RESUMO
Prior studies have established an inverse association between cigarette smoking and the risk of developing Parkinson's disease (PD), and currently, the disease-modifying potential of the nicotine patch is being tested in clinical trials. To identify genes that interact with the effect of smoking/nicotine, we conducted genome-wide interaction studies in humans and in Drosophila. We identified SV2C, which encodes a synaptic-vesicle protein in PD-vulnerable substantia nigra (P=1 × 10(-7) for gene-smoking interaction on PD risk), and CG14691, which is predicted to encode a synaptic-vesicle protein in Drosophila (P=2 × 10(-11) for nicotine-paraquat interaction on gene expression). SV2C is biologically plausible because nicotine enhances the release of dopamine through synaptic vesicles, and PD is caused by the depletion of dopamine. Effect of smoking on PD varied by SV2C genotype from protective to neutral to harmful (P=5 × 10(-10)). Taken together, cross-validating evidence from humans and Drosophila suggests SV2C is involved in PD pathogenesis and it might be a useful marker for pharmacogenomics studies involving nicotine.
Assuntos
Nicotina/efeitos adversos , Doença de Parkinson/etiologia , Fumar/efeitos adversos , Animais , Dopamina/metabolismo , Drosophila , Expressão Gênica , Interação Gene-Ambiente , Estudo de Associação Genômica Ampla , Humanos , Modelos Biológicos , Doença de Parkinson/genética , Doença de Parkinson/metabolismoRESUMO
Bicoid directs anterior development in Drosophila embryos by activating different genes along the anterior-posterior axis. However, its activity is down-regulated at the anterior tip of the embryo, in a process known as retraction. Retraction is under the control of the terminal polarity system, and results in localized repression of Bicoid target genes. Here, we describe a Drosophila homolog of human SAP18, a member of the Sin3A/Rpd3 histone deacetylase complex. dSAP18 interacts with Bicoid in yeast and in vitro, and is expressed early in development coincident with Bicoid. In tissue culture cells, dSAP18 inhibits the ability of Bicoid to activate reporter genes. These results suggest a model in which dSAP18 interacts with Bicoid to silence expression of Bicoid target genes in the anterior tip of the embryo.
Assuntos
Proteínas de Transporte , Drosophila/embriologia , Histona Desacetilases/metabolismo , Proteínas de Homeodomínio/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Sequência de Aminoácidos , Animais , Proteínas Relacionadas à Autofagia , Sequência de Bases , Mapeamento Cromossômico , Proteínas Correpressoras , DNA , Proteínas de Drosophila , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Homeodomínio/antagonistas & inibidores , Humanos , Dados de Sequência Molecular , Ligação Proteica , Proteínas de Ligação a RNA , Proteínas de Saccharomyces cerevisiae , Homologia de Sequência de Aminoácidos , Transativadores/antagonistas & inibidores , Fatores de Transcrição/química , Fatores de Transcrição/genética , Transcrição GênicaRESUMO
Cooperative DNA binding is thought to contribute to the ability of the Drosophila melanogaster protein, Bicoid, to stimulate transcription of target genes in precise sub-domains within the embryo. As a first step toward testing this idea, we devised a genetic screen to isolate mutations in Bicoid that specifically disrupt cooperative interactions, but do not disrupt DNA recognition or transcription activation. The screen was carried out in Saccharomyces cerevisiae and 12 cooperativity mutants were identified. The mutations map across most of the Bicoid protein, with some located within the DNA-binding domain (homeodomain). Four homeodomain mutants were characterized in yeast and shown to activate a single-site reporter gene to levels comparable to that of wild-type, indicating that DNA binding per se is not affected. However, these mutants failed to show cooperative coupling between high and low-affinity sites, and showed reduced activation of a reporter gene carrying a natural Drosophila enhancer. Homology modeling indicated that none of the four mutations is in residues that contact DNA. Instead, these residues are likely to interact with other DNA-bound Bicoid monomers or other parts of the Bicoid protein. In vitro, the isolated homeodomains did not show strong cooperativity defects, supporting the idea that other regions of Bicoid are also important for cooperativity. This study describes the first systematic screen to identify cooperativity mutations in a eukaryotic DNA-binding protein.
Assuntos
Sítio Alostérico , DNA/metabolismo , Proteínas de Drosophila , Drosophila melanogaster , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Mutação/genética , Transativadores/genética , Transativadores/metabolismo , Regulação Alostérica/genética , Substituição de Aminoácidos/genética , Animais , DNA/genética , Proteínas de Ligação a DNA/genética , Drosophila melanogaster/embriologia , Drosophila melanogaster/genética , Elementos Facilitadores Genéticos/genética , Regulação da Expressão Gênica no Desenvolvimento , Genes de Insetos/genética , Genes Reporter/genética , Proteínas de Homeodomínio/química , Proteínas de Insetos/química , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Modelos Moleculares , Mutagênese/genética , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crescimento & desenvolvimento , Termodinâmica , Transativadores/química , Fatores de Transcrição/genética , Ativação TranscricionalRESUMO
The authors investigated the use of a commercially available gentamicin enzyme-linked immunosorbent assay (ELISA) for the quantitative analysis of gentamicin concentrations in microdialysis samples. The assay demonstrated good accuracy and precision in the concentration range of 100 to 967 pg/mL. The developed quantitative ELISA is a highly sensitive and valid method for measuring gentamicin concentrations in microdialysis samples. This assay may be a useful alternative to other assay methodologies where analysis may be restricted by sample volume requirements and limited sensitivity.
Assuntos
Antibacterianos/análise , Gentamicinas/análise , Microdiálise , Animais , Cromatografia Líquida de Alta Pressão , Ensaio de Imunoadsorção Enzimática , SuínosRESUMO
Transcription of protein-coding genes by RNA polymerase II (pol II) is a highly coordinated process that requires the stepwise association of distinct protein complexes with the C-terminal domain (CTD) of Rpbl, the largest subunit of RNA pol II. Interaction of these complexes with the CTD might be subject to regulation by proteins such as Ess1 and Rsp5. Ess1, a prolyl-isomerase, binds the CTD and is thought to play a positive role in pol II transcription by generating conformational isomers of the CTD. Rsp5, a ubiquitin ligase, binds the CTD and is thought to play a negative role in transcription by mediating Rpbl ubiquitination and degradation. In this paper, we demonstrate that ESS1 and RSP5 interact genetically and that these interactions occur via RPBI. We show that over-expression of RSP5 enhances the growth defect of ess1ts cells and this effect is reversed by introducing extra copies of RPB1. Over-expression of RSP5 also mimics the sensitivity of ess1ts mutant cells to the toxicity of plasmids carrying dominant-negative CTD mutations, whereas mutations in RSP5 suppress this effect. Using a modified two-hybrid assay, we also demonstrate that Essl and Rsp5 compete directly for binding to the CTD. The results suggest a model in which Essl and Rsp5 act opposingly on pol II function to control the level of pol II available for transcription.
Assuntos
Proteínas Fúngicas/metabolismo , Peptidilprolil Isomerase/metabolismo , Proteínas Quinases , RNA Polimerase II/metabolismo , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/metabolismo , Complexos Ubiquitina-Proteína Ligase , Ligação Competitiva , Quinase 8 Dependente de Ciclina , Quinases Ciclina-Dependentes/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte , Proteínas Fúngicas/genética , Modelos Genéticos , Mutação , Peptidilprolil Isomerase de Interação com NIMA , Peptídeos/metabolismo , Peptidilprolil Isomerase/genética , Fosfoproteínas Fosfatases/metabolismo , Ligação Proteica , Estrutura Terciária de Proteína , RNA Polimerase II/química , RNA Polimerase II/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crescimento & desenvolvimento , TemperaturaRESUMO
BACKGROUND: Selective spatial regulation of gene expression lies at the core of pattern formation in the embryo. In the fruit fly Drosophila, localized transcriptional regulation accounts for much of the embryonic pattern. RESULTS: We identified a gene, partner of paired (ppa), whose properties suggest that localized receptors for protein degradation are integrated into regulatory networks of transcription factors to ensure robust spatial regulation of gene expression. We found that the Ppa protein interacts with the Pax transcription factor Paired (Prd) and contains an F-box, a motif found in receptors for ubiquitin-mediated protein degradation. In normal development, Prd functions only in cells in which ppa mRNA expression has been repressed by another segmentation protein, Even-skipped (Eve). When ppa was expressed ectopically in these cells, Prd protein, but not mRNA, levels diminished. When ppa function was removed from cells that express prd mRNA, Prd protein levels increased. CONCLUSIONS: Ppa co-ordinates Prd degradation and is important for expression of Prd to be correctly localized. In the presence of Ppa, Prd protein is targeted for degradation at sites where its mis-expression would disrupt development. In the absence of Ppa, Prd is longer-lived and regulates downstream target genes.
Assuntos
Proteínas de Transporte/genética , Proteínas de Drosophila , Drosophila/embriologia , Drosophila/genética , Proteínas de Homeodomínio/metabolismo , Sequência de Aminoácidos , Animais , Embrião não Mamífero/fisiologia , Gástrula/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Insetos/genética , Dados de Sequência Molecular , RNA Mensageiro/genética , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Transcrição GênicaRESUMO
BACKGROUND: The adequacy of intermittent and continuous infusion ceftazidime for the treatment of nosocomial pneumonia in critically ill trauma patients was assessed by analyzing ceftazidime pharmacokinetics in relation to the minimum inhibitory concentration (MIC) and treatment outcome. METHODS: Serial blood samples were obtained during ceftazidime therapy in 31 trauma patients. Ceftazidime pharmacokinetics were compared with that of previously studied healthy volunteers. Ceftazidime pharmacokinetics were analyzed according to the time above the MIC and treatment outcome. RESULTS: Critically ill trauma patients had a significantly increased volume of distribution and clearance (0.32 +/- 0.14 L/kg and 2.35 +/- 0.89 mL. min(-1). kg(-1), respectively) compared with healthy volunteers (0.21 +/- 0.03 and 1.58 +/- 0.23 mL. min(-1). kg(-1)). The time above the MIC was >/=92% of the dosing interval for all patients and treatment outcomes were similar between the two treatment groups. CONCLUSIONS: Ceftazidime pharmacokinetics are significantly altered in critically ill trauma patients. Both intermittent and continuous ceftazidime regimens were equally effective for the treatment of nosocomial pneumonia caused by less virulent bacteria.
Assuntos
Ceftazidima/administração & dosagem , Infecção Hospitalar/tratamento farmacológico , Bactérias Gram-Negativas/efeitos dos fármacos , Infecções por Bactérias Gram-Negativas/tratamento farmacológico , Pneumonia Bacteriana/tratamento farmacológico , Ferimentos e Lesões/microbiologia , Adolescente , Adulto , Idoso , Ceftazidima/farmacocinética , Cefalosporinas/administração & dosagem , Cefalosporinas/farmacocinética , Distribuição de Qui-Quadrado , Estado Terminal , Infecção Hospitalar/diagnóstico , Esquema de Medicação , Feminino , Seguimentos , Infecções por Bactérias Gram-Negativas/diagnóstico , Humanos , Infusões Intravenosas , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Pneumonia Bacteriana/diagnóstico , Probabilidade , Resultado do TratamentoRESUMO
The Ess1/Pin1 peptidyl-prolyl isomerase (PPIase) is thought to control mitosis by binding to cell cycle regulatory proteins and altering their activity. Here we isolate temperature-sensitive ess1 mutants and identify six multicopy suppressors that rescue their mitotic-lethal phenotype. None are cell cycle regulators. Instead, five encode proteins involved in transcription that bind DNA, modify chromatin structure or are regulatory subunits of RNA polymerase II. A sixth suppressor, cyclophilin A, is a member of a distinct family of PPIases that are targets of immuno suppressive drugs. We show that the expression of some but not all genes is decreased in ess1 mutants, and that Ess1 interacts with the C-terminal domain (CTD) of RNA polymerase II in vitro and in vivo. The results forge a strong link between PPIases and the transcription machinery and suggest a new model for how Ess1/Pin1 controls mitosis. In this model, Ess1 binds and isomerizes the CTD of RNA polymerase II, thus altering its interaction with proteins required for transcription of essential cell cycle genes.
Assuntos
Cromatina/metabolismo , Peptidilprolil Isomerase/metabolismo , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/genética , Fatores de Transcrição/metabolismo , Transcrição Gênica , Sequência de Aminoácidos , Animais , Cromatina/química , Cromatina/genética , Drosophila/enzimologia , Proteínas de Drosophila , Proteínas Fúngicas/metabolismo , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Humanos , Imunofilinas/metabolismo , Complexo Mediador , Mitose , Modelos Biológicos , Modelos Moleculares , Dados de Sequência Molecular , Peptidilprolil Isomerase de Interação com NIMA , Peptidilprolil Isomerase/química , Peptidilprolil Isomerase/genética , Fenótipo , Ligação Proteica , Estrutura Terciária de Proteína , RNA Polimerase II/química , RNA Polimerase II/metabolismo , Saccharomyces cerevisiae/citologia , Alinhamento de Sequência , Relação Estrutura-Atividade , Supressão Genética/genética , Proteínas de Ligação a Tacrolimo , Transcrição Gênica/genéticaRESUMO
Three families of prolyl isomerases have been identified: cyclophilins, FK506-binding proteins (FKBPs) and parvulins. All 12 cyclophilins and FKBPs are dispensable for growth in yeast, whereas the one parvulin homolog, Ess1, is essential. We report here that cyclophilin A becomes essential when Ess1 function is compromised. We also show that overexpression of cyclophilin A suppresses ess1 conditional and null mutations, and that cyclophilin A enzymatic activity is required for suppression. These results indicate that cyclophilin A and Ess1 function in parallel pathways and act on common targets by a mechanism that requires prolyl isomerization. Using genetic and biochemical approaches, we found that one of these targets is the Sin3-Rpd3 histone deacetylase complex, and that cyclophilin A increases and Ess1 decreases disruption of gene silencing by this complex. We show that conditions that favor acetylation over deacetylation suppress ess1 mutations. Our findings support a model in which Ess1 and cyclophilin A modulate the activity of the Sin3-Rpd3 complex, and excess histone deacetylation causes mitotic arrest in ess1 mutants.
Assuntos
Inativação Gênica , Histona Desacetilases/metabolismo , Peptidilprolil Isomerase/metabolismo , Proteínas Repressoras , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/enzimologia , Fatores de Transcrição/metabolismo , Acetilação , DNA Ribossômico/genética , Proteínas de Ligação a DNA/metabolismo , Estabilidade Enzimática , Regulação Enzimológica da Expressão Gênica , Regulação Fúngica da Expressão Gênica , Genes Fúngicos/genética , Genes Letais/genética , Inibidores de Histona Desacetilases , Histona Desacetilases/genética , Imunofilinas/genética , Imunofilinas/fisiologia , Mitose , Modelos Biológicos , Peptidilprolil Isomerase de Interação com NIMA , Peptidilprolil Isomerase/genética , RNA Polimerase II/metabolismo , Saccharomyces cerevisiae/genética , Supressão Genética , Proteínas de Ligação a Tacrolimo , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genéticaRESUMO
Bicoid directs pattern formation in the developing Drosophila embryo, and does so by performing two seemingly unrelated tasks; it activates transcription and represses translation. To understand how Bicoid carries out this dual role, we sought to identify Bicoid-ancillary proteins that might mediate Bicoid's function in transcription or translation. We used a customized version of the two-hybrid method and found two Bicoid-interacting proteins, Bin1 and Bin3, both of which interact with Bicoid in vitro. Bin1 is similar to a human protein (SAP18) involved in transcription regulation, and Bin3, described in this paper, is similar to a family of protein methyltransferases that modify RNA-binding proteins. Given that Bicoid's role as a translation regulator requires RNA binding, we suggest that the Bicoid-interacting methyltransferase might be important for that role. The custom two-hybrid method we used, in which Bicoid is bound to DNA via its own DNA binding domain, rather than via a fusion-protein tether, should be generally applicable to other DNA binding proteins.
Assuntos
Proteínas de Transporte/genética , Proteínas de Drosophila , Drosophila/genética , Proteínas de Homeodomínio/metabolismo , Proteínas de Insetos/genética , Transativadores/metabolismo , Fatores de Transcrição , Técnicas do Sistema de Duplo-Híbrido , Sequência de Aminoácidos , Animais , Proteínas Relacionadas à Autofagia , Proteínas de Transporte/metabolismo , Mapeamento Cromossômico , DNA Complementar/química , DNA Complementar/genética , DNA Complementar/isolamento & purificação , Drosophila/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Engenharia Genética , Proteínas de Homeodomínio/genética , Proteínas de Insetos/metabolismo , Metiltransferases/genética , Metiltransferases/metabolismo , Dados de Sequência Molecular , Ligação Proteica , Proteínas Serina-Treonina Quinases , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Transativadores/genéticaRESUMO
Respiratory depression secondary to morphine intoxication occurred in an elderly patient with chronic renal failure (CRF). It was reversed with a continuous infusion of naloxone. Approximately 11 hours after the infusion was discontinued, the patient relapsed into respiratory depression consistent with opioid intoxication. He was rechallenged with a naloxone infusion with resolution of the opioid effects. This case suggests prolonged antagonism of opioid effects inconsistent with naloxone's reported pharmacologic effects. Serum naloxone concentrations measured after the end of the infusion suggest that the drug's pharmacokinetics were significantly altered. Further research is necessary to characterize pharmacokinetic changes that occur in CRF. In the absence of this information, similar patients should be closely monitored for relapse of respiratory depression after naloxone is discontinued.
Assuntos
Falência Renal Crônica , Naloxona/uso terapêutico , Antagonistas de Entorpecentes/uso terapêutico , Idoso , Analgésicos Opioides/efeitos adversos , Humanos , Infusões Intravenosas , Masculino , Morfina/efeitos adversos , Insuficiência Respiratória/induzido quimicamente , Insuficiência Respiratória/tratamento farmacológico , Fatores de TempoRESUMO
A high-performance liquid chromatography procedure was developed to analyze ceftazidime concentrations in plasma. The procedure consisted of solid phase extraction followed by ion-pairing reverse-phase chromatography. An excellent linear relationship between ceftazidime peak height measurements and concentrations was demonstrated over the concentration range of 1-200 microg ml(-1). The advantage of this assay is the elimination of interference at the ceftazidime elution time that has been noted in previous studies and in our experience. Thus, this study describes an alternative, simple methodology that is clinically useful for analyzing ceftazidime in the research setting.
Assuntos
Ceftazidima/sangue , Cefalosporinas/sangue , Cromatografia Líquida de Alta Pressão/métodos , Humanos , Reprodutibilidade dos Testes , Espectrofotometria UltravioletaRESUMO
The Bicoid morphogen directs pattern formation along the anterior-posterior (A-P) axis of the Drosophila embryo. Bicoid is distributed in a concentration gradient that decreases exponentially from the anterior pole, however, it transcribes target genes such as hunchback in a step-function-like pattern; the expression domain is uniform and has a sharply defined posterior boundary. A 'gradient-affinity' model proposed to explain Bicoid action states that (i) cooperative gene activation by Bicoid generates the sharp on/off switch for target gene transcription and (ii) target genes with different affinities for Bicoid are expressed at different positions along the A-P axis. Using an in vivo yeast assay and in vitro methods, we show that Bicoid binds DNA with pairwise cooperativity; Bicoid bound to a strong site helps Bicoid bind to a weak site. These results support the first aspect of the model, providing a mechanism by which Bicoid generates sharp boundaries of gene expression. However, contrary to the second aspect of the model, we find no significant difference between the affinity of Bicoid for the anterior gene hunchback and the posterior gene knirps. We propose, instead, that the arrangement of Bicoids bound to the target gene presents a unique signature to the transcription machinery that, in combination with overall affinity, regulates the extent of gene transcription along the A-P axis.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Proteínas de Drosophila , Drosophila/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Genes de Insetos , Proteínas de Homeodomínio/fisiologia , Proteínas de Insetos/fisiologia , Transativadores/fisiologia , Animais , Sítios de Ligação/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Drosophila/genética , Proteínas de Homeodomínio/metabolismo , Proteínas de Insetos/metabolismo , Ligação Proteica/genética , Proteínas Repressoras/genética , Proteínas Repressoras/fisiologia , Saccharomyces cerevisiae , Transativadores/metabolismo , Ativação TranscricionalRESUMO
Pharmacological therapy, present and future, will undoubtedly continue to play a large role within the overall management of patients with severe head injury. Nevertheless, limited clinical data are available to evaluate the effect of severe head injury on pharmacokinetics. The disruption of the blood-brain barrier secondary to trauma and/or subsequent hyperosmolar therapy can be expected to result in higher than expected brain drug concentrations. Aggressive dietary protein supplementation may result in increased oxidative drug metabolism. These effects may counterbalance inhibitory influences on drug metabolism secondary to cytokine release during the acute phase response. Alterations in protein binding can also be anticipated with the hypoalbuminaemia and increases in alpha 1-acid glycoprotein typically observed in these patients. Based on studies in other patient populations, moderate hypothermia, a treatment strategy in patients with head injury, can decrease drug metabolism. The pharmacokinetics of the following drugs in patients with severe head injury have been studied: phenytoin, pentobarbital (pentobarbitone), thiopental (thiopentone), tirilazad, and the agents used as marker substrates, antipyrine, lorazepam and indocynanine green (ICG). Several studies have documented increase in metabolism over time with phenytoin, pentobarbital, thiopental, antipyrine and lorazepam. Increases in tirilazad clearance were also observed but attributed to concurrent phenytoin therapy. No changes in the pharmacokinetics of ICG were apparent following head injury. With the frequent use of potent inhibitors of drug metabolism (e.g., cimetidine, ciprofloxacin) the potential for drug interaction is high in patients with severe head injury. Additional pharmacokinetic investigations are recommended to optimise pharmacological outcomes in patients with severe head injury.
Assuntos
Anti-Inflamatórios não Esteroides/farmacocinética , Anticonvulsivantes/farmacocinética , Traumatismos Craniocerebrais/metabolismo , Hipnóticos e Sedativos/farmacocinética , Aminopirina/farmacocinética , Barreira Hematoencefálica , Interações Medicamentosas , Humanos , Lorazepam/farmacocinética , Pentobarbital/farmacocinética , Fenitoína/farmacocinética , Tiopental/farmacocinéticaRESUMO
The vast majority of patients with DiGeorge syndrome (DGS) and velocardiofacial syndrome (VCFS) have deletions of chromosomal region 22q11.2. These patients exhibit broad and variable phenotypes that include conotruncal cardiac defects, hypocalcemia, palatal and facial anomalies and developmental delay. Most of these abnormalities are thought to be due to defects in neural crest cell migration or differentiation. We have identified a homeobox-containing gene, Goosecoid-like (GSCL), that is in the region within 22q11 that is deleted most consistently in patients with DGS/VCFS. The GSCL gene is expressed in a limited number of adult tissues as well as in early human development, and is a member of a family of homeobox genes in vertebrates that includes Goosecoid and GSX. In this report, we present functional studies of the GSCL protein and determine the expression pattern of the GSCL gene in mouse embryos. We demonstrate that GSCL exhibits DNA sequence-specific recognition of sites bound by the Drosophila anterior morphogen, Bicoid. Several of these sites (TAATCCC) were found in the 5' upstream region of the GSCL gene itself, and we present evidence suggesting that GSCL might regulate its own transcription. In situ hybridization revealed that the mouse ortholog of GSCL, Gscl, is expressed in the brain starting as early as embryonic day 9.5, and expression continues in adults. This expression pattern is consistent with GSCL having either an indirect role in the development of neural crest-derived structures or a direct role in a subset of the phenotype observed in DGS/VCFS, such as learning disorders or psychiatric disease.
Assuntos
Anormalidades Múltiplas/genética , DNA/genética , Síndrome de DiGeorge/genética , Deleção de Genes , Genes Homeobox , Adulto , Sequência de Aminoácidos , Animais , Sequência de Bases , Sítios de Ligação/genética , Encéfalo/embriologia , Encéfalo/metabolismo , Primers do DNA/genética , Drosophila/genética , Proteínas de Drosophila , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Genes de Insetos , Proteínas de Homeodomínio/genética , Humanos , Hibridização In Situ , Camundongos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Gravidez , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico , Síndrome , Transativadores/genéticaRESUMO
The pharmacokinetics of fluconazole in nine adult patients with severe (30 to 95% total body surface area) burns were studied. There was no significant difference in half-life (t1/2), clearance (CL), or volume of distribution (V) over time in five patients on days 3 and 8 of the study (P > 0.05). Combined parameter estimates (means +/- standard deviations) for all nine patients for the two study periods were as follows: t1/2, 24.4 +/- 5.8 h; CL, 0.36 +/- 0.09 ml/min/kg; and V, 0.72 +/- 0.12 liters/kg. These estimates of t1/2 and CL in burn patients were approximately 13% shorter and 30% more rapid, respectively, than the most extreme estimates reported for other populations.
Assuntos
Antifúngicos/farmacocinética , Queimaduras/metabolismo , Fluconazol/farmacocinética , Adulto , Idoso , Área Sob a Curva , Cromatografia Líquida de Alta Pressão , Método Duplo-Cego , Feminino , Meia-Vida , Humanos , Masculino , Pessoa de Meia-IdadeAssuntos
Divisão Celular , Proteínas de Drosophila , Peptidilprolil Isomerase/genética , Peptidilprolil Isomerase/fisiologia , Animais , Drosophila , Proteínas Fúngicas/genética , Proteínas Fúngicas/fisiologia , Humanos , Proteínas de Insetos/genética , Proteínas de Insetos/fisiologia , Peptidilprolil Isomerase de Interação com NIMARESUMO
OBJECTIVE: To determine the incidence of thrombocytopenia (< 100 platelets x 10(3)/mm3) and potential risk factors, including medications, associated with the development of thrombocytopenia in critically ill trauma patients. DESIGN: Prospective, observational study. SETTING: A 20-bed trauma intensive care unit (ICU) at a university hospital. PATIENTS: Sixty-three critically ill trauma patients without baseline thrombocytopenia admitted to the trauma ICU for at least 48 hours. INTERVENTIONS: Patients were followed for up to 14 days. Platelet counts were determined daily. The following data were collected and analyzed as potential risk factors for the development of thrombocytopenia: medications, age, sex, race, trauma score, mode and type of injury, alcohol history, units of packed red blood cells (PRBC) and platelets transfused, surgical procedures, duration of ICU stay, and the development of systemic inflammatory response syndrome or disseminated intravascular coagulation. RESULTS: Thrombocytopenia occurred in 26 (41%) of the patients. Among risk factors studied, nonhead injury, age, trauma score, duration of ICU stay, and the number of PRBC transfusions were significantly associated with the development of thrombocytopenia (p < 0.05). However, nonhead injury, age, and trauma score were useful variables in predicting the development of thrombocytopenia by using multivariate analysis. Medications were not associated with the development of thrombocytopenia. CONCLUSIONS: The type of injury sustained, the quantity of platelet-deficient, transfusions, and age are the greatest risk factors associated with the development of thrombocytopenia in critically ill trauma patients. Drug-induced thrombocytopenia appears to play a minor role in the development of thrombocytopenia; therefore, medications should not be automatically discontinued or substituted when thrombocytopenia occurs.
Assuntos
Trombocitopenia/epidemiologia , Ferimentos e Lesões/sangue , Adolescente , Adulto , Fatores Etários , Idoso , Criança , Traumatismos Craniocerebrais/sangue , Traumatismos Craniocerebrais/complicações , Cuidados Críticos , Estado Terminal , Feminino , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Fatores de Risco , Trombocitopenia/etiologia , Reação Transfusional , Ferimentos e Lesões/complicaçõesRESUMO
Recent studies in yeast, Drosophila and humans have revealed the existence of a highly conserved gene encoding a novel protein, Dodo, comprised of four modules: a WW domain, involved in protein-protein interactions, a peptidyl-prolyl cis-trans isomerase (PPIase) domain belonging to a recently described third family of PPIases involved in protein folding and unfolding, a nuclear localization motif and finally, a long, surface-exposed alpha-helix that is likely to be involved in binding to a cell cycle serine/threonine kinase. The genetic, molecular, biochemical and structural data are reviewed in the context of the potential biological properties of this new protein family.