blood system.

Amino Acid Sequence↗

Spatial patterns of metallothionein mRNA expression in the sea urchin embryo.

Metallothioneins (MTs) are small, cysteine-rich proteins that bind heavy metals which induce their synthesis. Tissue fractionation of embryos at pluteus stage previously demonstrated that in the absence of added zinc, basal expression of MT mRNA is confined to ectoderm, whereas induction by zinc results in increased expression in the endoderm + mesoderm tissue fraction. Using in situ hybridization we now show that expression in the pluteus larva is restricted almost exclusively to the single cell type comprising the aboral ectoderm. Induction by Zn results in a marked accumulation of MT mRNA in gut and oral ectoderm to levels at least as high as that in aboral ectoderm. MT mRNA is also expressed in presumptive aboral ectoderm at earlier stages of normal development. In addition it is transiently expressed at variable levels in oral ectoderm and, to a lesser extent, in presumptive gut.

Animals↗

Spore coat protein synthesis during development of Dictyostelium discoideum requires a low-molecular-weight inducer and continued multicellularity.

The major spore coat proteins of Dictyostelium discoideum are synthesized during the culmination stage of development. In an attempt to examine the regulatory mechanisms involved, spore coat protein synthesis by pseudoplasmodia harvested prior to culmination and incubated in submerged culture under various environmental conditions has been monitored. It is reported that the synthesis of spore coat proteins SP170, SP103, SP94, SP82, SP76, and SP55 is dependent upon the presence of a low-molecular-weight (Mr approx 100), heat-stable factor secreted by cells incubated at high density in buffer. Previous studies have implicated cyclic AMP, ammonia, and amino acids in spore cell differentiation. Partial purification of the spore coat protein inducing factor (SPIF), together with attempts to mimic its activity, indicate that SPIF is not identical with any of these molecules and it is probably also distinct from DIF and "fruit juice," two other factors which regulate the spore-stalk decision and the initiation of culmination, respectively, in D. discoideum. In addition to SPIF, the continued expression of the spore coat protein genes also requires that the integrity of the pseudoplasmodium be maintained. Unlike the expression of many other genes after aggregation, this latter requirement cannot be replaced by exogenous cyclic AMP. Termination of spore coat protein gene expression occurs despite the presence of excess exogenous SPIF and hence involves mechanisms other than the destruction or depletion of SPIF.

Age Factors↗

Primary differentiation and ectoderm-specific gene expression in the animalized sea urchin embryo.

Primary differentiation in sea urchin embryos, animalized by zinc, has been gauged by the formation of characteristic endodermal and mesodermal tissue derivatives and by the accumulation of the ectoderm-specific Spec 1 mRNA. Increasing the dosage of zinc diminishes the differentiation of secondary mesenchyme, primary mesenchyme, endoderm, and ectoderm, in decreasing order. Treatment is effective only during the blastula stages, involving successive periods of sensitivity for these tissues. Removal of zinc with chelator results in the resumption of differentiation to increasing degree for this series of tissues. The developmental initiation of Spec 1 gene expression, normally at the earliest blastula stage, can be delayed by zinc for at least 30 hr before being implemented by treatment of the animalized embryos with a chelator. We conclude (1) that those processes in the blastula which are required for differentiation and are suppressed by zinc are distinguishable from the determinative processes, which are not affected by the animalizing agent and occur earlier during midcleavage; (2) that animalization by zinc involves a graded failure of primary tissues to form; and (3) that animalization involves a pause in the schedule of differentiation, which can be reinstated by removal of the animalizing agent, thereby providing a survival value inherent in a flexible schedule of development.

Animals↗