RESUMO
OBJECTIVES: Pharmacogenetics (PGx) is increasingly important in individualizing therapeutic management plans, but is often implemented apart from other types of medication clinical decision support (CDS). The lack of integration of PGx into existing CDS may result in incomplete interaction information, which may pose patient safety concerns. We sought to develop a cloud-based orchestrated medication CDS service that integrates PGx with a broad set of drug screening alerts and evaluate it through a clinician utility study. METHODS: We developed the PillHarmonics service for implementation per the CDS Hooks protocol, algorithmically integrating a wide range of drug interaction knowledge using cloud-based screening services from First Databank (drug-drug/allergy/condition), PharmGKB (drug-gene), and locally curated content (drug-renal/hepatic/race). We performed a user study, presenting 13 clinicians and pharmacists with a prototype of the system's usage in synthetic patient scenarios. We collected feedback via a standard questionnaire and structured interview. RESULTS: Clinician assessment of PillHarmonics via the Technology Acceptance Model questionnaire shows significant evidence of perceived utility. Thematic analysis of structured interviews revealed that aggregated knowledge, concise actionable summaries, and information accessibility were highly valued, and that clinicians would use the service in their practice. CONCLUSION: Medication safety and optimizing efficacy of therapy regimens remain significant issues. A comprehensive medication CDS system that leverages patient clinical and genomic data to perform a wide range of interaction checking and presents a concise and holistic view of medication knowledge back to the clinician is feasible and perceived as highly valuable for more informed decision-making. Such a system can potentially address many of the challenges identified with current medication-related CDS.
Assuntos
Sistemas de Apoio a Decisões Clínicas , Farmacogenética , Humanos , Computação em NuvemRESUMO
Excessive phototransduction signaling is thought to be involved in light-induced and inherited retinal degeneration. Using knockout mice with defects in rhodopsin shut-off and transducin signaling, we show that two different pathways of photoreceptor-cell apoptosis are induced by light. Bright light induces apoptosis that is independent of transducin and accompanied by induction of the transcription factor AP-1. By contrast, low light induces an apoptotic pathway that requires transducin. We also provide evidence that additional genetic factors regulate sensitivity to light-induced damage. Our use of defined mouse mutants resolves some of the complexity underlying the mechanisms that regulate susceptibility to retinal degeneration.
Assuntos
Apoptose , Proteínas do Olho , Luz/efeitos adversos , Retina/efeitos da radiação , Animais , Arrestina/genética , Arrestina/metabolismo , Proteínas de Transporte , Dexametasona/metabolismo , Receptor Quinase 1 Acoplada a Proteína G , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Mutação , Células Fotorreceptoras de Vertebrados/fisiologia , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Proteínas/metabolismo , Retina/metabolismo , Retina/fisiopatologia , Rodopsina/metabolismo , Transdução de Sinais , Fator de Transcrição AP-1/antagonistas & inibidores , Fator de Transcrição AP-1/metabolismo , Transducina/metabolismo , cis-trans-IsomerasesRESUMO
Fragile X syndrome (FXS) is the most common form of hereditary mental retardation. FXS patients have a deficit for the fragile X mental retardation protein (FMRP) that results in abnormal neuronal dendritic spine morphology and behavioral phenotypes, including sleep abnormalities. In a Drosophila model of FXS, flies lacking the dfmr1 protein (dFMRP) have abnormal circadian rhythms apparently as a result of altered clock output. In this study, we present biochemical and genetic evidence that dFMRP interacts with a known clock output component, the LARK RNA-binding protein. Our studies demonstrate physical interactions between dFMRP and LARK, that the two proteins are present in a complex in vivo, and that LARK promotes the stability of dFMRP. Furthermore, we show genetic interactions between the corresponding genes indicating that dFMRP and LARK function together to regulate eye development and circadian behavior.
Assuntos
Comportamento Animal/fisiologia , Ritmo Circadiano/fisiologia , Proteínas de Drosophila/metabolismo , Drosophila/fisiologia , Olho/crescimento & desenvolvimento , Proteína do X Frágil da Deficiência Intelectual/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Ritmo Circadiano/genética , Modelos Animais de Doenças , Drosophila/crescimento & desenvolvimento , Proteínas de Drosophila/genética , Proteína do X Frágil da Deficiência Intelectual/genética , Síndrome do Cromossomo X Frágil , Larva/metabolismo , Proteínas de Ligação a RNA/genéticaRESUMO
Loss of Fragile X mental retardation protein (FMRP) function causes the highly prevalent Fragile X syndrome [1 and 2]. Identifying targets for the RNA binding FMRP is a major challenge and an important goal of research into the pathology of the disease. Perturbations in neuronal development and circadian behavior are seen in Drosophila dfmr1 mutants. Here we show that regulation of the actin cytoskeleton is under dFMRP control. dFMRP binds the mRNA of the Drosophila profilin homolog and negatively regulates Profilin protein expression. An increase in Profilin mimics the phenotype of dfmr1 mutants. Conversely, decreasing Profilin levels suppresses dfmr1 phenotypes. These data place a new emphasis on actin misregulation as a major problem in fmr1 mutant neurons.
Assuntos
Actinas/metabolismo , Encéfalo/fisiologia , Proteínas Contráteis/genética , Proteínas de Drosophila/metabolismo , Proteínas dos Microfilamentos/genética , Proteínas de Ligação a RNA/metabolismo , Animais , Células Cultivadas , Proteínas Contráteis/metabolismo , Proteínas de Drosophila/genética , Proteína do X Frágil da Deficiência Intelectual , Regulação da Expressão Gênica , Proteínas dos Microfilamentos/metabolismo , Mutação , Neurônios/metabolismo , Neurônios/patologia , Profilinas , Proteínas de Ligação a RNA/genética , Colículos Superiores/metabolismo , Colículos Superiores/patologia , Proteínas rac de Ligação ao GTP/genética , Proteínas rac de Ligação ao GTP/metabolismoRESUMO
Mitogen-activated protein kinases/extracellular signal regulated kinases (MAPKs/ERKs) are typically thought to be soluble cytoplasmic enzymes that translocate to the nucleus subsequent to their phosphorylation by their activating kinases or mitogen-activated protein/extracellular signal regulated kinase kinase. We report here the first example of nuclear translocation of a MAPK that occurs via temporally regulated exit from a membranous organelle. Confocal microscopy examining the subcellular localization of ERK3 in several cell lines indicated that this enzyme was targeted to the Golgi/endoplasmic reticulum Golgi intermediate compartment. Deletion analysis of green fluorescent protein (GFP)-ERK3 uncovered a nuclear form that was carboxy-terminally truncated and established a Golgi targeting motif at the carboxy terminus. Immunoblot analysis of cells treated with the proteasome inhibitor MG132 further revealed two cleavage products, suggesting that in vivo, carboxy-terminal cleavage of the full-length protein controls its subcellular localization. In support of this hypothesis, we found that deletion of a small region rich in acidic residues within the carboxy terminus eliminated both the cleavage and nuclear translocation of GFP-ERK3. Finally, cell cycle synchronization studies revealed that the subcellular localization of ERK3 is temporally regulated. These data suggest a novel mechanism for the localization of an MAPK family member, ERK3, in which cell cycle-regulated, site-specific proteolysis generates the nuclear form of the protein.
Assuntos
Transporte Ativo do Núcleo Celular/fisiologia , Proteína Quinase 6 Ativada por Mitógeno/metabolismo , Motivos de Aminoácidos , Animais , Ciclo Celular , Linhagem Celular , Núcleo Celular/metabolismo , Complexo de Golgi/metabolismo , Humanos , Lisina/metabolismo , Proteína Quinase 6 Ativada por Mitógeno/genética , Sinais Direcionadores de Proteínas , Ratos , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismoRESUMO
Explicit biochemical models have been elaborated for the circadian oscillators of cyanobacterial, fungal, insect, and mammalian species. In contrast, much remains to be learned about how such circadian oscillators regulate rhythmic physiological processes. This article summarizes contemporary genetic and biochemical strategies that are useful for identifying gene products that have a role in circadian control.
Assuntos
Relógios Biológicos/efeitos dos fármacos , Ritmo Circadiano/efeitos dos fármacos , Genes de Insetos/fisiologia , Animais , Relógios Biológicos/genética , Química Encefálica , Ritmo Circadiano/genética , AMP Cíclico/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Proteínas de Ligação a DNA/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/fisiologia , Drosophila melanogaster/genética , Proteína do X Frágil da Deficiência Intelectual/genética , Proteína do X Frágil da Deficiência Intelectual/fisiologia , Larva/fisiologia , Sistema de Sinalização das MAP Quinases/fisiologia , Análise em Microsséries/métodos , Atividade Motora/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/fisiologia , Núcleo Supraquiasmático/fisiologiaRESUMO
Both host and viral factors play an important role in the pathogenesis of human immunodeficiency virus (HIV)-associated bran injury. In this study, the authors examined the interactions between tumor necrosis factor (TNF)-alpha, CXCR4, the alpha chemokine receptor, and three HIV isolates, including the T-tropic viruses, HIV-1(MN) and HIV-1(IIIB), and the dual tropic virus, HIV-1(89.6). The authors show by flow cytometry that treatment of differentiated SK-N-MC cells with TNF-alpha induces a significant increase in the cell surface expression of CXCR4 in a time- and dose-dependent manner. The effect is partly regulated at the level of transcription. To assess the biological significance of this finding, we show that TNF-alpha potentiates the ability of the above mentioned HIV isolates to induce neuronal apoptosis and that the effect is significantly reduced by pretreating cells with monoclonal antibodies to either CXCR4 and TNF-alpha. Together these results suggest that TNF-alpha may render neuronal cells vulnerable to the apoptotic effects of HIV by increasing the cell surface expression of CXCR4 and thus identify another mechanism by which TNF-alpha contributes to the pathogenesis of HIV-associated brain injury.
Assuntos
Receptores CXCR4/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Apoptose , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , HIV-1/patogenicidade , Humanos , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Neurônios/virologia , Fatores de TempoRESUMO
Past studies have implicated the Drosophila LARK protein in the circadian control of adult eclosion behavior. LARK has a broad tissue pattern of distribution, and is pan-neuronal in the differentiated brain. In certain peptidergic neurons, LARK abundance changes in a circadian manner. However, the precise cellular requirement for LARK, with respect to circadian behavior, is still not known. To explore this issue, we employed the GAL4/UAS binary expression system to increase LARK abundance in defined neuronal cell types. Interestingly, LARK expression in Crustacean Cardioactive Peptide (CCAP) neurons caused an early-eclosion phenotype, whereas a similar perturbation in the Eclosion Hormone (EH) cells resulted in abnormally late peaks of eclosion. Surprisingly, LARK expression in Pigment Dispersing Factor (PDF)- or TIMELESS (TIM)-containing clock neurons caused behavioral arrhythmicity, even though clock protein cycling was found to be normal in these flies. Although the observed effects of LARK expression mirrored those seen with genetic ablation of the relevant peptidergic populations, there was no evidence of defective cell development or morphology. This suggests that an alteration of cell function rather than cell death is the cause of the aberrant phenotypes. Diminished PDF immunoreactivity in flies expressing LARK in the PDF neurons suggests that an effect on neuropeptide synthesis, transport, or release may contribute to the observed arrhythmicity. Importantly, the expression of LARK in several other cell populations did not have detectable effects on development, viability or behavior, indicating a specificity of action within certain cell types.