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K J Fowler

Publications and source records attributed to K J Fowler.

At least 19 recordsLinked to original sources
PubMed2004

Centromere protein b-null mice display decreasing reproductive performance through successive generations of breeding due to diminishing endometrial glands.

Centromere protein B is a highly conserved constitutive protein found at centromeres. Gene knockout studies in mice have unexpectedly identified Cenpb as a candidate gene involved in uterine function. The present study further explores the role of Cenpb in mice by intermating Cenpb-null mice over several generations. Breeding studies and analysis of uterine tissue indicate that Cenpb-null mice lose reproductive fitness over a number of generations due to a significant reduction in endometrial glands. These results suggest that Cenpb may play an important function in the short- and long-term maintenance of uterine integrity.

Animals↗
PubMed2000

Survivin and the inner centromere protein INCENP show similar cell-cycle localization and gene knockout phenotype.

BACKGROUND: Survivin is a mammalian protein that carries a motif typical of the inhibitor of apoptosis (IAP)proteins, first identified in baculoviruses. Although baculoviral IAP proteins regulate cell death, the yeast Survivin homolog Bir1 is involved in cell division. To determine the function of Survivin in mammals, we analyzed the pattern of localization of Survivin protein during the cell cycle, and deleted its gene by homologous recombination in mice. RESULTS: In human cells, Survivin appeared first on centromeres bound to a novel para-polar axis during prophase/metaphase, relocated to the spindle midzone during anaphase/telophase, and disappeared at the end of telophase. In the mouse, Survivin was required for mitosis during development. Null embryos showed disrupted microtubule formation, became polyploid, and failed to survive beyond 4.5days post coitum. This phenotype, and the cell-cycle localization of Survivin, resembled closely those of INCENP. Because the yeast homolog of INCENP, Sli15, regulates the Aurora kinase homolog Ipl1p, and the yeast Survivin homolog Bir1 binds to Ndc10p, a substrate of Ipl1p, yeast Survivin, INCENP and Aurora homologs function in concert during cell division. CONCLUSIONS: In vertebrates, Survivin and INCENP have related roles in mitosis, coordinating events such as microtubule organization, cleavage-furrow formation and cytokinesis. Like their yeast homologs Bir1 and Sli15, they may also act together with the Aurora kinase.

Amino Acid Sequence↗
PubMed2000

Bub3 gene disruption in mice reveals essential mitotic spindle checkpoint function during early embryogenesis.

Bub3 is a conserved component of the mitotic spindle assembly complex. The protein is essential for early development in Bub3 gene-disrupted mice, evident from their failure to survive beyond day 6.5-7.5 postcoitus (pc). Bub3 null embryos appear normal up to day 3.5 pc but accumulate mitotic errors from days 4.5-6.5 pc in the form of micronuclei, chromatin bridging, lagging chromosomes, and irregular nuclear morphology. Null embryos treated with a spindle-depolymerising agent fail to arrest in metaphase and show an increase in mitotic disarray. The results confirm Bub3 as a component of the essential spindle checkpoint pathway that operates during early embryogenesis.

Animals↗
PubMed2000

Early disruption of centromeric chromatin organization in centromere protein A (Cenpa) null mice.

Centromere protein A (Cenpa for mouse, CENP-A for other species) is a histone H3-like protein that is thought to be involved in the nucleosomal packaging of centromeric DNA. Using gene targeting, we have disrupted the mouse Cenpa gene and demonstrated that the gene is essential. Heterozygous mice are healthy and fertile whereas null mutants fail to survive beyond 6.5 days postconception. Affected embryos show severe mitotic problems, including micronuclei and macronuclei formation, nuclear bridging and blebbing, and chromatin fragmentation and hypercondensation. Immunofluorescence analysis of interphase cells at day 5.5 reveals complete Cenpa depletion, diffuse Cenpb foci, absence of discrete Cenpc signal on centromeres, and dispersion of Cenpb and Cenpc throughout the nucleus. These results suggest that Cenpa is essential for kinetochore targeting of Cenpc and plays an early role in organizing centromeric chromatin at interphase. The evidence is consistent with the proposal of a critical epigenetic function for CENP-A in marking a chromosomal region for centromere formation.

Animals↗
PubMed2000

Uterine dysfunction and genetic modifiers in centromere protein B-deficient mice.

Centromere protein B (CENP-B) binds constitutively to mammalian centromere repeat DNA and is highly conserved between humans and mouse. Cenpb null mice appear normal but have lower body and testis weights. We demonstrate here that testis-weight reduction is seen in male null mice generated on three different genetic backgrounds (denoted R1, W9.5, and C57), whereas body-weight reduction is dependent on the genetic background as well as the gender of the animals. In addition, Cenpb null females show 31%, 33%, and 44% reduced uterine weights on the R1, W9.5, and C57 backgrounds, respectively. Production of "revertant" mice lacking the targeted frameshift mutation but not the other components of the targeting construct corrected these differences, indicating that the observed phenotype is attributable to Cenpb gene disruption rather than a neighbouring gene effect induced by the targeting construct. The R1 and W9.5 Cenpb null females are reproductively competent but show age-dependent reproductive deterioration leading to a complete breakdown at or before 9 months of age. Reproductive dysfunction is much more severe in the C57 background as Cenpb null females are totally incompetent or are capable of producing no more than one litter. These results implicate a further genetic modifier effect on female reproductive performance. Histology of the uterus reveals normal myometrium and endometrium but grossly disrupted luminal and glandular epithelium. Tissue in situ hybridization demonstrates high Cenpb expression in the uterine epithelium of wild-type animals. This study details the first significant phenotype of Cenpb gene disruption and suggests an important role of Cenpb in uterine morphogenesis and function that may have direct implications for human reproductive pathology.

Animals↗
PubMed1999

Defective chromosome segregation, microtubule bundling and nuclear bridging in inner centromere protein gene (Incenp)-disrupted mice.

INCENP is a chromosomal passenger protein which relocates from the centromere to thel spindle midzone during the metaphase-anaphase transition, ultimately being discarded in the cell midbody at the completion of cytokinesis. Using homologous recombination, we have generated Incenp gene-targeted heterozygous mice that are phenotypically indistinguishable from their wild-type littermates. Intercrossing the hetero-zygotes results in no live-born homozygous Incenp -disrupted progeny, indicating an early lethality. Day 3.5 affected pre-implantation embryos contain large, morphologically abnormal cells that fail to fully develop a blastocoel cavity or thrive in utero and in culture. Chromatin and tubulin immunocytochemical stainings of these and day 2.5 affected embryos reveal a high mitotic index, no discernible metaphase or anaphase stages, complete absence of midbodies, micronuclei formation, morphologically irregular macronuclei with large chromosome complements, multipolar mitotic configurations, binucleated cells, internuclear bridges and abnormal spindle bundling. The phenotype is consistent with a defect in the modulation of microtubule dynamics, severely affecting chromosome segregation and resulting in poorly resolved chromatin masses, aberrant karyokinesis and internuclear bridge formation. These latter occurrences could pose a physical barrier blocking cytokinesis.

Animals↗
PubMed1998

Centromere protein B null mice are mitotically and meiotically normal but have lower body and testis weights.

CENP-B is a constitutive centromere DNA-binding protein that is conserved in a number of mammalian species and in yeast. Despite this conservation, earlier cytological and indirect experimental studies have provided conflicting evidence concerning the role of this protein in mitosis. The requirement of this protein in meiosis has also not previously been described. To resolve these uncertainties, we used targeted disruption of the Cenpb gene in mouse to study the functional significance of this protein in mitosis and meiosis. Male and female Cenpb null mice have normal body weights at birth and at weaning, but these subsequently lag behind those of the heterozygous and wild-type animals. The weight and sperm content of the testes of Cenpb null mice are also significantly decreased. Otherwise, the animals appear developmentally and reproductively normal. Cytogenetic fluorescence-activated cell sorting and histological analyses of somatic and germline tissues revealed no abnormality. These results indicate that Cenpb is not essential for mitosis or meiosis, although the observed weight reduction raises the possibility that Cenpb deficiency may subtly affect some aspects of centromere assembly and function, and result in reduced rate of cell cycle progression, efficiency of microtubule capture, and/or chromosome movement. A model for a functional redundancy of this protein is presented.

Animals↗
PubMed1998

Targeted disruption of mouse centromere protein C gene leads to mitotic disarray and early embryo death.

Centromere protein C (CENPC) is a key protein that has been localized to the inner kinetochore plate of active mammalian centromeres. Using gene targeting techniques, we have disrupted the mouse Cenpc gene and shown that the gene is essential for normal mouse embryonic development. Heterozygous mice carrying one functional copy of the gene are healthy and fertile, whereas homozygous embryos fail to thrive. In these embryos, mitotic arrest and gross morphological degeneration become apparent as early as the morula stage of development. The degenerating embryos demonstrate highly irregular cell and nuclear morphologies, including the presence of a large number of micronuclei. Mitotic chromosomes of these embryos display a scattered and often highly condensed configuration and do not segregate in an ordered fashion. These results describing the phenotype of the mutant mouse embryos indicate that CENPC has a direct role in the mitotic progression from metaphase to anaphase.

Animals↗
PubMed1998

Genetic mapping of mouse centromere protein (Incenp and Cenpe) genes.

Inner centromere protein (INCENP) and centromere protein E (CENPE) are two functionally important proteins of the higher eukaryotic centromere. Using a mouse Incenp genomic DNA and a mouse Cenpe cDNA to analyze recombinant inbred mouse sets, as well as interspecific backcross panels, we have mapped these genes to the proximal regions of mouse Chromosomes 19 and 6, respectively. Comparison of Cenpe and human CENPE, which maps to chromosome region 4q24-->q25, has further identified a new region of homology between the two species.

Animals↗
PubMed1997

Chromosomal localization of mouse Cenpa gene.

Using a previously isolated mouse centromere protein A (Cenpa) probe, we have localized the gene to the proximal region of mouse Chromosome 5, between the known Il6 and Yes1 loci near [Adra2C-D5H4S43-Hdh]. Comparison of this localization with that of human CENPA, which maps to chromosome 2, is consistent with the presence of a new region of conserved synteny between the two species.

Animals↗
PubMed1995

A mutation in the epidermal growth factor receptor in waved-2 mice has a profound effect on receptor biochemistry that results in impaired lactation.

The mutant mouse waved-2 (wa-2) is strikingly similar to transforming growth factor alpha-deficient mice generated by gene targeting in embryonic stem cells. We confirm that wa-2 is a point mutation (T-->G resulting in a valine-->glycine substitution at residue 743) in the gene encoding the epidermal growth factor (EGF) receptor. wa-2 fibroblastic cells lack high-affinity binding sites for EGF, and the rate of internalization of EGF is retarded. Although the tyrosine kinase activity of wa-2 EGF receptors is significantly impaired, NIH 3T3 cells lacking endogenous EGF receptors but overexpressing recombinant wa-2 EGF receptor cDNA are mitogenically responsive to EGF. While young and adult wa-2 mice are healthy and fertile, 35% of wa-2 mice born of homozygous wa-2 mothers die of malnutrition because of impaired maternal lactation.

3T3 Cells↗
PubMed1994

Mice lacking granulocyte colony-stimulating factor have chronic neutropenia, granulocyte and macrophage progenitor cell deficiency, and impaired neutrophil mobilization.

Mice lacking granulocyte colony-stimulating factor (G-CSF) were generated by targeted disruption of the G-CSF gene in embryonal stem cells. G-CSF-deficient mice (genotype G-CSF-/-) are viable, fertile, and superficially healthy, but have a chronic neutropenia. Peripheral blood neutrophil levels were 20% to 30% of wild-type mice (genotype G-CSF+/+) and mice heterozygous for the null mutation had intermediate neutrophil levels, suggesting a gene-dosage effect. In the marrow of G-CSF-/- mice, granulopoietic precursor cells were reduced by 50% and there were reduced levels of granulocyte, macrophage, and blast progenitor cells. Despite G-CSF deficiency, mature neutrophils were still present in the blood and marrow, indicating that other factors can support neutrophil production in vivo. G-CSF-/- mice had reduced numbers of neutrophils available for rapid mobilization into the circulation by a single dose of G-CSF. G-CSF administration reversed the granulopoietic defect of G-CSF-/- mice. One day of G-CSF administration to G-CSF-/- mice elevated circulating neutrophil levels to normal, and after 4 days of G-CSF administration, G-CSF+/+ and G-CSF-/- marrows were morphologically indistinguishable. G-CSF-/- mice had a markedly impaired ability to control infection with Listeria monocytogenes, with diminished neutrophil and delayed monocyte increases in the blood and reduced infection-driven granulopoiesis. Collectively, these observations indicate that G-CSF is indispensible for maintaining the normal quantitative balance of neutrophil production during "steady-state" granulopoiesis in vivo and also implicate G-CSF in "emergency" granulopoiesis during infections.

Animals↗
PubMed1994

Insights into the physiology of TGF alpha and signaling through the EGF receptor revealed by gene targeting and acts of nature.

Transforming growth factor alpha (TGF alpha) is one of a group of structurally-related growth factors (the epidermal growth factor family of ligands) that interact with one or other members of the epidermal growth factor family of protein tyrosine kinase receptors (EGF-R's). A number of excellent reviews detailing our knowledge of this area have been recently published (Carpenter and Wahl, 1991; Derynck, 1992; Prigent and Lemoine, 1992). Rather than add to their number, this review focuses on new insights into the importance of TGF alpha and signaling through the EGF receptor considered in the context of the laboratory mouse. The new information has emerged from analysis of mutant mice generated either by classical gene targeting in embryonic stem (ES) cells or by accidents of nature. In addition to their intrinsic interest, these mice are proving invaluable in determining the importance of EGF receptor signaling in wound healing and as a contributing factor in the conversion of a normal cell into its tumorigenic counterpart.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1993

Mice with a null mutation of the TGF alpha gene have abnormal skin architecture, wavy hair, and curly whiskers and often develop corneal inflammation.

Mice homozygous for a disrupted transforming growth factor alpha (TGF alpha) gene are healthy and fertile, although some older mice show evidence of corneal inflammation. In contrast with TGF alpha +/- and +/+ animals, TGF alpha -/- mice have a pronounced waviness of the coat. Histological examination of the skin from TGF alpha -/- mice reveals a dramatic derangement of hair follicles. Mice with a disrupted TGF alpha gene also have curly whiskers, first evident on the day of birth. The phenotype of TGF alpha -/- mice is remarkably similar to that of the mouse mutant waved-1 (wa-1). Offspring resulting from crosses between TGF alpha -/- and wa-1 mice display the curly whisker-coat phenotype, indicating that the basis of the wa-1 phenotype is a mutation in the TGF alpha gene. These observations suggest that TGF alpha plays a pivotal role in determining skin architecture and in regulating hair development.

Animals↗