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2.
Cell ; 142(2): 309-19, 2010 Jul 23.
Artículo en Inglés | MEDLINE | ID: mdl-20655471

RESUMEN

Global energy balance in mammals is controlled by the actions of circulating hormones that coordinate fuel production and utilization in metabolically active tissues. Bone-derived osteocalcin, in its undercarboxylated, hormonal form, regulates fat deposition and is a potent insulin secretagogue. Here, we show that insulin receptor (IR) signaling in osteoblasts controls osteoblast development and osteocalcin expression by suppressing the Runx2 inhibitor Twist2. Mice lacking IR in osteoblasts have low circulating undercarboxylated osteocalcin and reduced bone acquisition due to decreased bone formation and deficient numbers of osteoblasts. With age, these mice develop marked peripheral adiposity and hyperglycemia accompanied by severe glucose intolerance and insulin resistance. The metabolic abnormalities in these mice are improved by infusion of undercarboxylated osteocalcin. These results indicate the existence of a bone-pancreas endocrine loop through which insulin signaling in the osteoblast ensures osteoblast differentiation and stimulates osteocalcin production, which in turn regulates insulin sensitivity and pancreatic insulin secretion.


Asunto(s)
Osteoblastos/metabolismo , Osteogénesis , Receptor de Insulina/metabolismo , Adiposidad , Animales , Diferenciación Celular , Proliferación Celular , Supervivencia Celular , Células Cultivadas , Resistencia a la Insulina , Masculino , Ratones , Osteoblastos/citología , Osteocalcina/metabolismo , Proteínas Represoras/metabolismo , Transducción de Señal , Proteína 1 Relacionada con Twist/metabolismo
3.
Circulation ; 146(2): 94-109, 2022 Jul 12.
Artículo en Inglés | MEDLINE | ID: mdl-35652342

RESUMEN

BACKGROUND: Cardiovascular health (CVH) from young adulthood is strongly associated with an individual's future risk of cardiovascular disease (CVD) and total mortality. Defining epigenomic biomarkers of lifelong CVH exposure and understanding their roles in CVD development may help develop preventive and therapeutic strategies for CVD. METHODS: In 1085 CARDIA study (Coronary Artery Risk Development in Young Adults) participants, we defined a clinical cumulative CVH score that combines body mass index, blood pressure, total cholesterol, and fasting glucose measured longitudinally from young adulthood through middle age over 20 years (mean age, 25-45). Blood DNA methylation at >840 000 methylation markers was measured twice over 5 years (mean age, 40 and 45). Epigenome-wide association analyses on the cumulative CVH score were performed in CARDIA and compared in the FHS (Framingham Heart Study). We used penalized regression to build a methylation-based risk score to evaluate the risk of incident coronary artery calcification and clinical CVD events. RESULTS: We identified 45 methylation markers associated with cumulative CVH at false discovery rate <0.01 (P=4.7E-7-5.8E-17) in CARDIA and replicated in FHS. These associations were more pronounced with methylation measured at an older age. CPT1A, ABCG1, and SREBF1 appeared as the most prominent genes. The 45 methylation markers were mostly located in transcriptionally active chromatin and involved lipid metabolism, insulin secretion, and cytokine production pathways. Three methylation markers located in genes SARS1, SOCS3, and LINC-PINT statistically mediated 20.4% of the total effect between CVH and risk of incident coronary artery calcification. The methylation risk score added information and significantly (P=0.004) improved the discrimination capacity of coronary artery calcification status versus CVH score alone and showed association with risk of incident coronary artery calcification 5 to 10 years later independent of cumulative CVH score (odds ratio, 1.87; P=9.66E-09). The methylation risk score was also associated with incident clinical CVD in FHS (hazard ratio, 1.28; P=1.22E-05). CONCLUSIONS: Cumulative CVH from young adulthood contributes to midlife epigenetic programming over time. Our findings demonstrate the role of epigenetic markers in response to CVH changes and highlight the potential of epigenomic markers for precision CVD prevention, and earlier detection of subclinical CVD, as well.


Asunto(s)
Enfermedades Cardiovasculares , Enfermedad de la Arteria Coronaria , Adulto , Biomarcadores , Enfermedades Cardiovasculares/diagnóstico , Enfermedades Cardiovasculares/epidemiología , Enfermedades Cardiovasculares/genética , Metilación de ADN , Humanos , Incidencia , Persona de Mediana Edad , Medición de Riesgo , Factores de Riesgo , Adulto Joven
4.
Proc Natl Acad Sci U S A ; 117(46): 28847-28858, 2020 11 17.
Artículo en Inglés | MEDLINE | ID: mdl-33127760

RESUMEN

CHD7 encodes an ATP-dependent chromatin remodeling factor. Mutation of this gene causes multiple developmental disorders, including CHARGE (Coloboma of the eye, Heart defects, Atresia of the choanae, Retardation of growth/development, Genital abnormalities, and Ear anomalies) syndrome, in which conotruncal anomalies are the most prevalent form of heart defects. How CHD7 regulates conotruncal development remains unclear. In this study, we establish that deletion of Chd7 in neural crest cells (NCCs) causes severe conotruncal defects and perinatal lethality, thus providing mouse genetic evidence demonstrating that CHD7 cell-autonomously regulates cardiac NCC development, thereby clarifying a long-standing controversy in the literature. Using transcriptomic analyses, we show that CHD7 fine-tunes the expression of a gene network that is critical for cardiac NCC development. To gain further molecular insights into gene regulation by CHD7, we performed a protein-protein interaction screen by incubating recombinant CHD7 on a protein array. We find that CHD7 directly interacts with several developmental disorder-mutated proteins including WDR5, a core component of H3K4 methyltransferase complexes. This direct interaction suggested that CHD7 may recruit histone-modifying enzymes to target loci independently of its remodeling functions. We therefore generated a mouse model that harbors an ATPase-deficient allele and demonstrates that mutant CHD7 retains the ability to recruit H3K4 methyltransferase activity to its targets. Thus, our data uncover that CHD7 regulates cardiovascular development through ATP-dependent and -independent activities, shedding light on the etiology of CHD7-related congenital disorders. Importantly, our data also imply that patients carrying a premature stop codon versus missense mutations will likely display different molecular alterations; these patients might therefore require personalized therapeutic interventions.


Asunto(s)
Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Corazón/embriología , Adenosina Trifosfato/metabolismo , Alelos , Animales , Síndrome CHARGE/genética , Ensamble y Desensamble de Cromatina/genética , ADN Helicasas/metabolismo , Modelos Animales de Enfermedad , Embrión de Mamíferos/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Cardiopatías Congénitas/genética , Ratones , Ratones Noqueados , Mutación , Cresta Neural/embriología , Cresta Neural/metabolismo , Organogénesis/fisiología
5.
Lab Invest ; 102(11): 1236-1244, 2022 11.
Artículo en Inglés | MEDLINE | ID: mdl-35907952

RESUMEN

Given the gut microbiome's rise as a potential frontier in cancer pathogenesis and therapy, leveraging microbial analyses in the study of breast tumor progression and treatment could unveil novel interactions between commensal bacteria and disease outcomes. In breast cancer, the Hedgehog (Hh) signaling pathway is a potential target for treatment due to its aberrant activation leading to poorer prognoses and drug resistance. There are limited studies that have investigated the influences of orally administered cancer therapeutics, such as Vismodegib (a pharmacological, clinically used Hh inhibitor) on the gut microbiota. Using a 4T1 mammary carcinoma mouse model and 16 S rRNA sequencing, we longitudinally mapped alterations in immunomodulating gut microbes during mammary tumor development. Next, we identified changes in the abundance of commensal microbiota in response to Vismodegib treatment of 4T1 mammary tumor-bearing mice. In addition to remodeling gut microbiota, Vismodegib treatment elicited an increase in proliferative CD8+ T cells in the colonic immune network, without any remarkable gastrointestinal-associated side effects. To our knowledge, this is the first study to assess longitudinal changes in the gut microbiome during mammary tumor development and progression. Our study also pioneers an investigation of the dynamic effects of an orally delivered Hh inhibitor on the gut microbiome and the gut-associated immune-regulatory adaptive effector CD8+ T cells. These findings inform future comprehensive studies on the consortium of altered microbes that can impact potential systemic immunomodulatory roles of Vismodegib.


Asunto(s)
Carcinoma , Microbioma Gastrointestinal , Ratones , Animales , Microbioma Gastrointestinal/fisiología , Proteínas Hedgehog , Linfocitos T CD8-positivos , Modelos Animales de Enfermedad
6.
Nucleic Acids Res ; 48(18): 10342-10352, 2020 10 09.
Artículo en Inglés | MEDLINE | ID: mdl-32894284

RESUMEN

Ribosomal DNA (rDNA) consists of highly repeated sequences that are prone to incurring damage. Delays or failure of rDNA double-strand break (DSB) repair are deleterious, and can lead to rDNA transcriptional arrest, chromosomal translocations, genomic losses, and cell death. Here, we show that the zinc-finger transcription factor GLI1, a terminal effector of the Hedgehog (Hh) pathway, is required for the repair of rDNA DSBs. We found that GLI1 is activated in triple-negative breast cancer cells in response to ionizing radiation (IR) and localizes to rDNA sequences in response to both global DSBs generated by IR and site-specific DSBs in rDNA. Inhibiting GLI1 interferes with rDNA DSB repair and impacts RNA polymerase I activity and cell viability. Our findings tie Hh signaling to rDNA repair and this heretofore unknown function may be critically important in proliferating cancer cells.


Asunto(s)
ADN Ribosómico/genética , Proteínas Hedgehog/genética , ARN Polimerasa I/genética , Neoplasias de la Mama Triple Negativas/radioterapia , Proteína con Dedos de Zinc GLI1/genética , Proteínas de Ciclo Celular/genética , Nucléolo Celular/genética , Nucléolo Celular/efectos de la radiación , Proliferación Celular/efectos de la radiación , Supervivencia Celular/efectos de la radiación , Roturas del ADN de Doble Cadena/efectos de la radiación , Daño del ADN/efectos de la radiación , Reparación del ADN/efectos de la radiación , ADN Ribosómico/efectos de la radiación , Regulación de la Expresión Génica/genética , Regulación de la Expresión Génica/efectos de la radiación , Humanos , ARN Polimerasa I/efectos de la radiación , Radiación Ionizante , Ribosomas/genética , Ribosomas/efectos de la radiación , Transducción de Señal/efectos de la radiación , Transcripción Genética/genética , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/patología
7.
Breast Cancer Res ; 23(1): 30, 2021 03 04.
Artículo en Inglés | MEDLINE | ID: mdl-33663560

RESUMEN

BACKGROUND: Poly (ADP-ribose)-polymerase inhibitors (PARPi) have been approved for cancer patients with germline BRCA1/2 (gBRCA1/2) mutations, and efforts to expand the utility of PARPi beyond BRCA1/2 are ongoing. In preclinical models of triple-negative breast cancer (TNBC) with intact DNA repair, we have previously shown an induced synthetic lethality with combined EGFR inhibition and PARPi. Here, we report the safety and clinical activity of lapatinib and veliparib in patients with metastatic TNBC. METHODS: A first-in-human, pilot study of lapatinib and veliparib was conducted in metastatic TNBC (NCT02158507). The primary endpoint was safety and tolerability. Secondary endpoints were objective response rates and pharmacokinetic evaluation. Gene expression analysis of pre-treatment tumor biopsies was performed. Key eligibility included TNBC patients with measurable disease and prior anthracycline-based and taxane chemotherapy. Patients with gBRCA1/2 mutations were excluded. RESULTS: Twenty patients were enrolled, of which 17 were evaluable for response. The median number of prior therapies in the metastatic setting was 1 (range 0-2). Fifty percent of patients were Caucasian, 45% African-American, and 5% Hispanic. Of evaluable patients, 4 demonstrated a partial response and 2 had stable disease. There were no dose-limiting toxicities. Most AEs were limited to grade 1 or 2 and no drug-drug interactions noted. Exploratory gene expression analysis suggested baseline DNA repair pathway score was lower and baseline immunogenicity was higher in the responders compared to non-responders. CONCLUSIONS: Lapatinib plus veliparib therapy has a manageable safety profile and promising antitumor activity in advanced TNBC. Further investigation of dual therapy with EGFR inhibition and PARP inhibition is needed. TRIAL REGISTRATION: ClinicalTrials.gov , NCT02158507 . Registered on 12 September 2014.


Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/patología , Adulto , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Bencimidazoles/administración & dosificación , Bencimidazoles/farmacocinética , Manejo de la Enfermedad , Monitoreo de Drogas , Femenino , Humanos , Lapatinib/administración & dosificación , Lapatinib/farmacocinética , Persona de Mediana Edad , Metástasis de la Neoplasia , Estadificación de Neoplasias , Proyectos Piloto , Resultado del Tratamiento , Neoplasias de la Mama Triple Negativas/diagnóstico por imagen
8.
Am J Epidemiol ; 190(10): 1977-1992, 2021 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-33861317

RESUMEN

Genotype-phenotype association studies often combine phenotype data from multiple studies to increase statistical power. Harmonization of the data usually requires substantial effort due to heterogeneity in phenotype definitions, study design, data collection procedures, and data-set organization. Here we describe a centralized system for phenotype harmonization that includes input from phenotype domain and study experts, quality control, documentation, reproducible results, and data-sharing mechanisms. This system was developed for the National Heart, Lung, and Blood Institute's Trans-Omics for Precision Medicine (TOPMed) program, which is generating genomic and other -omics data for more than 80 studies with extensive phenotype data. To date, 63 phenotypes have been harmonized across thousands of participants (recruited in 1948-2012) from up to 17 studies per phenotype. Here we discuss challenges in this undertaking and how they were addressed. The harmonized phenotype data and associated documentation have been submitted to National Institutes of Health data repositories for controlled access by the scientific community. We also provide materials to facilitate future harmonization efforts by the community, which include 1) the software code used to generate the 63 harmonized phenotypes, enabling others to reproduce, modify, or extend these harmonizations to additional studies, and 2) the results of labeling thousands of phenotype variables with controlled vocabulary terms.


Asunto(s)
Estudios de Asociación Genética/métodos , Fenómica/métodos , Medicina de Precisión/métodos , Agregación de Datos , Humanos , Difusión de la Información , National Heart, Lung, and Blood Institute (U.S.) , Fenotipo , Evaluación de Programas y Proyectos de Salud , Estados Unidos
9.
Antimicrob Agents Chemother ; 65(10): e0093221, 2021 09 17.
Artículo en Inglés | MEDLINE | ID: mdl-34339269

RESUMEN

High attrition rates in tuberculosis (TB) drug development have been largely attributed to safety, which is likely due to the use of endpoint assays measuring cell viability to detect drug cytotoxicity. In drug development for cancer, metabolic, and neurological disorders and for antibiotics, cytotoxicity is increasingly being assessed using extracellular flux (XF) analysis, which measures cellular bioenergetic metabolism in real time. Here, we adopt the XF platform to investigate the cytotoxicity of drugs currently used in TB treatment on the bioenergetic metabolism of HepG2 cells, THP-1 macrophages, and human monocyte-derived macrophages (hMDMs). We found that the XF analysis reveals earlier drug-induced effects on the cells' bioenergetic metabolism prior to cell death, measured by conventional viability assays. Furthermore, each cell type has a distinct response to drug treatment, suggesting that more than one cell type should be considered to examine cytotoxicity in TB drug development. Interestingly, chemically unrelated drugs with different modes of action on Mycobacterium tuberculosis have similar effects on the bioenergetic parameters of the cells, thus discouraging the prediction of potential cytotoxicity based on chemical structure and mode of action of new chemical entities. The clustering of the drug-induced effects on the hMDM bioenergetic parameters are reflected in the clustering of the effects of the drugs on cytokine production in hMDMs, demonstrating concurrence between the effects of the drugs on the metabolism and functioning of the macrophages. These findings can be used as a benchmark to establish XF analysis as a new tool to assay cytotoxicity in TB drug development.


Asunto(s)
Mycobacterium tuberculosis , Tuberculosis , Antituberculosos/toxicidad , Metabolismo Energético , Humanos , Macrófagos
10.
Hum Mol Genet ; 27(16): 2940-2953, 2018 08 15.
Artículo en Inglés | MEDLINE | ID: mdl-29878111

RESUMEN

C-reactive protein (CRP) is a circulating biomarker indicative of systemic inflammation. We aimed to evaluate genetic associations with CRP levels among non-European-ancestry populations through discovery, fine-mapping and conditional analyses. A total of 30 503 non-European-ancestry participants from 6 studies participating in the Population Architecture using Genomics and Epidemiology study had serum high-sensitivity CRP measurements and ∼200 000 single nucleotide polymorphisms (SNPs) genotyped on the Metabochip. We evaluated the association between each SNP and log-transformed CRP levels using multivariate linear regression, with additive genetic models adjusted for age, sex, the first four principal components of genetic ancestry, and study-specific factors. Differential linkage disequilibrium patterns between race/ethnicity groups were used to fine-map regions associated with CRP levels. Conditional analyses evaluated for multiple independent signals within genetic regions. One hundred and sixty-three unique variants in 12 loci in overall or race/ethnicity-stratified Metabochip-wide scans reached a Bonferroni-corrected P-value <2.5E-7. Three loci have no (HACL1, OLFML2B) or only limited (PLA2G6) previous associations with CRP levels. Six loci had different top hits in race/ethnicity-specific versus overall analyses. Fine-mapping refined the signal in six loci, particularly in HNF1A. Conditional analyses provided evidence for secondary signals in LEPR, IL1RN and HNF1A, and for multiple independent signals in CRP and APOE. We identified novel variants and loci associated with CRP levels, generalized known CRP associations to a multiethnic study population, refined association signals at several loci and found evidence for multiple independent signals at several well-known loci. This study demonstrates the benefit of conducting inclusive genetic association studies in large multiethnic populations.


Asunto(s)
Proteína C-Reactiva/genética , Estudio de Asociación del Genoma Completo , Metagenómica , Epidemiología Molecular/métodos , Liasas de Carbono-Carbono , Enoil-CoA Hidratasa/genética , Femenino , Glicoproteínas/genética , Fosfolipasas A2 Grupo VI/genética , Humanos , Desequilibrio de Ligamiento , Masculino , Polimorfismo de Nucleótido Simple , Población Blanca/genética
11.
BMC Infect Dis ; 20(1): 111, 2020 Feb 10.
Artículo en Inglés | MEDLINE | ID: mdl-32039707

RESUMEN

After publication of the original article [1], we were notified that Fig. 3 has "Fig. 1" posted on the top of it and Figs. 4 and 5 have "Genomic Position" in a different spot.

12.
Proc Natl Acad Sci U S A ; 114(38): E7949-E7958, 2017 09 19.
Artículo en Inglés | MEDLINE | ID: mdl-28855339

RESUMEN

Posttranslational histone modifications play important roles in regulating chromatin-based nuclear processes. Histone H2AK119 ubiquitination (H2Aub) is a prevalent modification and has been primarily linked to gene silencing. However, the underlying mechanism remains largely obscure. Here we report the identification of RSF1 (remodeling and spacing factor 1), a subunit of the RSF complex, as a H2Aub binding protein, which mediates the gene-silencing function of this histone modification. RSF1 associates specifically with H2Aub, but not H2Bub nucleosomes, through a previously uncharacterized and obligatory region designated as ubiquitinated H2A binding domain. In human and mouse cells, genes regulated by RSF1 overlap significantly with those controlled by RNF2/Ring1B, the subunit of Polycomb repressive complex 1 (PRC1) which catalyzes the ubiquitination of H2AK119. About 82% of H2Aub-enriched genes, including the classic PRC1 target Hox genes, are bound by RSF1 around their transcription start sites. Depletion of H2Aub levels by Ring1B knockout results in a significant reduction of RSF1 binding. In contrast, RSF1 knockout does not affect RNF2/Ring1B or H2Aub levels but leads to derepression of H2Aub target genes, accompanied by changes in H2Aub chromatin organization and release of linker histone H1. The action of RSF1 in H2Aub-mediated gene silencing is further demonstrated by chromatin-based in vitro transcription. Finally, RSF1 and Ring1 act cooperatively to regulate mesodermal cell specification and gastrulation during Xenopus early embryonic development. Taken together, these data identify RSF1 as a H2Aub reader that contributes to H2Aub-mediated gene silencing by maintaining a stable nucleosome pattern at promoter regions.


Asunto(s)
Silenciador del Gen/fisiología , Histonas/metabolismo , Proteínas Nucleares/metabolismo , Nucleosomas/metabolismo , Transactivadores/metabolismo , Ubiquitinación/fisiología , Animales , Células HeLa , Histonas/genética , Humanos , Ratones , Proteínas Nucleares/genética , Nucleosomas/genética , Complejo Represivo Polycomb 1/genética , Complejo Represivo Polycomb 1/metabolismo , Regiones Promotoras Genéticas/fisiología , Transactivadores/genética , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo
13.
BMC Infect Dis ; 19(1): 1046, 2019 Dec 10.
Artículo en Inglés | MEDLINE | ID: mdl-31822287

RESUMEN

BACKGROUND: Congenital cytomegalovirus (cCMV) infection is the most common congenital infection and a leading cause of long-term neurological and sensory sequelae, the most common being sensorineural hearing loss (SNHL). Despite extensive research, clinical or laboratory markers to identify CMV infected children with increased risk for disease have not been identified. This study utilizes viral whole-genome next generation-sequencing (NGS) of specimens from congenitally infected infants to explore viral diversity and specific viral variants that may be associated with symptomatic infection and SNHL. METHODS: CMV DNA from urine specimens of 30 infants (17 asymptomatic, 13 symptomatic) was target enriched and next generation sequenced resulting in 93% coverage of the CMV genome allowing analysis of viral diversity. RESULTS: Variant frequency distribution was compared between children with symptomatic and asymptomatic cCMV and those with (n = 13) and without (n = 17) hearing loss. The CMV genes UL48A, UL88, US19 and US22 were found to have an increase in nucleotide diversity in symptomatic children; while UL57, UL20, UL104, US14, UL115, and UL35 had an increase in diversity in children with hearing loss. An analysis of single variant differences between symptomatic and asymptomatic children found UL55 to have the highest number, while the most variants associated with SNHL were in the RL11 gene family. In asymptomatic infants with SNHL, mutations were observed more frequently in UL33 and UL20. CONCLUSION: CMV genomes from infected newborns can be mapped to 93% of the genome at a depth allowing accurate and reproducible analysis of polymorphisms for variant and gene discovery that may be linked to symptomatic and hearing loss outcomes.


Asunto(s)
Infecciones por Citomegalovirus/diagnóstico , Citomegalovirus/genética , ADN Viral/genética , Pérdida Auditiva Sensorineural/diagnóstico , Niño , Citomegalovirus/clasificación , Citomegalovirus/aislamiento & purificación , Infecciones por Citomegalovirus/complicaciones , Infecciones por Citomegalovirus/virología , ADN Viral/metabolismo , ADN Viral/orina , Femenino , Pérdida Auditiva Sensorineural/complicaciones , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Lactante , Recién Nacido , Masculino , Mutación , Filogenia , Análisis de Componente Principal
14.
Proc Natl Acad Sci U S A ; 113(1): E51-60, 2016 Jan 05.
Artículo en Inglés | MEDLINE | ID: mdl-26699484

RESUMEN

Epigenetic mechanisms play important regulatory roles in hematopoiesis and hematopoietic stem cell (HSC) function. Subunits of polycomb repressive complex 1 (PRC1), the major histone H2A ubiquitin ligase, are critical for both normal and pathological hematopoiesis; however, it is unclear which of the several counteracting H2A deubiquitinases functions along with PRC1 to control H2A ubiquitination (ubH2A) level and regulates hematopoiesis in vivo. Here we investigated the function of Usp16 in mouse hematopoiesis. Conditional deletion of Usp16 in bone marrow resulted in a significant increase of global ubH2A level and lethality. Usp16 deletion did not change HSC number but was associated with a dramatic reduction of mature and progenitor cell populations, revealing a role in governing HSC lineage commitment. ChIP- and RNA-sequencing studies in HSC and progenitor cells revealed that Usp16 bound to many important hematopoietic regulators and that Usp16 deletion altered the expression of genes in transcription/chromosome organization, immune response, hematopoietic/lymphoid organ development, and myeloid/leukocyte differentiation. The altered gene expression was partly rescued by knockdown of PRC1 subunits, suggesting that Usp16 and PRC1 counterbalance each other to regulate cellular ubH2A level and gene expression in the hematopoietic system. We further discovered that knocking down Cdkn1a (p21cip1), a Usp16 target and regulated gene, rescued the altered cell cycle profile and differentiation defect of Usp16-deleted HSCs. Collectively, these studies identified Usp16 as one of the histone H2A deubiquitinases, which coordinates with the H2A ubiquitin ligase PRC1 to regulate hematopoiesis, and revealed cell cycle regulation by Usp16 as key for HSC differentiation.


Asunto(s)
Hematopoyesis/fisiología , Células Madre Hematopoyéticas/citología , Ubiquitina Tiolesterasa/fisiología , Proteasas Ubiquitina-Específicas/fisiología , Animales , Recuento de Células , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Endopeptidasas/genética , Endopeptidasas/fisiología , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Genes Letales , Hematopoyesis/genética , Histonas/metabolismo , Ratones , Ratones Endogámicos C57BL , Proteínas del Grupo Polycomb/genética , Proteínas del Grupo Polycomb/fisiología , Transactivadores , Ubiquitina Tiolesterasa/genética , Proteasas Ubiquitina-Específicas/genética
15.
Stem Cells ; 35(2): 473-484, 2017 02.
Artículo en Inglés | MEDLINE | ID: mdl-27539014

RESUMEN

Long-term self-renewing hematopoietic stem cell (LT-HSC) homeostasis within the bone marrow (BM) of adult mammals is regulated by complex interactions between LT-HSC and a number of niche-associated cell types including mesenchymal stromal/stem cells (MSC), osteoblasts (OB), macrophage, and neuronal cells in close proximity with the vasculature. Here, we cloned and functionally characterized a murine BM MSC subpopulation that was uniformly Nestin+ Lepr + Sca-1+ CD146+ and could be stably propagated with high colony-forming unit fibroblast re-cloning efficiency. MSC synergized with SCF and IL-11 to support a 20-fold expansion in true LT-HSC after 10-days of in vitro coculture. Optimal stimulation of LT-HSC expansion was minimally dependent on Notch signaling but was significantly enhanced by global inhibition of Wnt signaling. The self-renewal-promoting activity of MSC was progressively lost when MSC clones were differentiated into mature OB. This suggests that the stage of osteoblast development may significantly impact the ability of osteolineage cells to support LT-HSC homeostasis in vivo. Stem Cells 2017;35:473-484.


Asunto(s)
Autorrenovación de las Células , Células Madre Hematopoyéticas/citología , Células Madre Mesenquimatosas/citología , Osteogénesis , Animales , Células de la Médula Ósea/citología , Diferenciación Celular , Células Cultivadas , Células Clonales , Técnicas de Cocultivo , Células Madre Hematopoyéticas/metabolismo , Ratones Endogámicos C57BL , Osificación Heterotópica/patología , Receptores Notch/metabolismo , Transducción de Señal
16.
Immunity ; 30(1): 92-107, 2009 Jan 16.
Artículo en Inglés | MEDLINE | ID: mdl-19119024

RESUMEN

Development of T helper (Th) 17 cells requires transforming growth factor (TGF)-beta and interleukin (IL)-6 and is independent of the Th1 pathway. Although T cells that produce interferon (IFN)-gamma are a recognized feature of Th17 cell responses, mice deficient for STAT4 and T-bet-two prototypical Th1 transcription factors-are protected from autoimmunity associated with Th17 pathogenesis. To examine the fate and pathogenic potential of Th17 cells and origin of IFN-gamma-producing T cells that emerge during Th17 immunity, we developed IL-17F reporter mice that identify cells committed to expression of IL-17F and IL-17A. Th17 cells required TGF-beta for sustained expression of IL-17F and IL-17A. In the absence of TGF-beta, both IL-23 and IL-12 acted to suppress IL-17 and enhance IFN-gamma production in a STAT4- and T-bet-dependent manner, albeit with distinct efficiencies. These results support a model of late Th17 developmental plasticity with implications for autoimmunity and host defense.


Asunto(s)
Linaje de la Célula/inmunología , Interleucina-17/metabolismo , Subgrupos de Linfocitos T/inmunología , Células TH1/inmunología , Células Th2/inmunología , Factor de Crecimiento Transformador beta/metabolismo , Animales , Células Cultivadas , Citometría de Flujo , Interferón gamma/metabolismo , Interleucina-23/metabolismo , Ratones , Ratones Noqueados
17.
J Cell Biochem ; 118(9): 2968-2976, 2017 09.
Artículo en Inglés | MEDLINE | ID: mdl-28247948

RESUMEN

The low-density lipoprotein receptor-related protein 6 (LRP6) is an essential Wnt co-receptor of the Wnt/ß-catenin signaling pathway. Although studies have shown an increased expression of LRP6 in several types of cancer, its function in tumor development and progression remains to be elucidated. We herein demonstrated that LRP6 expression is up-regulated in human triple negative breast cancer (TNBC) patients and human TNBC cell lines, and that knockdown of LRP6 expression and treatment of recombinant Mesd protein (a specific inhibitor of LRP6) significantly decreased cell migration and invasion of TNBC MDA-MB-231 and BT549 cells. Interestingly, the effects of LRP6 knockdown and Mesd treatment on TNBC cell migration and invasion were more prominent than on TNBC cell proliferation/viability. Mechanistically, LRP6 knockdown and Mesd treatment inhibited Wnt/ß-catenin signaling and decreased the expression of S100A4, a mediator of cancer metastasis and a specific target of Wnt/ß-catenin signaling, in TNBC cells. Together, our data suggest that LRP6 promotes TNBC cell migration and invasion by regulating the expression and function of S100A4 via the Wnt/ß-catenin signaling pathway. J. Cell. Biochem. 118: 2968-2976, 2017. © 2017 Wiley Periodicals, Inc.


Asunto(s)
Neoplasias de la Mama/metabolismo , Movimiento Celular , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/metabolismo , Proteínas de Neoplasias/metabolismo , Vía de Señalización Wnt , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Femenino , Técnicas de Silenciamiento del Gen , Humanos , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/genética , Células MCF-7 , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Invasividad Neoplásica , Proteínas de Neoplasias/genética
18.
Breast Cancer Res ; 19(1): 73, 2017 Jun 21.
Artículo en Inglés | MEDLINE | ID: mdl-28637482

RESUMEN

BACKGROUND: Members of the microRNA (miR)-200 family, which are involved in tumor metastasis, have potential as cancer biomarkers, but their regulatory mechanisms remain elusive. METHODS: We investigated FOXP3-inducible breast cancer cells, Foxp3 heterozygous Scurfy mutant (Foxp3 sf/+ ) female mice, and patients with breast cancer for characterization of the formation and regulation of the miR-200 family in breast cancer cells and circulation. Participants (259), including patients with breast cancer or benign breast tumors, members of breast cancer families, and healthy controls, were assessed for tumor and circulating levels of the miR-200 family. RESULTS: First, we identified a FOXP3-KAT2B-miR-200c/141 axis in breast cancer cells. Second, aging Foxp3 sf/+ female mice developed spontaneous breast cancers and lung metastases. Levels of miR-200c and miR-141 were lower in Foxp3 sf/+ tumor cells than in normal breast epithelial cells, but plasma levels of miR-200c and miR-141 in the Foxp3 sf/+ mice increased during tumor progression and metastasis. Third, in patients with breast cancer, the levels of miR-200c and 141 were lower in FOXP3 low relative to those with FOXP3 high breast cancer cells, especially in late-stage and metastatic cancer cells. The levels of miR-200c and miR-141 were higher in plasma from patients with metastatic breast cancer than in plasma from those with localized breast cancer, with benign breast tumors, with a family history of breast cancer, or from healthy controls. Finally, in Foxp3 sf/+ mice, plasma miR-200c and miR-141 appeared to be released from tumor cells. CONCLUSIONS: miR-200c and miR-141 are regulated by a FOXP3-KAT2B axis in breast cancer cells, and circulating levels of miR-200c and miR-141 are potential biomarkers for early detection of breast cancer metastases.


Asunto(s)
Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Factores de Transcripción Forkhead/genética , Regulación Neoplásica de la Expresión Génica , MicroARNs/genética , Factores de Transcripción p300-CBP/genética , Adulto , Anciano , Animales , Biomarcadores de Tumor , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , MicroARN Circulante , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Femenino , Humanos , Ratones , Persona de Mediana Edad , Clasificación del Tumor , Metástasis de la Neoplasia , Estadificación de Neoplasias , Curva ROC , Transcripción Genética
19.
Proteomics ; 16(3): 516-31, 2016 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-26572242

RESUMEN

We have applied a serologic proteomic workflow involving three complementary MS approaches to a tissue-specific Kras(G12D) -knockin mouse model of pancreatic cancer that consistently forms precancerous lesions by 4 months of age. The three proteomics applications were highly complementary and allowed us to survey the entire range of low to high molecular weight serologic proteins. Combined, we identified 121 (49↓, 72↑) unique and statistically relevant serologic biomarkers with 88% previously reported to be associated with cancer and 38% specifically correlated with pancreatic cancer. Four markers, lysozyme C2, cytokeratin 19, Serpina1A and Pcf11, were further verified by Western blotting. When applying systems analysis, the top-associated gene ontology functions were tied to wound healing, RXR signaling, growth, differentiation and innate immune activation through the JAK/STAT pathway. Upon further investigation of the apparent immune response using a multiplex cytokine screen, we found that IFN-γ, VEGF and GM-CSF were significantly increased in serum from the Kras(G12D) animals compared to littermate controls. By combining three complementary MS applications, we were able to survey the native intact peptidome and the global proteome in parallel, unveiling pathways that may be biologically relevant to promotion of pancreatic cancer progression and serologic markers of noninvasive early-stage neoplasia.


Asunto(s)
Biomarcadores de Tumor/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias Pancreáticas/genética , Proteoma/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Animales , Biomarcadores de Tumor/sangre , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Técnicas de Sustitución del Gen , Factor Estimulante de Colonias de Granulocitos y Macrófagos/sangre , Factor Estimulante de Colonias de Granulocitos y Macrófagos/genética , Interferón gamma/sangre , Interferón gamma/genética , Queratina-19/sangre , Queratina-19/genética , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Muramidasa/sangre , Muramidasa/genética , Neoplasias Pancreáticas/sangre , Neoplasias Pancreáticas/patología , Proteoma/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/sangre , Transducción de Señal , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Factor A de Crecimiento Endotelial Vascular/sangre , Factor A de Crecimiento Endotelial Vascular/genética , alfa 1-Antitripsina/sangre , alfa 1-Antitripsina/genética , Factores de Escisión y Poliadenilación de ARNm/sangre , Factores de Escisión y Poliadenilación de ARNm/genética
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