RESUMO
PURPOSE: Our aim was to generate and phenotypically characterize a transgenic mouse line expressing a constitutively active variant of the transcription regulatory protein serum response factor (SRF), namely the SRF-VP16 protein. This new mouse strain has been registered under the designation Gt(ROSA)26Sor(tm1(SRF-VP16)Antu). We found phenotypic changes upon ectopic expression of SRF-VP16, especially in the mouse retina. METHODS: Using homologous recombination, we integrated an SRF-VP16 conditional (i.e., "flox-STOP" repressed) expression transgene into the Rosa26 locus of murine embryonic stem (ES) cells. These engineered ES cells were used to derive the Gt(ROSA)26Sor(tm1(SRF-VP16)Antu) mouse strain. Semiquantitative real-time PCR was used to determine expression of the SRF-VP16 transgene at the mRNA level, both in young (P20 and P30) and adult (six months old) Gt(ROSA)26Sor(tm1(SRF-VP16)Antu) mice. We also investigated the transcript levels of endogenous Srf and several SRF target genes. Retinal function was tested by electroretinography in both young and adult mice. Morphological abnormalities could be visualized by hematoxylin and eosin staining of sectioned, paraffin-embedded eye tissue samples. Scanning-laser ophthalmoscopy was used to investigate retinal vascularization and degeneration in adult mice. RESULTS: We show that the SRF-VP16 mRNA is expressed to a low but significant degree in the retinas of young and adult animals of the Gt(ROSA)26Sor(tm1(SRF-VP16)Antu) mouse strain, even in the absence of Cre-mediated deletion of the "flox-STOP" cassette. In the retinas of these transgenic mice, endogenous Srf displays elevated transcript levels. Ectopic retinal expression of constitutively active SRF-VP16 is correlated with the malfunction of retinal neurons in both heterozygous and homozygous animals of both age groups (P20 and adult). Additionally, mislamination of retinal cell layers and cellular rosette formations are found in retinas of both heterozygous and homozygous animals of young age. In homozygous individuals, however, the cellular rosettes are more widespread over the fundus. At adult age, retinas both from animals that are heterozygous and homozygous for the floxSTOP/SRF-VP16 transgene display severe degeneration, mainly of the photoreceptor cell layer. Wild-type age-matched littermates, however, do not show any degeneration. The severity of the observed effects correlates with dosage of the transgene. CONCLUSIONS: This is the first report suggesting an influence of the transcription factor SRF on the development and function of the murine retina. Ectopic SRF-VP16 mRNA expression in the retinas of young animals is correlated with photoreceptor layer mislamination and impaired retinal function. At an advanced age of six months, degenerative processes are detected in SRF-VP16 transgenic retinas accompanied by impaired retinal function. The Gt(ROSA)26Sor(tm1(SRF-VP16)Antu) mouse strain represents a genetic SRF gain-of-function mouse model that will complement the current SRF loss-of-function models. It promises to provide new insight into the hitherto poorly defined role of SRF in retinal development and function, including potential contributions to ophthalmologic disorders. Furthermore, using conditional Cre-mediated activation of SRF-VP16, the described mouse strain will enable assessment of the impact of dysregulated SRF activity on the physiologic functions of various other organs.
Assuntos
Camundongos Mutantes/genética , Retina/patologia , Degeneração Retiniana/genética , Fator de Resposta Sérica , Animais , Modelos Animais de Doenças , Eletrorretinografia , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Amarelo de Eosina-(YS) , Dosagem de Genes , Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Hematoxilina , Proteína Vmw65 do Vírus do Herpes Simples/genética , Proteína Vmw65 do Vírus do Herpes Simples/metabolismo , Camundongos , Camundongos Mutantes/crescimento & desenvolvimento , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Retina/metabolismo , Degeneração Retiniana/patologia , Fator de Resposta Sérica/genética , Fator de Resposta Sérica/metabolismo , Transfecção , TransgenesRESUMO
The German Mouse Clinic (GMC) is a large scale phenotyping center where mouse mutant lines are analyzed in a standardized and comprehensive way. The result is an almost complete picture of the phenotype of a mouse mutant line--a systemic view. At the GMC, expert scientists from various fields of mouse research work in close cooperation with clinicians side by side at one location. The phenotype screens comprise the following areas: allergy, behavior, clinical chemistry, cardiovascular analyses, dysmorphology, bone and cartilage, energy metabolism, eye and vision, host-pathogen interactions, immunology, lung function, molecular phenotyping, neurology, nociception, steroid metabolism, and pathology. The German Mouse Clinic is an open access platform that offers a collaboration-based phenotyping to the scientific community (www.mouseclinic.de). More than 80 mutant lines have been analyzed in a primary screen for 320 parameters, and for 95% of the mutant lines we have found new or additional phenotypes that were not associated with the mouse line before. Our data contributed to the association of mutant mouse lines to the corresponding human disease. In addition, the systemic phenotype analysis accounts for pleiotropic gene functions and refines previous phenotypic characterizations. This is an important basis for the analysis of underlying disease mechanisms. We are currently setting up a platform that will include environmental challenge tests to decipher genome-environmental interactions in the areas nutrition, exercise, air, stress and infection with different standardized experiments. This will help us to identify genetic predispositions as susceptibility factors for environmental influences.
Assuntos
Pesquisa Biomédica/métodos , Modelos Animais de Doenças , Camundongos Mutantes/genética , Fenótipo , Criação de Animais Domésticos , Animais , Pesquisa Biomédica/normas , Alemanha , Camundongos , Camundongos Mutantes/crescimento & desenvolvimento , Controle de QualidadeRESUMO
Tail DNA genotyping of fetal and neonatal mice from C/EBPbeta heterozygous parents was performed to determine whether the decreased number of surviving C/EBPbeta mutants was caused by prenatal or postnatal death. Eighty-four 3-week-old mice born of heterozygous parents had significantly lower numbers of C/EBPbeta-deficient offspring than the expected Mendelian ratio (29.8%+/+, 65.5%+/-, 4.76%-/-, P<0.05). The genotypes of 72 fetal mice from intercrossed heterozygotes showed approximately the expected 1:2:1 Mendelian ratio (18.1% +/+, 52.8% +/-, 29.2% -/-, P>0.05). No difference in the proportions by sex could be detected in these perinates. This data indicates that C/EBPbeta-deficient mice have unknown lethal problems between the embryonic stage and weaning.
Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/deficiência , Genes Letais/fisiologia , Camundongos Mutantes/crescimento & desenvolvimento , Animais , Proteína beta Intensificadora de Ligação a CCAAT/genética , Feminino , Genótipo , Masculino , Camundongos , Camundongos Mutantes/genéticaRESUMO
Regulation of telomere length maintenance and capping are a critical cell functions in both normal and tumor cells. Tankyrase 2 (Tnks2) is a poly(ADP-ribose) polymerase (PARP) that has been shown to modify itself and TRF1, a telomere-binding protein. We show here by overexpression studies that tankyrase 2, like its closely related homolog tankyrase 1, can function as a positive regulator of telomere length in human cells, dependent on its catalytic PARP activity. To study the role of Tnks2 in vivo, we generated mice with the Tnks2 PARP domain deleted. These mice are viable and fertile but display a growth retardation phenotype. Telomere analysis by quantitative fluorescence in situ hybridization (FISH), flow-FISH, and restriction fragment analysis showed no change in telomere length or telomere capping in these mice. To determine the requirement for Tnks2 in long-term maintenance of telomeres, we generated embryonic stem cells with the Tnks2 PARP domain deleted and observed no change, even upon prolonged growth, in telomere length or telomere capping. Together, these results suggest that Tnks2 has a role in normal growth and development but is not essential for telomere length maintenance or telomere capping in mice.
Assuntos
Crescimento/genética , Tanquirases/genética , Tanquirases/metabolismo , Telômero/fisiologia , Animais , Tamanho Corporal/genética , Células Cultivadas , Feminino , Humanos , Masculino , Camundongos , Camundongos Mutantes/crescimento & desenvolvimento , Poli(ADP-Ribose) Polimerases/metabolismo , Estrutura Terciária de Proteína , Valores de Referência , Células-Tronco/citologia , Células-Tronco/fisiologiaRESUMO
The Splotch mouse, a Pax 3 mutation, represents a model of Waardenburg syndrome I. We show that the homozygous Splotch mutation (Sp(2H)) is associated with severe defects that prevent the formation of the cochlea and vestibulo-cochlear ganglion. To clarify the role of Pax 3 in inner ear formation, we examined the expression of polysialic acid (PSA) associated with neural cell adhesion molecule (NCAM). In accordance with the occurrence of phenotypic abnormalities, PSA NCAM was expressed early in otocyst development in the otic epithelium and the vestibulo-cochlear anlage. During the period of vestibular and cochlear ganglia formation, PSA NCAM expression was decreased. In the late phase of embryonic development, the expression of calcium binding proteins (S100) in the vestibulo-cochlear ganglion was also decreased. Minor differences in S100 immunostaining were found postnatally between the cochleas of heterozygous and wild type animals.
Assuntos
Proteínas de Ligação a DNA/genética , Orelha Interna/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento , Camundongos Mutantes/crescimento & desenvolvimento , Mutação , Fatores de Transcrição/genética , Fatores Etários , Animais , Animais Recém-Nascidos , Contagem de Células/métodos , Morte Celular/genética , Proteínas de Ligação a DNA/metabolismo , Orelha Interna/anatomia & histologia , Orelha Interna/metabolismo , Embrião de Mamíferos , Imuno-Histoquímica/métodos , Marcação In Situ das Extremidades Cortadas , Camundongos , Camundongos Mutantes/anatomia & histologia , Camundongos Mutantes/metabolismo , Molécula L1 de Adesão de Célula Nervosa/metabolismo , Fator de Transcrição PAX3 , Fatores de Transcrição Box Pareados , Proteínas S100/metabolismo , Ácidos Siálicos/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Toll receptors in Drosophila contribute to host defense and establish the body plan. Mammalian homologues of Toll, the Toll-like receptors (TLRs), are thought to function only in host defense. Here, we report that mice harboring mutations in TLR4 or in CD14, a co-receptor for TLR4, have an "ideal" body plan consisting of increased bone mineral content, density, and size as well as decreased body fat. These mutant mice live long lives, have normal activity and fertility, and show no evidence of infection. Unlike many strains of caged wild-type mice, they do not become obese. Although all mice continue to gain body fat, bone content, and overall weight, the difference in bone content and body fat between mutant and wild-type mice increases with age. Thus, defects in TLR4/CD14 complex generate an "Adonis" phenotype, characterized by this ideal body type, and this function could potentially be exploited for the treatment of osteoporosis and obesity.
Assuntos
Constituição Corporal/genética , Padronização Corporal/genética , Densidade Óssea/genética , Receptores de Lipopolissacarídeos/fisiologia , Camundongos Mutantes/genética , Receptores de Superfície Celular/deficiência , Magreza/genética , Tecido Adiposo/anatomia & histologia , Envelhecimento , Animais , Osso e Ossos/anatomia & histologia , Feminino , Fertilidade/genética , Deleção de Genes , Receptores de Lipopolissacarídeos/genética , Longevidade/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Mutantes/anatomia & histologia , Camundongos Mutantes/crescimento & desenvolvimento , Atividade Motora/genética , Obesidade/genética , Osteoporose/genética , Fenótipo , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/fisiologia , Deleção de Sequência , Receptor 4 Toll-Like , Suporte de CargaRESUMO
To gain insight into the pathways by which caloric restriction (CR) slows aging, gene expression levels were assessed for each of 2,352 genes in the livers of 9-month-old CR and control mice. A total of 352 genes were found to be significantly increased or decreased by CR. The distribution of affected genes among functional classes was similar to the distribution of genes within the test set. Surprisingly, a disruption or knockout of the gene for the GH receptor (GHR-KO), which also produces life extension, had a much smaller effect on gene expression, with no more than 10 genes meeting the selection criterion. There was, however, an interaction between the GHR-KO mutation and the CR diet: the effects of CR on gene expression were significantly lower in GHR-KO mice than in control mice. Of the 352 genes altered significantly by CR, 29 had shown a significant and parallel alteration in expression in a previous study of liver gene expression that compared mice of the long-lived Snell dwarf stock (dw/dw) to controls. These 29 genes, altered both by CR and in dwarf mice, provide a list of biochemical features common to both models of delayed aging, and thus merit confirmation and more detailed study.
Assuntos
Dieta Redutora , Regulação da Expressão Gênica no Desenvolvimento , Longevidade/fisiologia , Camundongos Mutantes/crescimento & desenvolvimento , Animais , Ingestão de Energia , Camundongos , Proteínas/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
In bronx waltzer mouse mutants, inner hair cells die at an early stage in their development, from around 17.5 days of gestation onwards. In contrast, outer hair cells appear to develop normally. Vestibular hair cells also degenerate, but the earliest signs of vestibular abnormalities have not yet been described. We looked at prenatal and early postnatal stages of vestibular development by scanning electron microscopy in the mutants, and established that vestibular hair cells (types I and II) never reach beyond the middle stages of differentiation (at least up to P2) and instead show signs of degeneration. Thus, it appears that the bronx waltzer gene product is required for the continued survival and differentiation of inner and vestibular hair cells past a set point in their development.
Assuntos
Células Ciliadas Auditivas/crescimento & desenvolvimento , Células Ciliadas Auditivas/patologia , Camundongos Mutantes/anatomia & histologia , Camundongos Mutantes/crescimento & desenvolvimento , Vestíbulo do Labirinto/crescimento & desenvolvimento , Vestíbulo do Labirinto/patologia , Animais , Animais Recém-Nascidos , Feminino , Genótipo , Idade Gestacional , Células Ciliadas Auditivas/embriologia , Células Ciliadas Auditivas Internas/embriologia , Células Ciliadas Auditivas Internas/crescimento & desenvolvimento , Células Ciliadas Auditivas Internas/patologia , Masculino , Camundongos , Camundongos Mutantes/embriologia , Microscopia Eletrônica de Varredura , Gravidez , Vestíbulo do Labirinto/embriologiaRESUMO
ATM, the gene mutated in the human immunodeficiency disorder ataxia-telangiectasia (A-T), plays a central role in recognizing ionizing radiation damage in DNA and in controlling several cell cycle checkpoints. We describe here a murine model in which a nine-nucleotide in-frame deletion has been introduced into the Atm gene by homologous recombination followed by removal of the selectable marker cassette by Cre-loxP site-specific, recombination-mediated excision. This mouse, Atm-DeltaSRI, was designed as a model of one of the most common deletion mutations (7636del9) found in A-T patients. The murine Atm deletion results in the loss of three amino acid residues (SRI; 2556-2558) but produces near full-length detectable Atm protein that lacks protein kinase activity. Radiosensitivity was observed in Atm-DeltaSRI mice, whereas the immunological profile of these mice showed greater heterogeneity of T-cell subsets than observed in Atm(-/-) mice. The life span of Atm-DeltaSRI mice was significantly longer than that of Atm(-/-) mice when maintained under nonspecific pathogen-free conditions. This can be accounted for by a lower incidence of thymic lymphomas in Atm-DeltaSRI mice up to 40 weeks, after which time the animals died of other causes. The thymic lymphomas in Atm-DeltaSRI mice were characterized by extensive apoptosis, which appears to be attributable to an increased number of cells expressing Fas ligand. A variety of other tumors including B-cell lymphomas, sarcomas, and carcinomas not seen in Atm(-/-) mice were observed in older Atm-DeltaSRI animals. Thus, expression of mutant protein in Atm-DeltaSRI knock-in mice gives rise to a discernibly different phenotype to Atm(-/-) mice, which may account for the heterogeneity seen in A-T patients with different mutations.
Assuntos
Camundongos Mutantes/genética , Proteínas Serina-Treonina Quinases/genética , Deleção de Sequência , Animais , Apoptose/genética , Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia , Sequência de Bases , Proteínas de Ciclo Celular , Cruzamentos Genéticos , DNA/genética , Proteínas de Ligação a DNA , Feminino , Humanos , Linfoma/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Mutantes/crescimento & desenvolvimento , Camundongos Mutantes/imunologia , Mutagênese Sítio-Dirigida , Fenótipo , Neoplasias do Timo/genética , Proteínas Supressoras de Tumor , Regulação para CimaRESUMO
It has been reported that the arrival of primary olfactory axons is required to induce the development of the olfactory bulb (OB). On the other hand, the Sey(Neu)/Sey(Neu) mutant mouse (Small eye) has been previously described as a model for the absence of olfactory bulbs, owing to the lack of olfactory epithelium (OE). In the present report, we take advantage of this mutant and study a neural structure in the rostral pole of the telencephalon that phenotypically resembles the prospective OB. We named this formation olfactory bulb-like structure (OBLS). We also report the occurrence, in the mutants, of small epithelial vesicles in the malformed craneofacial pits, resembling an atrophic OE, although a mature olfactory nerve was not identified. Axonal tracing, birthdating, immunohistochemistry, and in situ hybridization using antibodies and probes expressed in the olfactory system, indicated that two distinct structures observed in the OBLS correspond to the main and accessory olfactory bulbs of the control mouse. We propose that the OBLS has developed independently of the external influences exerted by the olfactory nerve. The presence of a prospective OB in the mutants, without intervening olfactory fibers, suggests that intrinsic factors could define brain territories even in absence of the proper afferent innervation. The intrinsic mechanisms and environmental cues in the telencephalon could be sufficient to promote axonogenesis in the projection neurons of the OB and guide their axons in a lateral prospective tract, in the absence of olfactory axons.
Assuntos
Proteínas de Homeodomínio/genética , Camundongos Mutantes/embriologia , Camundongos Mutantes/crescimento & desenvolvimento , Vias Neurais/embriologia , Vias Neurais/crescimento & desenvolvimento , Bulbo Olfatório/embriologia , Bulbo Olfatório/crescimento & desenvolvimento , Animais , Biomarcadores/análise , Divisão Celular/genética , Embrião de Mamíferos/citologia , Embrião de Mamíferos/embriologia , Embrião de Mamíferos/metabolismo , Proteínas do Olho , Camundongos , Camundongos Mutantes/anatomia & histologia , Camundongos Mutantes/genética , Proteínas do Tecido Nervoso/genética , Vias Neurais/citologia , Neurônios/citologia , Neurônios/metabolismo , Neuropilina-1 , Bulbo Olfatório/citologia , Mucosa Olfatória/citologia , Mucosa Olfatória/embriologia , Mucosa Olfatória/crescimento & desenvolvimento , Fator de Transcrição PAX6 , Fatores de Transcrição Box Pareados , RNA Mensageiro/metabolismo , Receptores Imunológicos/genética , Proteínas Repressoras , Proteínas RoundaboutRESUMO
Nongenetic factors such as nutrition modulate the effects of genes responsible for overgrowth in animals. The goal of this study was to examine the importance of genotype x diet interactions on the effects of a major locus that regulates growth in the mouse. We have examined the phenotype of high growth (hg), a partially recessive autosomal locus that increases growth rate and mature body size. C57BL/6J (C57) and congenic C57BL/6J-hg/hg (HG) mice were fed three experimental diets differing in protein and energy content from 3 to 12 wk of age. HG mice grew faster and were, on average, 51% heavier than C57 at 12 wk of age. Feed intake was higher in HG mice but proportional to the increase in body weight. The magnitude of the differences in body size and composition between lines depended on the interaction between genotype and the protein/energy ratio of the diet. In C57, the diets modified the level of fatness without changing adult lean mass. However, in HG the diets differentially affected both linear growth and body composition. In general, HG had higher plasma levels of insulin-like growth factor I at 3 and 12 wk than C57. Plasma insulin did not differ between lines, but leptin was higher for C57 mice fed a high-energy diet. These results show that the effects of hg on growth are modulated by diet composition. Therefore, this mutation could be a valuable model with which to study the genetic and nutritional aspects of overgrowth disorders.
Assuntos
Composição Corporal/fisiologia , Alimentos Formulados , Transtornos do Crescimento/metabolismo , Camundongos Mutantes/crescimento & desenvolvimento , Aumento de Peso/fisiologia , Animais , Glicemia , Composição Corporal/genética , Proteínas Alimentares/metabolismo , Modelos Animais de Doenças , Ingestão de Alimentos , Ingestão de Energia/fisiologia , Transtornos do Crescimento/sangue , Transtornos do Crescimento/genética , Homozigoto , Insulina/sangue , Fator de Crescimento Insulin-Like I/metabolismo , Leptina/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes/genética , Camundongos Mutantes/metabolismo , Tamanho do Órgão/fisiologia , Fenótipo , Aumento de Peso/genéticaRESUMO
The actions of insulin-like growth factor-I (IGF-I) are modulated by IGF binding proteins (IGFBPs). The effects of IGFBP-1 in vivo are insufficiently known, with respect to inhibitory or stimulatory actions on IGF-induced growth of specific organs. Therefore, we studied the effects of IGFBP-1 on IGF-I-induced somatic and organ growth in pituitary-deficient Snell dwarf mice. Human GH, IGF-I, IGFBP-1, and a preequilibrated combination of equimolar amounts of IGF-I and IGFBP-1 were administered sc during 4 weeks. Treatment with IGF-I alone induced a significant increase in body length (108% of control) and weight (112%) as well as an increase in weight of the submandibular salivary glands (135%), kidneys (124%), femoral muscles (111%), testes (129%), and spleen (126%) compared with saline-treated controls. IGFBP-1 alone induced a significant increase in weight of the kidneys (152% of control). Coadministration of IGF-I with IGFBP-1 neutralized the stimulating effects of IGF-I on body length and weight as well as on the femoral muscles and testes. In contrast, the weights of the submandibular salivary glands (143%) were not significantly different from those of IGF-I-treated animals, whereas the weights of the kidneys (171%) and spleen (156%) were significantly increased compared with IGF-I-treated mice. The effect of IGFBP-1 plus IGF-I on kidney weight was not significantly greater than the effect of IGFBP-1 alone. Western ligand blotting showed induction of the IGFBP-3 doublet as well as IGFBPs with molecular masses of 24 kDa, most probably IGFBP-4, by human GH, IGF-I alone, and IGF-I in combination with IGFBP-1. Our data show that coadministration of IGFBP-1 inhibits IGF-I-induced body growth of GH-deficient mice but significantly stimulates the growth promoting effects of IGF-I on the kidneys and the spleen. These data warrant further investigation because differences in concentrations of IGFBP-1 occurring in vivo may influence IGF-I-induced anabolic processes.
Assuntos
Peso Corporal/efeitos dos fármacos , Nanismo/fisiopatologia , Proteína 1 de Ligação a Fator de Crescimento Semelhante à Insulina/farmacologia , Fator de Crescimento Insulin-Like I/farmacologia , Rim/efeitos dos fármacos , Rim/crescimento & desenvolvimento , Animais , Glicemia/análise , Nanismo/genética , Nanismo/patologia , Glândulas Endócrinas/efeitos dos fármacos , Glândulas Endócrinas/crescimento & desenvolvimento , Humanos , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/sangue , Camundongos , Camundongos Mutantes/crescimento & desenvolvimento , Somatomedinas/análiseRESUMO
The three mammalian Raf serine/threonine protein kinases mediate the transduction of proliferative and differentiative signals from a variety of cell surface receptors to the nucleus. We report here that Craf-1 is essential for mouse development, as its mutation results in embryonic lethality. Developmental defects are found in mutant placentas as well as in the skin and in the lungs of mutant embryos. Craf-1 mutants also display a generalized growth retardation which is consistent with the ubiquitous expression of Craf-1 and which could be due to the reduced proliferation of mutant cells. Interestingly, the time-point of embryonal death varies depending on the genetic background. This suggests that Craf-1-mediated signaling is affected by genetic background-specific alleles of other genes.
Assuntos
Camundongos Mutantes/crescimento & desenvolvimento , Camundongos Mutantes/genética , Mutação , Proteínas/genética , Proteínas/metabolismo , Anormalidades Múltiplas/genética , Animais , Sequência de Bases , Divisão Celular , Morte Fetal/genética , Fibroblastos , Regulação da Expressão Gênica no Desenvolvimento , Camundongos , Camundongos Endogâmicos , Camundongos Mutantes/embriologia , Dados de Sequência Molecular , Especificidade da Espécie , Fator 3 Associado a Receptor de TNFRESUMO
The cation-dependent and cation-independent mannose 6-phosphate receptors (CD- and CI-MPRs) bind the phosphomannosyl recognition marker of lysosomal hydrolases, but in mammals the latter also interacts with insulin-like growth factor II (IGF-II). While IGF signaling is mediated by the type 1 IGF receptor (IGF1R), the type 2 receptor (IGF2R/CI-MPR) serves IGF-II turnover. Mouse mutants inheriting maternally a targeted disruption of the imprinted Igf2r gene, which is normally expressed only from the maternal allele, have increased serum and tissue levels of IGF-II and exhibit overgrowth (135% of normal birthweight) and generalized organomegaly, kinky tail, postaxial polydactyly, heart abnormalities, and edema. These mutants usually die perinatally, but a small minority can survive depending on genetic background and can occasionally reproduce, except for some females characterized by an imperforate vagina and hydrometrocolpos. Consistent with the hypothesis that lethality in the absence of IGF2R-mediated turnover is caused by excess of IGF-II overstimulating IGF1R, Igf2r mutants are completely rescued when they carry a second mutation eliminating either IGF-II or IGF1R. Normal embryonic development of the Igf1r/Igf2r double mutants, which differ from wild-type siblings only in the pattern of postnatal growth, appears to occur by signaling of IGF-II, being in excess, through a genetically identified unknown receptor, since triple mutants lacking IGF1R, IGF2R, and IGF-II are nonviable dwarfs (30% of normal size). In contrast with the Igf2r/Igf2 double mutants, mice lacking IGF2R/CI-MPR and CD-MPR survive in an IGF-II null background at a very low frequency and only for a few postnatal weeks, indicating that the mannose 6-phosphate-mediated lysosomal enzyme trafficking is essential for viability.
Assuntos
Animais Recém-Nascidos/genética , Genes Letais/genética , Camundongos Mutantes/genética , Receptor IGF Tipo 2/genética , Animais , Cruzamento/métodos , Cardiopatias Congênitas/genética , Cardiopatias Congênitas/patologia , Manosefosfatos/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Mutantes/crescimento & desenvolvimento , Mutação , Fenótipo , Receptor IGF Tipo 2/deficiência , Receptores de Somatomedina/deficiência , Receptores de Somatomedina/genética , Reprodução/genética , Taxa de SobrevidaRESUMO
IL-12 is a cytokine that can exert regulatory effects on T and NK cells and promote Th1 responses. To delineate further the physiologic role of IL-12 in immunity, mice deficient for this cytokine were generated. IL-12-deficient mice were impaired but not completely lacking in the ability to produce IFN gamma following endotoxin administration and to mount a Th1 response in vivo, as measured by antigen-induced IFN gamma secretion by immune lymph node cells in vitro. In contrast, secretion of IL-4 was enhanced, while proliferation and secretion of IL-2 and IL-10 were normal following antigen stimulation. DTH responses were significantly reduced in IL-12-deficient mice, but no defect in allogeneic CTL responses was observed. These results indicate that IL-12 plays an essential role in regulating IFN gamma production and in facilitating normal DTH responses. However, other phenomena associated with Th1 responses and cell-mediated immunity, i.e., IL-2 secretion and CTL generation, were not compromised in the absence of IL-12.
Assuntos
Interferon gama/biossíntese , Interleucina-12/deficiência , Interleucina-12/genética , Linfocinas/biossíntese , Camundongos Mutantes/imunologia , Animais , Sequência de Bases , Vetores Genéticos/imunologia , Células Matadoras Naturais/imunologia , Ativação Linfocitária/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Mutantes/crescimento & desenvolvimento , Camundongos Mutantes/metabolismo , Dados de Sequência Molecular , Células Th1/imunologia , Células Th1/metabolismoRESUMO
Mice were inoculated with 10 LD50 of venezuelan equine encephalitis virus and 1.5 mgs, daily of tetracycline. At the end of the experiment almost half of the group which had received virus and antibiotic were alive while those with virus only had all died
Assuntos
Animais , Camundongos , Camundongos Mutantes/crescimento & desenvolvimento , Tetraciclina/administração & dosagem , Tetraciclina/classificação , Tetraciclina/uso terapêuticoRESUMO
The immediate-early gene NGFI-B encodes an orphan nuclear receptor that binds DNA as a monomer and activates transcription through a canonical response element (NBRE). NGFI-B is expressed under basal conditions and in response to external stimuli in many mammalian tissues. In particular, NGFI-B expression is dramatically elevated in the adrenal cortex in response to stress and in Y1 adrenocortical cells in response to adrenocorticotropin. NGFI-B activates transcription through an NBRE of the gene encoding 21-hydroxylase (P450c21) in Y1 cells. Steroidogenic factor 1 (SF-1), a homolog of NGFI-B, also activates the P450c21 promoter. To examine the influence of these factors on P450c21 expression in vivo and the function of the hypothalamic-pituitary-adrenocortical axis as a whole, we generated NGFI-B (-/-) mice. These mice thrive and reproduce normally and maintain normal basal adrenocorticotropin, corticosterone, and P450c21 mRNA levels. In response to increases in adrenocorticotropin, NGFI-B (-/-) and wild-type mice demonstrated equivalent increases in serum corticosterone levels. Furthermore, and in contrast to in vitro results, no increases in P450c21 mRNA levels were observed in response to increases in adrenocorticotropin in NGFI-B (-/-) or wild-type mice. While SF-1 mRNA levels were not increased with increased steroidogenic demand, adrenal expression of Nurr1, a close homolog of NGFI-B, was induced to a greater extent by lipopolysaccharide in NGFI-B (-/-) mice than in wild-type mice. Finally, when the administration of dexamethasone for suppression was stopped, P450c21 mRNA and serum corticosterone levels recovered at the same rate in wild-type and NGFI-B (-/-) mice. Thus, while NGFI-B appears poised to affect the structure and function of the adrenal gland, the gland functions normally in its absence, suggesting that other factors, including Nurr1 and SF-1, are sufficient to drive P450c21 expression in mice and maintain normal steroidogenesis.
Assuntos
Córtex Suprarrenal/enzimologia , Proteínas de Ligação a DNA/metabolismo , Regulação Enzimológica da Expressão Gênica , Receptores de Esteroides/metabolismo , Esteroide 21-Hidroxilase/metabolismo , Fatores de Transcrição/metabolismo , Hormônio Adrenocorticotrópico/biossíntese , Hormônio Adrenocorticotrópico/genética , Hormônio Adrenocorticotrópico/farmacologia , Animais , Corticosterona/biossíntese , Corticosterona/sangue , Corticosterona/genética , Proteínas de Ligação a DNA/genética , Dexametasona/farmacologia , Fatores de Transcrição Fushi Tarazu , Proteínas de Homeodomínio , Sistema Hipotálamo-Hipofisário/fisiologia , Lipopolissacarídeos/farmacologia , Metirapona/farmacologia , Camundongos , Camundongos Mutantes/crescimento & desenvolvimento , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares , Sistema Hipófise-Suprarrenal/fisiologia , RNA Mensageiro/análise , Receptores Citoplasmáticos e Nucleares/genética , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptores de Esteroides/deficiência , Receptores de Esteroides/genética , Esteroide 21-Hidroxilase/genética , Fator Esteroidogênico 1 , Fatores de Transcrição/genética , Transcrição GênicaRESUMO
H1 histones bind to the linker DNA between nucleosome core particles and facilitate the folding of chromatin into a 30-nm fiber. Mice contain at least seven nonallelic subtypes of H1, including the somatic variants H1a through H1e, the testis-specific variant H1t, and the replacement linker histone H1(0). H1(0) accumulates in terminally differentiating cells from many lineages, at about the time when the cells cease dividing. To investigate the role of H1(0) in development, we have disrupted the single-copy H1(0) gene by homologous recombination in mouse embryonic stem cells. Mice homozygous for the mutation and completely lacking H1(0) mRNA and protein grew and reproduced normally and exhibited no anatomic or histologic abnormalities. Examination of tissues in which H1(0) is normally present at high levels also failed to reveal any abnormality in cell division patterns. Chromatin from H1(0)-deficient animals showed no significant change in the relative proportions of the other H1 subtypes or in the stoichiometry between linker histones and nucleosomes, suggesting that the other H1 histones can compensate for the deficiency in H1(0) by occupying sites that normally contain H1(0). Our results indicate that despite the unique properties and expression pattern of H1(0), its function is dispensable for normal mouse development.
Assuntos
Quimera , Variação Genética , Histonas/deficiência , Histonas/genética , Camundongos Mutantes/crescimento & desenvolvimento , Animais , Sequência de Bases , Cromatina/metabolismo , Primers do DNA , Embrião de Mamíferos , Feminino , Genótipo , Homozigoto , Rim/metabolismo , Fígado/metabolismo , Pulmão/metabolismo , Masculino , Camundongos , Dados de Sequência Molecular , Especificidade de Órgãos , Reação em Cadeia da Polimerase , Mapeamento por Restrição , Células-Tronco/metabolismo , Testículo/metabolismoRESUMO
Glial fibrillary acidic protein (GFAP) is the main component of the intermediate filaments in cells of astroglial lineage, including astrocytes in the CNS, nonmyelin forming Schwann cells and enteric glia. To address the function of GFAP in vivo, we have disrupted the GFAP gene in mice via targeted mutation in embryonic stem cells. Mice lacking GFAP developed normally, reached adulthood and reproduced. We did not find any abnormalities in the histological architecture of the CNS, in their behavior, motility, memory, blood-brain barrier function, myenteric plexi histology or intestinal peristaltic movement. Comparisons between GFAP and S-100 immunohistochemical staining patterns in the hippocampus of wild-type and mutant mice suggested a normal abundance of astrocytes in GFAP-negative mice, however, in contrast to wild-types, GFAP-negative astrocytes of the hippocampus and in the white matter of the spinal cord were completely lacking intermediate filaments. This shows that the loss of GFAP intermediate filaments is not compensated for by the up-regulation of other intermediate filament proteins, such as vimentin. The GFAP-negative mice displayed post-traumatic reactive gliosis, which suggests that GFAP up-regulation, a hallmark of reactive gliosis, is not an obligatory requirement for this process.
Assuntos
Astrócitos/fisiologia , Proteína Glial Fibrilar Ácida/deficiência , Filamentos Intermediários/fisiologia , Camundongos Mutantes , Tecido Nervoso/fisiologia , Animais , Sequência de Bases , Barreira Hematoencefálica/fisiologia , Encéfalo/ultraestrutura , Química Encefálica , Feminino , Proteína Glial Fibrilar Ácida/genética , Gliose , Hipocampo/química , Hipocampo/ultraestrutura , Histocitoquímica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes/embriologia , Camundongos Mutantes/crescimento & desenvolvimento , Camundongos Mutantes/psicologia , Dados de Sequência Molecular , Plexo Mientérico/química , Plexo Mientérico/ultraestrutura , Tecido Nervoso/anatomia & histologia , Tecido Nervoso/química , Desempenho Psicomotor , RNA Mensageiro/análise , Proteínas S100/isolamento & purificação , Medula Espinal/química , Medula Espinal/ultraestruturaRESUMO
A phenotypic analysis was performed on two alleles at the pygmy locus which arose by insertional mutagenesis in transgenic mice. Similar to the spontaneous mutant pygmy, the adult insertional transgenic mutants are 40% of the size of wild-type litter-mates whereas adult heterozygotes are 80% of wild-type litter-mates. An analysis of the various organs revealed that, in general, there was a reduction in weight of each organ commensurate with the overall reduction in body size. However, two organs did not follow this pattern, the brain being disproportionately larger and the adrenals disproportionately smaller in the mutant mice. In addition, mini-mice have less adipose tissue than their wild-type or heterozygous litter-mates. A developmental analysis determined that mutants could first be identified on the basis of reduced body weight at day 15.5 of gestation. The small size is not due to a growth hormone deficiency so these mice differ from other known dwarf mouse mutants. Therefore they should provide insight into the growth hormone-resistant human dwarfisms and help in furthering our knowledge of mammalian growth and development.