RESUMEN
Loss of the CDK inhibitor p27(KIP1) is widely linked with poor prognosis in human cancer. In Wnt10b-expressing mammary tumors, levels of p27(KIP1) were extremely low; conversely, Wnt10b-null mammary cells expressed high levels of this protein, suggesting Wnt-dependent regulation of p27(KIP1). Interestingly we found that Wnt-induced turnover of p27(KIP1) was independent from classical SCF(SKP2)-mediated degradation in both mouse and human cells. Instead, turnover required Cullin 4A and Cullin 4B, components of an alternative E3 ubiquitin ligase induced in response to active Wnt signaling. We found that CUL4A was a novel Wnt target gene in both mouse and human cells and that CUL4A physically interacted with p27(KIP1) in Wnt-responding cells. We further demonstrated that both Cul4A and Cul4B were required for Wnt-induced p27(KIP1) degradation and S-phase progression. CUL4A and CUL4B are therefore components of a conserved Wnt-induced proteasome targeting (WIPT) complex that regulates p27(KIP1) levels and cell cycle progression in mammalian cells.
Asunto(s)
Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Proteínas Quinasas Asociadas a Fase-S/metabolismo , Transducción de Señal/fisiología , Proteínas Wnt/metabolismo , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Ciclo Celular/genética , Ciclo Celular/fisiología , Línea Celular , Línea Celular Tumoral , Inmunoprecipitación de Cromatina , Proteínas Cullin/genética , Proteínas Cullin/metabolismo , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/genética , Femenino , Humanos , Immunoblotting , Inmunoprecipitación , Carioferinas/genética , Carioferinas/metabolismo , Masculino , Ratones , Ratones Transgénicos , Mutación , Complejo de la Endopetidasa Proteasomal/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Proteínas Quinasas Asociadas a Fase-S/genética , Transducción de Señal/genética , Proteínas Wnt/genética , Proteína Exportina 1RESUMEN
Converging evidence from basic and clinical studies suggests a role for proinflammatory cytokines in cancer-related fatigue, although the etiology of elevated inflammatory processes is unclear. We examined single nucleotide polymorphisms (SNPs) in the promoters of cytokine genes as genetic risk factors for cytokine-related fatigue in 33 fatigued and 14 non-fatigued breast cancer survivors, focusing on promoter sequence polymorphisms in IL1B and IL6 associated with differential expression of proinflammatory cytokines. Predictors of fatigue included presence of at least one cytosine at IL1B -511 (95%CI=0.91-16.6, p=.007) and homozygosity for either variant of the IL6 -174 genotype (G/G or C/C; 95%CI=1.12-17.9, p=.027). Associations between fatigue status and IL1B genotype remained significant after covariate adjustment for demographic, biobehavioral and treatment-related factors. These findings provide preliminary evidence that polymorphisms in IL1B may serve as a potential risk factor for persistent fatigue in the aftermath of cancer.
Asunto(s)
Neoplasias de la Mama/genética , Fatiga/genética , Interleucina-1beta/genética , Interleucina-6/genética , Neoplasias de la Mama/complicaciones , Distribución de Chi-Cuadrado , Fatiga/etiología , Femenino , Predisposición Genética a la Enfermedad , Genotipo , Humanos , Persona de Mediana Edad , Oportunidad Relativa , Selección de Paciente , Polimorfismo de Nucleótido Simple , Encuestas y Cuestionarios , SobrevivientesRESUMEN
BACKGROUND: Inflammation is associated with increased risk of cardiovascular disorders, arthritis, diabetes mellitus, and mortality. The effects of sleep loss on the cellular and genomic mechanisms that contribute to inflammatory cytokine activity are not known. METHODS: In 30 healthy adults, monocyte intracellular proinflammatory cytokine production was repeatedly assessed during the day across 3 baseline periods and after partial sleep deprivation (awake from 11 pm to 3 am). We analyzed the impact of sleep loss on transcription of proinflammatory cytokine genes and used DNA microarray analyses to characterize candidate transcription-control pathways that might mediate the effects of sleep loss on leukocyte gene expression. RESULTS: In the morning after a night of sleep loss, monocyte production of interleukin 6 and tumor necrosis factor alpha was significantly greater compared with morning levels following uninterrupted sleep. In addition, sleep loss induced a more than 3-fold increase in transcription of interleukin 6 messenger RNA and a 2-fold increase in tumor necrosis factor alpha messenger RNA. Bioinformatics analyses suggested that the inflammatory response was mediated by the nuclear factor kappaB inflammatory signaling system as well as through classic hormone and growth factor response pathways. CONCLUSIONS: Sleep loss induces a functional alteration of the monocyte proinflammatory cytokine response. A modest amount of sleep loss also alters molecular processes that drive cellular immune activation and induce inflammatory cytokines; mapping the dynamics of sleep loss on molecular signaling pathways has implications for understanding the role of sleep in altering immune cell physiologic characteristics. Interventions that target sleep might constitute new strategies to constrain inflammation with effects on inflammatory disease risk.
Asunto(s)
Perfilación de la Expresión Génica , Interleucina-6/metabolismo , Privación de Sueño/genética , Privación de Sueño/inmunología , Factor de Necrosis Tumoral alfa/metabolismo , Adulto , Biomarcadores/metabolismo , Femenino , Citometría de Flujo , Humanos , Masculino , Persona de Mediana Edad , Monocitos/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Mensajero/metabolismo , Privación de Sueño/metabolismo , Transcripción GenéticaRESUMEN
PURPOSE: This study seeks to define immunologic and inflammatory variables associated with persistent post-treatment fatigue in breast cancer survivors. EXPERIMENTAL DESIGN: Leukocyte subsets, plasma inflammatory markers, and ex vivo proinflammatory cytokine production were assessed in 50 fatigued and nonfatigued breast cancer survivors recruited > or = 2 years after successful primary therapy. Multivariate statistical analyses were used to define a composite immunologic biomarker of fatigue risk. RESULTS: Fatigued breast cancer survivors were distinguished from nonfatigued survivors by increased ex vivo monocyte production of interleukin (IL)-6 and tumor necrosis factor-alpha following lipopolysaccharide stimulation, elevated plasma IL-1ra and soluble IL-6 receptor (sIL-6R/CD126), decreased monocyte cell-surface IL-6R, and decreased frequencies of activated T lymphocytes and myeloid dendritic cells in peripheral blood (all P < 0.05). An inverse correlation between sIL-6R and cell-surface IL-6R was consistent with inflammation-mediated shedding of IL-6R, and in vitro studies confirmed that proinflammatory cytokines induced such shedding. Multivariate linear discriminant function analysis identified two immunologic markers, the ratio of sIL-6R to monocyte-associated IL-6R and decreased circulating CD69+ T lymphocytes, as highly diagnostic of fatigue (P = 0.0005), with cross-validation estimates indicating 87% classification accuracy (sensitivity = 0.83; specificity = 0.83). CONCLUSION: These results extend links between fatigue and inflammatory markers to show a functional alteration in proinflammatory cytokine response to lipopolysaccharide and define a prognostic biomarker of behavioral fatigue.
Asunto(s)
Neoplasias de la Mama/fisiopatología , Fatiga/fisiopatología , Inflamación , Biomarcadores/sangre , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Citocinas/sangre , Femenino , Humanos , Recuento de Linfocitos , Persona de Mediana Edad , SobrevivientesRESUMEN
Type I interferons (IFN-alpha and -beta) play a key role in anti-viral immunity, and we sought to define the molecular mechanisms by which the sympathetic nervous system (SNS) inhibits their effects. In peripheral blood leukocytes and plasmacytoid dendritic cells (pDC2), induction of interferon anti-viral activity by double-stranded RNA (poly-I:C) or CpG DNA was substantially inhibited by norepinephrine and by pharmacologic activation of the cAMP/PKA signaling pathway. This effect was specific to Type I interferons and driven by PKA-mediated repression of IFNA and IFNB gene transcription. Luciferase reporter analyses identified tandem interferon response factor-binding sites in positive regulatory domains I and III of the IFNB promoter as a key target of PKA inhibition. PKA suppression of Type I interferons was associated with impaired transcription of interferon response genes supporting the "anti-viral state", and was sufficient to account for norepinephrine-induced enhancement of HIV-1 replication. Given the ubiquitous role of Type I interferons in containing viral replication, PKA-mediated inhibition of IFN transcription could explain the stimulatory effects of catecholamines on a broad range of viral pathogens.
Asunto(s)
Regulación de la Expresión Génica/inmunología , Infecciones por VIH/inmunología , VIH-1/inmunología , Interferón Tipo I/inmunología , Norepinefrina/fisiología , Replicación Viral/inmunología , Sistema Nervioso Autónomo/inmunología , Células Cultivadas , Islas de CpG/inmunología , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Células Dendríticas/virología , VIH-1/genética , Humanos , Inmunidad Innata/fisiología , Interferón Tipo I/genética , Interferón Tipo I/metabolismo , Leucocitos/inmunología , Leucocitos/metabolismo , Leucocitos/virología , Neuroinmunomodulación , Norepinefrina/inmunología , Regiones Promotoras Genéticas/genética , ARN Bicatenario/inmunología , Transducción de Señal/fisiología , Transcripción Genética/fisiología , Replicación Viral/genéticaRESUMEN
Reactivation of Kaposi's sarcoma-associated herpesvirus (KSHV) lytic replication is mediated by the viral RTA transcription factor, but little is known about the physiological processes controlling its expression or activity. Links between autonomic nervous system activity and AIDS-associated Kaposi's sarcoma led us to examine the potential influence of catecholamine neurotransmitters. Physiological concentrations of epinephrine and norepinephrine efficiently reactivated lytic replication of KSHV in latently infected primary effusion lymphoma cells via beta-adrenergic activation of the cellular cyclic AMP/protein kinase A (PKA) signaling pathway. Effects were blocked by PKA antagonists and mimicked by pharmacological and physiological PKA activators (prostaglandin E2 and histamine) or overexpression of the PKA catalytic subunit. PKA up-regulated RTA gene expression, enhanced activity of the RTA promoter, and posttranslationally enhanced RTA's trans-activating capacity for its own promoter and heterologous lytic promoters (e.g., the viral PAN gene). Mutation of predicted phosphorylation targets at RTA serines 525 and 526 inhibited PKA-mediated enhancement of RTA trans-activating capacity. Given the high catecholamine levels at sites of KSHV latency such as the vasculature and lymphoid organs, these data suggest that beta-adrenergic control of RTA might constitute a significant physiological regulator of KSHV lytic replication. These findings also suggest novel therapeutic strategies for controlling the activity of this oncogenic gammaherpesvirus in vivo.