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1.
Neuroscience ; 148(2): 510-21, 2007 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-17651901

RESUMO

Apoptosis-associated tyrosine kinase (AATYK) is a protein kinase that is predominantly expressed in the nervous system and is involved in apoptosis and neurite growth of cerebellar granule cells. In this study, we cloned three new members of the mouse AATYK family, AATYK1B, AATYK2 and AATYK3. AATYK1B is a splicing variant of the previously reported AATYK1 (referred to as AATYK1A hereafter). In comparison with AATYK1A, these three AATYK members were characterized by having an extra N-terminal region that consists of a signal peptide-like sequence and a predicted transmembrane (TM) region, which is followed by a kinase domain and a long C-terminal domain. Both TM-containing AATYK isoforms (AATYK(+)TM: AATYK1B, 2, and 3) and TM-lacking isoform (AATYK(-)TM: AATYK1A) were recovered in membrane fractions, suggesting that AATYK(+)TM and AATYK(-)TM are transmembrane- and peripheral-membrane protein kinases, respectively. AATYK1A was recovered in the soluble fraction when the cells were treated with 2-bromo palmitate, suggesting that AATYK1A associates with membrane via palmitoylation. The kinase domain was highly conserved among all AATYK members and was shown to be catalytically active. Three AATYK family members were predominantly expressed in adult mouse brains with almost similar expression profiles: widespread distribution over the various brain regions, especially in the cerebellum and hippocampus, and up-regulated expression during development of the cerebellum. In cultured cerebellar granule cells, AATYK1 was abundantly localized in both soma and axons, AATYK2 distribution was restricted to soma, and AATYK3 was punctately present over the cells. AATYK1 was concentrated in the central domain of growth cones of dorsal root ganglion neurons. Our results indicate that AATYK family members are brain-dominant and membrane-associated kinases with slightly different distribution patterns in the developing and adult mouse brain, which may be involved in fine regulation of neuronal functions including neurite extension and apoptosis.


Assuntos
Proteínas Reguladoras de Apoptose/classificação , Proteínas Reguladoras de Apoptose/fisiologia , Apoptose/fisiologia , Neurônios/metabolismo , Animais , Animais Recém-Nascidos , Encéfalo/citologia , Encéfalo/metabolismo , Células Cultivadas , Embrião de Galinha , Clonagem Molecular/métodos , Regulação Enzimológica da Expressão Gênica/genética , Camundongos , Fosfotransferases/metabolismo , Alinhamento de Sequência/métodos , Transfecção/métodos
2.
Eur J Neurosci ; 14(1): 57-63, 2001 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-11488949

RESUMO

The cerebellar Purkinje cell has been the focus of numerous studies involving the analysis of development and information processing in the nervous system. Purkinje cells represent less than 0.1% of the total cell content of the cerebellum. To facilitate studies of molecules that are expressed in such a small proportion of neurons, we have established procedures for the purification of these cells. Transgenic mice were developed in which the expression of green fluorescent protein (GFP) was controlled by the L7 promoter. In adult cerebellum, GFP fluorescence was only detected in Purkinje cells, where it filled dendrites, soma and axons. GFP fluorescence was detected in Purkinje cells as early as embryonic day 17 and increased during development in vivo and in dissociated cerebellar culture. Mirroring endogenous L7 expression, high levels of GFP were observed in retinal rod bipolar cells. Lower levels of GFP were seen in olfactory periglomerular cells, neurons in the interpeduncular nucleus, and superior colliculus neurons. Cerebella from transgenic mice were dissociated by mild enzymatic treatment and Purkinje cells were isolated by fluorescence-activated cell sorting (FACS). By selecting optimal parameters, a fraction of viable Purkinje cells that was 94% pure was obtained. These results indicate that FACS is a powerful tool for isolating Purkinje cells from postnatal L7-GFP transgenic mice. GFP-positive neurons will also be useful in the real-time observation of dendritic morphogenesis and axonal outgrowth during development, or after neuronal activity in vitro.


Assuntos
Técnicas de Cultura de Células/métodos , Células Cultivadas/citologia , Citometria de Fluxo/métodos , Indicadores e Reagentes/metabolismo , Proteínas Luminescentes/genética , Camundongos Transgênicos/metabolismo , Células de Purkinje/citologia , Animais , Células Cultivadas/metabolismo , Proteínas de Fluorescência Verde , Camundongos , Camundongos Transgênicos/anatomia & histologia , Camundongos Transgênicos/genética , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Bulbo Olfatório/citologia , Bulbo Olfatório/metabolismo , Regiões Promotoras Genéticas/fisiologia , Células de Purkinje/metabolismo , Retina/citologia , Retina/metabolismo , Transcrição Gênica/fisiologia
3.
Oncogene ; 20(9): 1022-32, 2001 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-11314040

RESUMO

We isolated three related cDNA clones from a mouse cerebellar library; the type I cDNA was identical to the gene encoding the apoptosis-associated tyrosine kinase (AATYK), whose expression in myeloid precursor cells is increased during growth arrest or apoptosis. Low levels of AATYK mRNA expression were seen in adult mouse brains but not in embryos. In situ hybridization confirmed the widespread expression of AATYK mRNA in neurons throughout the adult brain. AATYK possessed tyrosine kinase activity and was autophosphorylated when expressed in 293 cells. AATYK mRNA expression was rapidly induced in cultured cerebellar granule cells during apoptosis induced by a low concentration of KCl (5 mM). Levels of endogenous AATYK protein were increased only slightly, but they were accompanied by an increase in molecular weight during apoptosis. Results of the tyrosine phosphatase treatments indicated that the increase in molecular weight was partly caused by tyrosine phosphorylation. The number of apoptotic granule cells overexpressing wild-type AATYK protein was significantly greater than the number of apoptotic granule cells overexpressing a mutant AATYK that lacked tyrosine kinase activity in low concentrations of KCl. These findings suggest that through its tyrosine kinase activity, AATYK is involved in the apoptosis of mature neurons.


Assuntos
Apoptose , Encéfalo/enzimologia , Proteínas Tirosina Quinases/biossíntese , RNA Mensageiro/biossíntese , Sequência de Aminoácidos , Animais , Proteínas Reguladoras de Apoptose , Sequência de Bases , Northern Blotting , Diferenciação Celular , Células Cultivadas , Primers do DNA/química , Vetores Genéticos , Técnicas Imunoenzimáticas , Camundongos , Dados de Sequência Molecular , Neurônios/enzimologia , Proteínas Tirosina Quinases/genética , Células de Purkinje/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Distribuição Tecidual , Transfecção
4.
Mol Genet Metab ; 68(3): 346-56, 1999 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-10562461

RESUMO

The expression of carbamoyl phosphate synthetase I (CPS) gene is suppressed in the liver of carnitine-deficient juvenile visceral steatosis (JVS) mice at weaning and under starvation at adult age. To clarify the suppression mechanism, we produced CPSL transgenic JVS mice carrying a transgene composed of the chloramphenicol acetyltransferase (CAT) gene with the upstream region (-12 kb to +138) of the rat CPS gene and CPSE transgenic JVS mice carrying a transgene composed of the luciferase gene with minimal promoter (299 bp from -161 to +138) and enhancer (469 bp around -6.3 kb) fragments of the rat gene. The expression of the CAT gene as well as the endogenous CPS was suppressed in CPSL transgenic JVS mice, but luciferase gene expression was not suppressed in CPSE transgenic JVS mice. We isolated the 5'-upstream region of the mouse CPS gene and identified an activator protein-1 (AP-1) site downstream of the minimum enhancer region of both rat and mouse CPS genes. In conjunction with the 313-bp mouse promoter region, the 714-bp mouse enhancer fragment conferred a cell-type-dependent hormone responsiveness. In rat primary cultured hepatocytes, the addition of oleic acid suppressed reporter gene expression induced by dexamethasone in the construct containing the enhancer fragment of 714 bp with the AP-1 site, but not in its AP-1 site mutants or in 519 bp without the AP-1 site. These results strongly suggest that direct protein-protein interaction between AP-1 and glucocorticoid receptor is not involved in the suppression of the CPS gene in JVS mice and that the AP-1 element is the cis-element which is responsible for the suppression.


Assuntos
Carbamoil-Fosfato Sintase (Amônia)/genética , Carnitina/deficiência , Regulação Enzimológica da Expressão Gênica , Fígado/enzimologia , Animais , Sítios de Ligação , Carbamoil-Fosfato Sintase (Amônia)/biossíntese , Cloranfenicol O-Acetiltransferase/genética , Clonagem Molecular , Modelos Animais de Doenças , Elementos Facilitadores Genéticos , Repressão Enzimática , Genes Reporter , Neoplasias Hepáticas Experimentais , Luciferases/genética , Camundongos , Camundongos Transgênicos , Regiões Promotoras Genéticas , Ratos , Mapeamento por Restrição , Fator de Transcrição AP-1/metabolismo , Células Tumorais Cultivadas
5.
J Mol Cell Cardiol ; 29(2): 571-8, 1997 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-9140816

RESUMO

To characterize cardiac hypertrophy in juvenile visceral steatosis (JVS) mice with systemic carnitine deficiency, we investigated how the hypertrophy develops and whether it is associated with altered expression of any specific genes, especially atrial natriuretic peptide (ANP) and contractile protein genes, in the hypertrophied ventricle. Cardiac hypertrophy in JVS mice became apparent at 10 days after birth and progressed during development. The hypertrophy was observed in the ventricles but not in the atria. ANP mRNA was more intensively expressed in JVS ventricles than in control even at 5 days. Carnitine administration ameliorated the cardiac hypertrophy and suppressed the augmentation of ANP mRNA in the ventricles. Isoform change of expression of alpha-actin genes from cardiac to skeletal was seen in the ventricles of JVS mice at 2 weeks. There was no difference in the ratio of beta-myosin heavy chain mRNA to alpha-myosin heavy chain mRNA between control and JVS mice at 5 days, but at 2 weeks the ratio was significantly lower in JVS mice than in control. These results suggest that the molecular characteristics of cardiac hypertrophy caused by carnitine deficiency are different from those of cardiac hypertrophy caused by aortic constriction.


Assuntos
Fator Natriurético Atrial/metabolismo , Cardiomegalia/metabolismo , Carnitina/deficiência , Ventrículos do Coração/patologia , Contração Miocárdica/genética , Actinas/genética , Actinas/metabolismo , Animais , Fator Natriurético Atrial/genética , Peso Corporal , Modelos Animais de Doenças , Regulação da Expressão Gênica no Desenvolvimento , Ventrículos do Coração/metabolismo , Camundongos , Camundongos Endogâmicos C3H , Camundongos Mutantes , Cadeias Pesadas de Miosina/genética , Cadeias Pesadas de Miosina/metabolismo , Tamanho do Órgão , RNA Mensageiro/metabolismo , Fatores de Tempo
7.
J Biochem ; 121(1): 172-7, 1997 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-9058209

RESUMO

Systemic carnitine-deficient juvenile visceral steatosis (JVS) mice exhibit decreased expression of some liver-selective genes including those for the urea cycle enzymes during the infantile period. At 25 days, carbamoylphosphate synthetase (CPS) mRNA level was remarkably low in the liver of JVS mice, and the HNF-4 and C/EBP-alpha mRNA contents were also reduced. HNF-3 alpha and C/EBP-beta mRNAs were slightly higher in the liver of JVS mice, and HNF-1 mRNA remained normal. These results, together with the developmental changes of these transcription factor mRNA levels, suggest that HNF-4 and C/EBP-alpha are involved in the suppression of CPS expression. If JVS mice survived the crisis at 4-5 weeks, their body weight caught up with that of control mice around 7 weeks. The steady-state levels of CPS and argininosuccinate synthetase (ASS) mRNAs in the liver of JVS mice were normalized by no later than 8 weeks. Starvation for 48 h caused an increase of about twofold in CPS and ASS mRNA levels in the liver of control mice, while the same treatment failed to increase their levels in the liver of JVS mice. The starvation similarly caused increases in HNF-4 and C/EBP-beta mRNA levels in the liver of both control and JVS mice, but the increases were significantly less in JVS mice than in control mice. Thus, the lack of induction of CPS and ASS mRNAs during development and under starvation in JVS mice correlated with the lower induction of HNF-4 and C/EBP-alpha mRNAs, and of HNF-4 and C/ EBP-beta mRNAs, respectively. Furthermore, all these changes seemed to correlate with the presence of fatty liver and the high serum free fatty acid levels, suggesting that disturbance of fatty acid metabolism affects nitrogen metabolism at least in part via altered gene expression of transcription factors such as HNF-4, C/EBP-alpha, and C/EBP-beta.


Assuntos
Carnitina/deficiência , Hipoglicemia/metabolismo , Fígado/enzimologia , Inanição/genética , Ureia/metabolismo , Fatores Etários , Animais , Argininossuccinato Sintase/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos , Peso Corporal , Proteínas Estimuladoras de Ligação a CCAAT , Carbamoil-Fosfato Sintase (Amônia)/genética , Carbamoil-Fosfato Sintase (Amônia)/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Ácidos Graxos/sangue , Fígado Gorduroso/enzimologia , Regulação Enzimológica da Expressão Gênica , Gliceraldeído-3-Fosfato Desidrogenases/genética , Fator 1 Nuclear de Hepatócito , Fator 1-alfa Nuclear de Hepatócito , Fator 1-beta Nuclear de Hepatócito , Fator 3-alfa Nuclear de Hepatócito , Fator 4 Nuclear de Hepatócito , Hormônios/sangue , Metabolismo dos Lipídeos , Lipídeos/química , Fígado/química , Fígado/crescimento & desenvolvimento , Camundongos , Camundongos Mutantes , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Tamanho do Órgão , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , RNA Mensageiro/metabolismo , Inanição/enzimologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
8.
FEBS Lett ; 399(3): 310-2, 1996 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-8985169

RESUMO

In order to test the possibility that free fatty acids are the mediator of the abnormal expression of urea cycle enzyme genes in carnitine-deficient juvenile visceral steatosis (JVS) mice, the effects of fatty acids on urea cycle enzyme, carbamoylphosphate synthetase (CPS) and argininosuccinate synthetase (ASS), mRNA levels were examined in rat primary cultured hepatocytes. Addition of a synthetic glucocorticoid hormone, dexamethasone, caused increases in CPS and ASS mRNAs. Further addition of oleic acid suppressed the induction of CPS and ASS mRNAs by dexamethasone. In contrast, the phosphoenolpyruvate carboxykinase (PEPCK) mRNA level induced by dexamethasone was enhanced in the presence of oleic acid. The effects were reversed on further addition of carnitine. The mRNA levels of these enzymes induced by dibutyryl cAMP were not affected by the addition of oleic acid. A study of the specificity of fatty acids revealed that long-chain fatty acids of more than 16 carbons chain length had a suppressive effect on the CPS mRNA level induced by dexamethasone and that the presence of double bonds enhanced the effect. The changes in gene expression of CPS, ASS and PEPCK caused by the fatty acids in the cultured hepatocytes were very similar to those observed in the liver of JVS mice. The AP-1 DNA binding activity in the presence of dexamethasone was slightly enhanced by the addition of oleic acid. These results suggest that the long-chain fatty acids not metabolized in JVS mice are mediators of the abnormal gene expression in the liver which results in hyperammonemia.


Assuntos
Argininossuccinato Sintase/genética , Carbamoil-Fosfato Sintase (Amônia)/genética , Dexametasona/farmacologia , Ácidos Graxos/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Fígado/efeitos dos fármacos , Animais , Células Cultivadas , Fígado/citologia , Fígado/enzimologia , Masculino , RNA Mensageiro/genética , Ratos , Ratos Wistar
10.
FEBS Lett ; 386(1): 26-8, 1996 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-8635596

RESUMO

Earlier we reported the primary structure of serum calcium-decreasing factor (caldecrin) from rat pancreas, a protein which is considered to be a member of the elastase family. In this report, we describe the isolation of the two homologous cDNA clones encoding caldecrin from human pancreas, the structures of which are identical except for one base and the corresponding amino acid residue. These human caldecrin isoforms are composed of a signal peptide of 16 amino acids, a propeptide of 13 amino acids, and a mature form of 239 amino acids. Both recombinant caldecrins showed the same chymotrypsin-type protease activity and hypocalcemic activity. The hypocalcemic activity of both remained intact even after treatment with PMSF to abolish their protease activity. These results suggest that human caldecrin possesses hypocalcemic activity that has no connection with its protease activity.


Assuntos
Serina Endopeptidases/genética , Sequência de Aminoácidos , Animais , Baculoviridae/genética , Sequência de Bases , Cálcio/sangue , Cálcio/metabolismo , Clonagem Molecular , Humanos , Dados de Sequência Molecular , Pâncreas/química , Pâncreas/enzimologia , Ratos , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Homologia de Sequência de Aminoácidos , Serina Endopeptidases/biossíntese , Serina Endopeptidases/química
11.
J Biol Chem ; 270(51): 30315-21, 1995 Dec 22.
Artigo em Inglês | MEDLINE | ID: mdl-8530454

RESUMO

We previously reported on the purification of a serum calcium-decreasing factor, referred to as caldecrin, from porcine pancreas, that is thought to be a serine protease (Tomomura, A., Fukushige, T., Noda, T., Noikura, T., and Saheki, T. (1992) FEBS Lett. 301, 277-281). In the present study, we purified caldecrin from rat pancreas and determined its primary structure by cDNA cloning. The predicted caldecrin protein is presumed to be synthesized as a preproenzyme of 268 amino acids with a signal peptide of 16 amino acids and an activation peptide of 13 amino acids, and is, with the exception of a central region, almost identical to the reported rat pancreatic elastase IV sequence. The caldecrin gene is selectively expressed in the pancreas, as judged by Northern blot analysis. After expression in BMT-10 cells, immunoreactive caldecrin was found in the culture supernatant, and it inhibited the parathyroid hormone-stimulated 45Ca release from cultured fetal long bones. Catalytic site mutants were synthesized in a baculovirus system, and recombinant mutants also decreased the serum calcium level of mice. These data implicate caldecrin, a protease closely related to elastase IV, in the regulation of blood calcium levels.


Assuntos
Reabsorção Óssea , Antagonistas de Hormônios/farmacologia , Pâncreas/metabolismo , Serina Endopeptidases/biossíntese , Serina Endopeptidases/farmacologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Bioensaio , Osso e Ossos/efeitos dos fármacos , Osso e Ossos/metabolismo , Cálcio/metabolismo , Clonagem Molecular , Primers do DNA , DNA Complementar , Feto , Expressão Gênica , Antagonistas de Hormônios/isolamento & purificação , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Técnicas de Cultura de Órgãos , Especificidade de Órgãos , Pâncreas/química , Elastase Pancreática/biossíntese , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/isolamento & purificação , Mutação Puntual , Reação em Cadeia da Polimerase , Sinais Direcionadores de Proteínas/biossíntese , Ratos , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/farmacologia , Homologia de Sequência de Aminoácidos , Serina Endopeptidases/isolamento & purificação , Suínos
12.
Biochim Biophys Acta ; 1226(3): 307-14, 1994 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-7914432

RESUMO

Carnitine-deficient jvs mice expressed reduced levels of a group of genes which are preferentially expressed in the liver, including urea cycle enzyme genes (Biochim. Biophys. Acta 1138, 167-171, 1992). The expression of alpha-fetoprotein and aldolase A was elevated, indicating that the liver of jvs mice is undifferentiated or dedifferentiated (FEBS Lett. 311, 63-66, 1992). Studies of the hormone signal transduction pathway showed that serum cortisol and plasma glucagon levels of jvs mice were 2 and 3 times higher, respectively, than those of normal mice, and that the hormone binding activity of glucocorticoid receptor (GR) in the cytosol of jvs liver was 50% of normal mice, which reflected the amount of receptor protein in the cytosol. On the other hand, GR protein accumulated in the nuclear fraction in jvs mice. Exogenously administrated dexamethasone induced carbamoyl phosphate synthetase (CPS) and tyrosine aminotransferase (TAT) mRNAs in jvs mice, indicating that CPS and TAT genes in jvs mice are responsive to induction by glucocorticoid and cAMP. Analysis of transacting factors by gel retardation assay revealed that HNF-1, COUP-TF and SP-1 were detected at almost the same level in the hepatic nuclear fraction of jvs mice as in normal littermates, and C/EBP and CREB were a little higher in jvs mice, suggesting that these factors are probably not targets of jvs mutation causing abnormal gene expression in the liver. On the other hand, AP-1 binding activity was much higher in jvs mice from an early age, preceding the abnormal expression of urea cycle enzyme, and carnitine administration normalized AP-1 binding activity. We suggest that elevated AP-1 binding induced by carnitine deficiency is closely connected with the abnormal gene expression in the liver.


Assuntos
Carnitina/deficiência , Proteínas de Ligação a DNA/metabolismo , Proteínas de Homeodomínio , Fígado/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2 , Proteínas Repressoras , Proteínas de Saccharomyces cerevisiae , Animais , Sequência de Bases , Carbamoil-Fosfato Sintase (Amônia)/genética , Carnitina/farmacologia , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica , Glucagon/sangue , Hidrocortisona/sangue , Masculino , Camundongos , Camundongos Mutantes , Antígenos de Histocompatibilidade Menor , Dados de Sequência Molecular , Receptores de Glucocorticoides/análise , Proteína de Replicação C , Tirosina Transaminase/genética
13.
FEBS Lett ; 335(2): 213-6, 1993 Dec 06.
Artigo em Inglês | MEDLINE | ID: mdl-8253199

RESUMO

Proform serum calcium-decreasing factor (procaldecrin) was purified from porcine pancreas acetone powder. Procaldecrin showed chymotrypsin activity after trypsin treatment in a time- and dose-dependent manner. Procaldecrin did not possess serum calcium-decreasing activity but acquired serum calcium-decreasing activity as well as protease activity after trypsin treatment. However, PMSF treatment after activation of procaldecrin by trypsin did not affect the serum calcium-decreasing activity, even though protease activity was nullified by treatment with PMSF. These findings suggest that the serum calcium-decreasing activity acquired by procaldecrin requires conformational change caused by trypsin treatment.


Assuntos
Cálcio/sangue , Precursores Enzimáticos/metabolismo , Serina Endopeptidases/metabolismo , Tripsina/metabolismo , Sequência de Aminoácidos , Animais , Ativação Enzimática , Dados de Sequência Molecular , Suínos
14.
FEBS Lett ; 326(1-3): 267-71, 1993 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-8325377

RESUMO

We have reported the clinical and biochemical findings in juvenile visceral steatosis (jvs) mice with systemic carnitine deficiency. This paper is the first report about cardiomyopathy in jvs mice. Adult jvs mice (at the age of 2-3 months) show cardiac hypertrophy which is caused by enlargement of the cardiac muscle cell associated with increases of non-collagen protein and DNA content. Carnitine administration (2 mg/head, twice a day, from 1 month of age) significantly suppresses the cardiac hypertrophy, showing that carnitine deficiency plays an important role in the development of the cardiac hypertrophy. The discovery of cardiac hypertrophy in carnitine-deficient jvs mice will lead to clarification of the pathophysiology of cardiomyopathy in systemic carnitine deficiency in human beings.


Assuntos
Cardiomegalia/etiologia , Carnitina/deficiência , Fígado Gorduroso/complicações , Animais , Cardiomegalia/patologia , Cardiomegalia/prevenção & controle , Carnitina/administração & dosagem , Carnitina/uso terapêutico , Núcleo Celular/patologia , Citoplasma/patologia , DNA/metabolismo , Fígado Gorduroso/patologia , Feminino , Homozigoto , Masculino , Camundongos , Camundongos Endogâmicos C3H , Camundongos Mutantes , Miocárdio/patologia , Tamanho do Órgão
15.
FEBS Lett ; 311(1): 63-6, 1992 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-1383034

RESUMO

We determined the mRNA levels of c-jun and c-fos in the liver of C3H-H-2 degrees jvs mice. Both were higher in jvs mice than in normal mice. The level of c-jun mRNA increased gradually after birth, but in the control mice there was almost no change. In addition, alpha-fetoprotein and aldolase A mRNA levels were also higher than in normal littermates. These results suggest that the pattern of the gene expression in jvs mice partly resembles the one that occurs in undifferentiated hepatocytes and/or hepatocellular carcinoma.


Assuntos
Carnitina/deficiência , Erros Inatos do Metabolismo Lipídico/genética , Fígado/metabolismo , Proto-Oncogenes , RNA Mensageiro/biossíntese , Envelhecimento , Animais , Animais Recém-Nascidos/metabolismo , Frutose-Bifosfato Aldolase/genética , Genes fos , Genes jun , Homozigoto , Camundongos , Camundongos Mutantes/metabolismo , Transcrição Gênica , alfa-Fetoproteínas/genética
16.
J Biol Chem ; 267(8): 5032-5, 1992 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-1544887

RESUMO

Previous studies in our laboratories have revealed that juvenile visceral steatosis mice show suppressed transcription of urea cycle enzyme genes during development and are systemically deficient in carnitine. It has not yet been explained, however, how this carnitine deficiency relates to the abnormal gene expression. We investigated the effect of carnitine on abnormal gene expression, growth retardation, and fatty liver. Carnitine administration relieved the suppression of the developmental induction of two urea cycle enzymes examined, carbamoyl-phosphate synthetase and argininosuccinate synthase, and kept the activities of enzymes normal. However, carnitine did not reduce accumulated lipid in the liver to the normal level. These results suggest that carnitine deficiency plays an important role in the abnormal expression of urea cycle enzyme genes and that the abnormal expression of the genes is not directly caused by lipid accumulation in the liver.


Assuntos
Argininossuccinato Sintase/genética , Carbamoil-Fosfato Sintase (Amônia)/genética , Carnitina/farmacologia , Fígado Gorduroso/enzimologia , Frutose-Bifosfato Aldolase/genética , Fígado/metabolismo , Ornitina Carbamoiltransferase/genética , RNA Mensageiro/metabolismo , Transcrição Gênica/efeitos dos fármacos , Ureia/metabolismo , Envelhecimento , Animais , Argininossuccinato Sintase/biossíntese , Northern Blotting , Peso Corporal/efeitos dos fármacos , Carbamoil-Fosfato Sintase (Amônia)/biossíntese , Repressão Enzimática/efeitos dos fármacos , Fígado Gorduroso/genética , Frutose-Bifosfato Aldolase/biossíntese , Fígado/efeitos dos fármacos , Fígado/enzimologia , Camundongos , Camundongos Endogâmicos C3H , Camundongos Mutantes , Ornitina Carbamoiltransferase/biossíntese , RNA Mensageiro/genética
17.
Biochim Biophys Acta ; 1138(2): 167-71, 1992 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-1540663

RESUMO

Juvenile visceral steatosis (jvs) mice from the C3H-H-2 degrees strain have markedly low levels of all the hepatic urea cycle enzymes (Imamura et al. (1990) FEBS Lett. 260, 119-121). The steady state levels of messenger RNA for the four urea cycle enzymes examined and also for albumin and serine dehydratase were severely reduced in the liver. The levels of mRNA for other liver-specific enzymes including aldolase B and phospho enol pyruvate carboxykinase did not vary significantly from normal littermates. As for extrahepatic expression of the urea cycle enzymes, only argininosuccinate synthetase in the kidney was decreased. Nuclear run-on experiments showed reduced transcription of the corresponding genes, which mostly accounts for the low mRNA levels. Furthermore, the time-course of mRNA accumulation from 5 days of age showed that the developmental induction of hepatic carbamyl phosphate synthetase and argininosuccinate synthetase mRNAs was strongly suppressed. These results suggest that jvs affects not only the regulation of the tissue-specific expression of the urea cycle enzymes but also the regulation of their developmental induction.


Assuntos
Argininossuccinato Liase/genética , Argininossuccinato Sintase/genética , Carbamoil-Fosfato Sintase (Amônia)/genética , Ornitina Carbamoiltransferase/genética , Animais , Argininossuccinato Liase/metabolismo , Argininossuccinato Sintase/metabolismo , Northern Blotting , Carbamoil-Fosfato Sintase (Amônia)/metabolismo , Frutose-Bifosfato Aldolase/genética , Frutose-Bifosfato Aldolase/metabolismo , Expressão Gênica , Rim/enzimologia , Camundongos , Ornitina Carbamoiltransferase/metabolismo , Fosfoenolpiruvato Carboxiquinase (GTP)/genética , Fosfoenolpiruvato Carboxiquinase (GTP)/metabolismo , RNA Mensageiro/genética , Transcrição Gênica
18.
Biochem Biophys Res Commun ; 171(2): 603-9, 1990 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-2403350

RESUMO

MK is a gene whose expression increases transiently during retinoic acid-induced differentiation of embryonal carcinoma cells. MK polypeptide was secreted by differentiating HM-1 embryonal carcinoma cells and by L-cells transfected with an MK cDNA under the control of the beta-actin promoter and Rous sarcoma virus enhancer. MK polypeptide was found to have heparin binding activity. Conditioned medium of the transfected L-cells promoted growth of PC-12 pheochromocytoma cells. These findings support the view that MK polypeptide is a secreted factor involved in regulation of growth and differentiation.


Assuntos
Proteínas de Transporte/genética , Diferenciação Celular/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Heparina/metabolismo , Tretinoína/farmacologia , Animais , Proteínas de Transporte/isolamento & purificação , Proteínas de Transporte/metabolismo , Linhagem Celular , Cromatografia de Afinidade , Cinética , Células L/citologia , Células L/efeitos dos fármacos , Células L/metabolismo , Regiões Promotoras Genéticas , Biossíntese de Proteínas , Coelhos , Reticulócitos/metabolismo , Teratoma , Transfecção
19.
J Biol Chem ; 265(16): 9441-3, 1990 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-2345177

RESUMO

MK is a gene that is expressed temporarily during the early stages of retinoic acid-induced differentiation of embryonal carcinoma (EC) cells and during the mid-gestation period of mouse embryogenesis. The 5'-regions of MK cDNAs and their mRNAs are heterogeneous; so far three kinds of MK cDNAs (MK1, MK2, and MK3) have been isolated. The MK gene was cloned from a genomic DNA library of a BALB/c mouse, and its structure was elucidated. 5'-Region sequences specific for MK1, MK2, and MK3 were arranged in the order of MK3, MK2, and MK1. Then, there was a sequence common to all MK cDNAs consisting of four exons. The results indicate that different species of MK mRNA are generated by the use of alternative promoters and different modes of splicing.


Assuntos
Diferenciação Celular/genética , Desenvolvimento Embrionário e Fetal/genética , Expressão Gênica/efeitos dos fármacos , Teratoma/patologia , Tretinoína/farmacologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , DNA/genética , Éxons , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , Células Tumorais Cultivadas
20.
J Biol Chem ; 265(18): 10765-70, 1990 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-2355021

RESUMO

A newly identified gene, MK, is transiently expressed in the early stages of embryonal carcinoma cell differentiation and in the mid-gestation period of mouse embryogenesis (Kadomatsu, K., Tomomura, M., and Muramatsu, T. (1988) Biochem. Biophys. Res. Commun. 151, 1312-1318). Analysis of various MK cDNA clones revealed differences in the 5'-region. So far three classes of cDNA clones (MK1, MK2, and MK3) have been identified; they were different in the 5'-untranslated region but shared the rest of the sequence. Ribonuclease protection, RNA blotting, and primer extension revealed that MK2-type RNA was the major MK RNA in retinoic acid-treated embryonal carcinoma cells. In addition, the number of A residues in an oligo(A) stretch in the 5'-side of the common sequence differed from 9 to 29. The number was 9 in the most frequent cases, when the putative MK polypeptide had a molecular weight of about 15,500 and had a signal peptide-like sequence. Hybrid selected MK RNA yielded the predicted polypeptide upon in vitro translation. When pancreatic microsomal membranes were included in the translation system, the translation product of MK RNA was processed and entered into the lumen of the membranes. These results suggest that the product of the MK gene is an extracellular polypeptide.


Assuntos
Expressão Gênica/efeitos dos fármacos , Genes/efeitos dos fármacos , Biossíntese de Proteínas , Teratoma/genética , Transcrição Gênica , Tretinoína/farmacologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Diferenciação Celular , Linhagem Celular , Clonagem Molecular , Sondas de DNA , DNA de Neoplasias/genética , DNA de Neoplasias/isolamento & purificação , Biblioteca Gênica , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Sondas de Oligonucleotídeos , RNA Neoplásico/genética , RNA Neoplásico/isolamento & purificação , Mapeamento por Restrição
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