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1.
Proc Natl Acad Sci U S A ; 114(19): E3816-E3822, 2017 05 09.
Artículo en Inglés | MEDLINE | ID: mdl-28439009

RESUMEN

As estrogen receptor ß-/- (ERß-/-) mice age, the ventral prostate (VP) develops increased numbers of hyperplastic, fibroplastic lesions and inflammatory cells. To identify genes involved in these changes, we used RNA sequencing and immunohistochemistry to compare gene expression profiles in the VP of young (2-mo-old) and aging (18-mo-old) ERß-/- mice and their WT littermates. We also treated young and old WT mice with an ERß-selective agonist and evaluated protein expression. The most significant findings were that ERß down-regulates androgen receptor (AR) signaling and up-regulates the tumor suppressor phosphatase and tensin homolog (PTEN). ERß agonist increased expression of the AR corepressor dachshund family (DACH1/2), T-cadherin, stromal caveolin-1, and nuclear PTEN and decreased expression of RAR-related orphan receptor c, Bcl2, inducible nitric oxide synthase, and IL-6. In the ERß-/- mouse VP, RNA sequencing revealed that the following genes were up-regulated more than fivefold: Bcl2, clusterin, the cytokines CXCL16 and -17, and a marker of basal/intermediate cells (prostate stem cell antigen) and cytokeratins 4, 5, and 17. The most down-regulated genes were the following: the antioxidant gene glutathione peroxidase 3; protease inhibitors WAP four-disulfide core domain 3 (WFDC3); the tumor-suppressive genes T-cadherin and caveolin-1; the regulator of transforming growth factor ß signaling SMAD7; and the PTEN ubiquitin ligase NEDD4. The role of ERß in opposing AR signaling, proliferation, and inflammation suggests that ERß-selective agonists may be used to prevent progression of prostate cancer, prevent fibrosis and development of benign prostatic hyperplasia, and treat prostatitis.


Asunto(s)
Envejecimiento/metabolismo , Regulación hacia Abajo , Receptor beta de Estrógeno/metabolismo , Próstata/metabolismo , Receptores Androgénicos/biosíntesis , Transducción de Señal , Envejecimiento/genética , Envejecimiento/patología , Andrógenos/metabolismo , Animales , Quimiocina CXCL16/biosíntesis , Quimiocina CXCL16/genética , Quimiocinas CXC/biosíntesis , Quimiocinas CXC/genética , Clusterina/biosíntesis , Clusterina/genética , Receptor beta de Estrógeno/genética , Interleucina-6/genética , Interleucina-6/metabolismo , Queratinas/biosíntesis , Queratinas/genética , Masculino , Ratones , Ratones Noqueados , Ubiquitina-Proteína Ligasas Nedd4/biosíntesis , Ubiquitina-Proteína Ligasas Nedd4/genética , Fosfohidrolasa PTEN/biosíntesis , Fosfohidrolasa PTEN/genética , Próstata/patología , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Receptores Androgénicos/genética , Proteína smad7/biosíntesis , Proteína smad7/genética
2.
Proc Natl Acad Sci U S A ; 113(27): 7614-9, 2016 07 05.
Artículo en Inglés | MEDLINE | ID: mdl-27335465

RESUMEN

The etiology of peripheral squamous cell lung cancer (PSCCa) remains unknown. Here, we show that this condition spontaneously develops in mice in which the genes for two oxysterol receptors, Liver X Receptor (LXR) α (Nr1h3) and ß (Nr1h2), are inactivated. By 1 y of age, most of these mice have to be euthanized because of severe dyspnea. Starting at 3 mo, the lungs of LXRα,ß(Dko) mice, but not of LXRα or LXRß single knockout mice, progressively accumulate foam cells, so that by 1 y, the lungs are covered by a "golden coat." There is infiltration of inflammatory cells and progressive accumulation of lipid in the alveolar wall, type 2 pneumocytes, and macrophages. By 14 mo, there are three histological lesions: one resembling adenomatous hyperplasia, one squamous metaplasia, and one squamous cell carcinoma characterized by expression of transformation-related protein (p63), sex determining region Y-box 2 (Sox2), cytokeratin 14 (CK14), and cytokeratin 13 (CK13) and absence of thyroid transcription factor 1 (TTF1), and prosurfactant protein C (pro-SPC). RNA sequencing analysis at 12 mo confirmed a massive increase in markers of M1 macrophages and lymphocytes. The data suggest a previously unidentified etiology of PSCCa: cholesterol dysregulation and M1 macrophage-predominant lung inflammation combined with damage to, and aberrant repair of, lung tissue, particularly the peripheral parenchyma. The results raise the possibility that components of the LXR signaling may be useful targets in the treatment of PSCCa.


Asunto(s)
Metabolismo de los Lípidos , Receptores X del Hígado/fisiología , Neoplasias Pulmonares/etiología , Pulmón/metabolismo , Neoplasias de Células Escamosas/etiología , Células Epiteliales Alveolares/metabolismo , Animales , Fibroblastos/metabolismo , Homeostasis , Pulmón/patología , Macrófagos/metabolismo , Ratones Endogámicos C57BL , Neumonía/etiología , Análisis de Secuencia de ARN
3.
Proc Natl Acad Sci U S A ; 112(45): 14006-11, 2015 Nov 10.
Artículo en Inglés | MEDLINE | ID: mdl-26504234

RESUMEN

The recent discovery of browning of white adipose tissue (WAT) has raised great research interest because of its significant potential in counteracting obesity and type 2 diabetes. Browning is the result of the induction in WAT of a newly discovered type of adipocyte, the beige cell. When mice are exposed to cold or several kinds of hormones or treatments with chemicals, specific depots of WAT undergo a browning process, characterized by highly activated mitochondria and increased heat production and energy expenditure. However, the mechanisms underlying browning are still poorly understood. Liver X receptors (LXRs) are one class of nuclear receptors, which play a vital role in regulating cholesterol, triglyceride, and glucose metabolism. Following our previous finding that LXRs serve as repressors of uncoupling protein-1 (UCP1) in classic brown adipose tissue in female mice, we found that LXRs, especially LXRß, also repress the browning process of subcutaneous adipose tissue (SAT) in male rodents fed a normal diet. Depletion of LXRs activated thyroid-stimulating hormone (TSH)-releasing hormone (TRH)-positive neurons in the paraventricular nucleus area of the hypothalamus and thus stimulated secretion of TSH from the pituitary. Consequently, production of thyroid hormones in the thyroid gland and circulating thyroid hormone level were increased. Moreover, the activity of thyroid signaling in SAT was markedly increased. Together, our findings have uncovered the basis of increased energy expenditure in male LXR knockout mice and provided support for targeting LXRs in treatment of obesity.


Asunto(s)
Tejido Adiposo Blanco/metabolismo , Encéfalo/metabolismo , Metabolismo Energético/fisiología , Mitocondrias/metabolismo , Receptores Nucleares Huérfanos/metabolismo , Hormonas Tiroideas/metabolismo , Análisis de Varianza , Animales , Composición Corporal/fisiología , Ensayo de Inmunoadsorción Enzimática , Perfilación de la Expresión Génica , Inmunohistoquímica , Receptores X del Hígado , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores Nucleares Huérfanos/genética , Hormona Liberadora de Tirotropina/metabolismo
4.
Proc Natl Acad Sci U S A ; 112(16): 5135-40, 2015 Apr 21.
Artículo en Inglés | MEDLINE | ID: mdl-25848008

RESUMEN

In 1998, an estrogen receptor ß (ERß) knockout (KO) mouse was created by interrupting the gene at the DNA binding domain (DBD) with a neocassette. The mutant females were subfertile and there were abnormalities in the brain, prostate, lung, colon, and immune system. In 2008, another ERß mutant mouse was generated by deleting ERß exon 3 which encodes the first zinc finger in the DBD. The female mice of this strain were unable to ovulate but were otherwise normal. The differences in the phenotypes of the two KO strains, have led to questions about the physiological function of ERß. In the present study, we created an ERß exon 3-deleted mouse (ERß-Δex3) and confirmed that the only observable defect was anovulation. Despite the two in-frame stop codons introduced by splicing between exons 2 and 4, an ERß protein was expressed in nuclei of prostate epithelial cells. Using two different anti-ERß antibodies, we showed that an in-frame ligand binding domain and C terminus were present in the ERß-Δex3 protein. Moreover, with nuclear extracts from ERß-Δex3 prostates, there was an ERß-dependent retardation of migration of activator protein-1 response elements in EMSA. Unlike the original knockout mouse, expression of Ki67, androgen receptor, and Dachshund-1 in prostate epithelium was not altered in the ERß-Δex3 mouse. We conclude that very little of ERß transcriptional activity depends on binding to classical estrogen response elements (EREs).


Asunto(s)
Receptor beta de Estrógeno/genética , Exones/genética , Elementos de Respuesta/genética , Eliminación de Secuencia/genética , Transducción de Señal/genética , Animales , ADN/metabolismo , Femenino , Regulación de la Expresión Génica , Marcación de Gen , Masculino , Ratones Mutantes , Ovario/fisiopatología , Próstata/metabolismo , Próstata/patología , Unión Proteica/genética , Transporte de Proteínas , Receptores Androgénicos/metabolismo , Reproducibilidad de los Resultados , Factor de Transcripción AP-1/metabolismo
5.
J Clin Invest ; 134(5)2024 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-38426503

RESUMEN

Tissue-intrinsic mechanisms that regulate severity of systemic pathogenic immune-mediated diseases, such as acute graft-versus-host disease (GVHD), remain poorly understood. Following allogeneic hematopoietic stem cell transplantation, autophagy, a cellular stress protective response, is induced in host nonhematopoietic cells. To systematically address the role of autophagy in various host nonhematopoietic tissues, both specific classical target organs of acute GVHD (intestines, liver, and skin) and organs conventionally not known to be targets of GVHD (kidneys and heart), we generated mice with organ-specific knockout of autophagy related 5 (ATG5) to specifically and exclusively inhibit autophagy in the specific organs. When compared with wild-type recipients, animals that lacked ATG5 in the gastrointestinal tract or liver showed significantly greater tissue injury and mortality, while autophagy deficiency in the skin, kidneys, or heart did not affect mortality. Treatment with the systemic autophagy inducer sirolimus only partially mitigated GVHD mortality in intestine-specific autophagy-deficient hosts. Deficiency of autophagy increased MHC class I on the target intestinal epithelial cells, resulting in greater susceptibility to damage by alloreactive T cells. Thus, autophagy is a critical cell-intrinsic protective response that promotes tissue tolerance and regulates GVHD severity.


Asunto(s)
Enfermedad Injerto contra Huésped , Trasplante de Células Madre Hematopoyéticas , Animales , Ratones , Enfermedad Injerto contra Huésped/genética , Enfermedad Injerto contra Huésped/patología , Intestinos/patología , Linfocitos T/patología , Células Epiteliales/patología
6.
Cell Stem Cell ; 31(10): 1447-1464.e6, 2024 Oct 03.
Artículo en Inglés | MEDLINE | ID: mdl-39232559

RESUMEN

It remains unknown whether and how intestinal stem cells (ISCs) adapt to inflammatory exposure and whether the adaptation leaves scars that will affect their subsequent regeneration. We investigated the consequences of inflammation on Lgr5+ ISCs in well-defined clinically relevant models of acute gastrointestinal graft-versus-host disease (GI GVHD). Utilizing single-cell transcriptomics, as well as organoid, metabolic, epigenomic, and in vivo models, we found that Lgr5+ ISCs undergo metabolic changes that lead to the accumulation of succinate, which reprograms their epigenome. These changes reduced the ability of ISCs to differentiate and regenerate ex vivo in serial organoid cultures and also in vivo following serial transplantation. Furthermore, ISCs demonstrated a reduced capacity for in vivo regeneration despite resolution of the initial inflammatory exposure, demonstrating the persistence of the maladaptive impact induced by the inflammatory encounter. Thus, inflammation imprints the epigenome of ISCs in a manner that persists and affects their sensitivity to adapt to future stress or challenges.


Asunto(s)
Epigénesis Genética , Inflamación , Intestinos , Células Madre , Animales , Inflamación/patología , Inflamación/genética , Células Madre/metabolismo , Células Madre/citología , Ratones , Intestinos/citología , Impresión Genómica , Ratones Endogámicos C57BL , Enfermedad Injerto contra Huésped , Regeneración , Diferenciación Celular , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Organoides/metabolismo
7.
Nat Cell Biol ; 26(4): 593-603, 2024 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-38553595

RESUMEN

Loss of protein function is a driving force of ageing. We have identified peptidyl-prolyl isomerase A (PPIA or cyclophilin A) as a dominant chaperone in haematopoietic stem and progenitor cells. Depletion of PPIA accelerates stem cell ageing. We found that proteins with intrinsically disordered regions (IDRs) are frequent PPIA substrates. IDRs facilitate interactions with other proteins or nucleic acids and can trigger liquid-liquid phase separation. Over 20% of PPIA substrates are involved in the formation of supramolecular membrane-less organelles. PPIA affects regulators of stress granules (PABPC1), P-bodies (DDX6) and nucleoli (NPM1) to promote phase separation and increase cellular stress resistance. Haematopoietic stem cell ageing is associated with a post-transcriptional decrease in PPIA expression and reduced translation of IDR-rich proteins. Here we link the chaperone PPIA to the synthesis of intrinsically disordered proteins, which indicates that impaired protein interaction networks and macromolecular condensation may be potential determinants of haematopoietic stem cell ageing.


Asunto(s)
Proteínas Intrínsecamente Desordenadas , Proteínas Intrínsecamente Desordenadas/química , Ciclofilina A/genética , Ciclofilina A/metabolismo , Proteínas de Unión al ARN , Células Madre Hematopoyéticas/metabolismo
8.
Res Sq ; 2023 Mar 07.
Artículo en Inglés | MEDLINE | ID: mdl-36945465

RESUMEN

Intestinal stem cells (ISC) encounter inflammatory insults in immune mediated gastro-intestinal (GI) diseases. It remains unknown whether, and how, they adapt, and if the adaptation leaves scars on the ISCs that affects their subsequent regeneration capacity. We investigated the consequences of inflammation on Lgr5+ISCs in well-defined clinically relevant models of gastro-intestinal acute graft-versus-host disease (GI GVHD). Utilizing single cell transcriptomics, organoid, metabolic, epigenomic and in vivo models we found that Lgr5+ISCs undergo metabolic changes that lead to accumulation of succinate, which reprograms its epigenome. These changes reduced the ability of ISCs to differentiate and regenerate ex vivo in serial organoid cultures demonstrating the persistence of the maladaptive impact of an in vivo inflammatory encounter by the ISCs. Thus, inflammation from GI GVHD leaves a memory of its effects on ISCs that persist and are likely to affect their sensitivity to adapt to future stress or challenges.

9.
iScience ; 26(9): 107596, 2023 Sep 15.
Artículo en Inglés | MEDLINE | ID: mdl-37664586

RESUMEN

Recent studies suggest that infection reprograms hematopoietic stem and progenitor cells (HSPCs) to enhance innate immune responses upon secondary infectious challenge, a process called "trained immunity." However, the specificity and cell types responsible for this response remain poorly defined. We established a model of trained immunity in mice in response to Mycobacterium avium infection. scRNA-seq analysis revealed that HSPCs activate interferon gamma-response genes heterogeneously upon primary challenge, while rare cell populations expand. Macrophages derived from trained HSPCs demonstrated enhanced bacterial killing and metabolism, and a single dose of recombinant interferon gamma exposure was sufficient to induce similar training. Mice transplanted with influenza-trained HSPCs displayed enhanced immunity against M. avium challenge and vice versa, demonstrating cross protection against antigenically distinct pathogens. Together, these results indicate that heterogeneous responses to infection by HSPCs can lead to long-term production of bone marrow derived macrophages with enhanced function and confer cross-protection against alternative pathogens.

10.
Science ; 381(6662): eabn4180, 2023 09 08.
Artículo en Inglés | MEDLINE | ID: mdl-37676964

RESUMEN

Despite substantial advances in targeting mutant KRAS, tumor resistance to KRAS inhibitors (KRASi) remains a major barrier to progress. Here, we report proteostasis reprogramming as a key convergence point of multiple KRASi-resistance mechanisms. Inactivation of oncogenic KRAS down-regulated both the heat shock response and the inositol-requiring enzyme 1α (IRE1α) branch of the unfolded protein response, causing severe proteostasis disturbances. However, IRE1α was selectively reactivated in an ER stress-independent manner in acquired KRASi-resistant tumors, restoring proteostasis. Oncogenic KRAS promoted IRE1α protein stability through extracellular signal-regulated kinase (ERK)-dependent phosphorylation of IRE1α, leading to IRE1α disassociation from 3-hydroxy-3-methylglutaryl reductase degradation (HRD1) E3-ligase. In KRASi-resistant tumors, both reactivated ERK and hyperactivated AKT restored IRE1α phosphorylation and stability. Suppression of IRE1α overcame resistance to KRASi. This study reveals a druggable mechanism that leads to proteostasis reprogramming and facilitates KRASi resistance.


Asunto(s)
Antineoplásicos , Resistencia a Antineoplásicos , Endorribonucleasas , Inhibidores Enzimáticos , Quinasas MAP Reguladas por Señal Extracelular , Factores de Transcripción del Choque Térmico , Neoplasias , Proteostasis , Proteínas Proto-Oncogénicas p21(ras) , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Proteínas Serina-Treonina Quinasas , Proteínas Proto-Oncogénicas p21(ras)/antagonistas & inhibidores , Proteínas Proto-Oncogénicas p21(ras)/genética , Inhibidores Enzimáticos/farmacología , Antineoplásicos/farmacología , Factores de Transcripción del Choque Térmico/metabolismo
11.
Cancers (Basel) ; 14(3)2022 Feb 04.
Artículo en Inglés | MEDLINE | ID: mdl-35159073

RESUMEN

The ubiquitin-proteasome pathway precisely controls the turnover of transcription factors in the nucleus, playing an important role in maintaining appropriate quantities of these regulatory proteins. The transcription factor c-MYC is essential for normal development and is a critical cancer driver. Despite being highly expressed in several tissues and malignancies, the c-MYC protein is also continuously targeted by the ubiquitin-proteasome pathway, which can either facilitate or inhibit c-MYC degradation. Deubiquitinating proteases can remove ubiquitin chains from target proteins and rescue them from proteasomal digestion. This study sought to determine novel elements of the ubiquitin-proteasome pathway that regulate c-MYC levels. We performed an overexpression screen with 41 human proteases to identify which deubiquitinases stabilize c-MYC. We discovered that the highly expressed Otubain-1 (OTUB1) protease increases c-MYC protein levels. Confirming its role in enhancing c-MYC activity, we found that elevated OTUB1 correlates with inferior clinical outcomes in the c-MYC-dependent cancer multiple myeloma, and overexpression of OTUB1 accelerates the growth of myeloma cells. In summary, our study identifies OTUB1 as a novel amplifier of the proto-oncogene c-MYC.

12.
Cancer Res Commun ; 2(12): 1693-1710, 2022 12.
Artículo en Inglés | MEDLINE | ID: mdl-36846090

RESUMEN

Proteasome inhibitors have become the standard of care for multiple myeloma (MM). Blocking protein degradation particularly perturbs the homeostasis of short-lived polypeptides such as transcription factors and epigenetic regulators. To determine how proteasome inhibitors directly impact gene regulation, we performed an integrative genomics study in MM cells. We discovered that proteasome inhibitors reduce the turnover of DNA-associated proteins and repress genes necessary for proliferation through epigenetic silencing. Specifically, proteasome inhibition results in the localized accumulation of histone deacetylase 3 (HDAC3) at defined genomic sites, which reduces H3K27 acetylation and increases chromatin condensation. The loss of active chromatin at super-enhancers critical for MM, including the super-enhancer controlling the proto-oncogene c-MYC, reduces metabolic activity and cancer cell growth. Epigenetic silencing is attenuated by HDAC3 depletion, suggesting a tumor-suppressive element of this deacetylase in the context of proteasome inhibition. In the absence of treatment, HDAC3 is continuously removed from DNA by the ubiquitin ligase SIAH2. Overexpression of SIAH2 increases H3K27 acetylation at c-MYC-controlled genes, increases metabolic output, and accelerates cancer cell proliferation. Our studies indicate a novel therapeutic function of proteasome inhibitors in MM by reshaping the epigenetic landscape in an HDAC3-dependent manner. As a result, blocking the proteasome effectively antagonizes c-MYC and the genes controlled by this proto-oncogene.


Asunto(s)
Cromatina , Mieloma Múltiple , Humanos , Inhibidores de Proteasoma/farmacología , Complejo de la Endopetidasa Proteasomal/metabolismo , Mieloma Múltiple/tratamiento farmacológico , Genes myc
13.
Sci Adv ; 8(46): eabq0615, 2022 11 18.
Artículo en Inglés | MEDLINE | ID: mdl-36383649

RESUMEN

Chronic exposure to airborne carbon black ultrafine (nCB) particles generated from incomplete combustion of organic matter drives IL-17A-dependent emphysema. However, whether and how they alter the immune responses to lung cancer remains unknown. Here, we show that exposure to nCB particles increased PD-L1+ PD-L2+ CD206+ antigen-presenting cells (APCs), exhausted T cells, and Treg cells. Lung macrophages that harbored nCB particles showed selective mitochondrial structure damage and decreased oxidative respiration. Lung macrophages sustained the HIF1α axis that increased glycolysis and lactate production, culminating in an immunosuppressive microenvironment in multiple mouse models of non-small cell lung cancers. Adoptive transfer of lung APCs from nCB-exposed wild type to susceptible mice increased tumor incidence and caused early metastasis. Our findings show that nCB exposure metabolically rewires lung macrophages to promote immunosuppression and accelerates the development of lung cancer.


Asunto(s)
Neoplasias Pulmonares , Hollín , Ratones , Animales , Hollín/metabolismo , Material Particulado/efectos adversos , Neoplasias Pulmonares/etiología , Neoplasias Pulmonares/metabolismo , Macrófagos , Pulmón/metabolismo , Carbono/metabolismo , Microambiente Tumoral
14.
Aging Cell ; 20(8): e13432, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-34247441

RESUMEN

The rise of life expectancy of the human population is accompanied by the drastic increases of age-associated diseases, in particular Alzheimer's disease (AD), and underscores the need to understand how aging influences AD development. The Forkhead box O transcription factor 3 (FoxO3) is known to mediate aging and longevity downstream of insulin/insulin-like growth factor signaling across species. However, its function in the adult brain under physiological and pathological conditions is less understood. Here, we report a region and cell-type-specific regulation of FoxO3 in the central nervous system (CNS). We found that FoxO3 protein levels were reduced in the cortex, but not hippocampus, of aged mice. FoxO3 was responsive to insulin/AKT signaling in astrocytes, but not neurons. Using CNS Foxo3-deficient mice, we reveal that loss of FoxO3 led to cortical astrogliosis and altered lipid metabolism. This is associated with impaired metabolic homoeostasis and ß-amyloid (Aß) uptake in primary astrocyte cultures. These phenotypes can be reversed by expressing a constitutively active FOXO3 but not a FOXO3 mutant lacking the transactivation domain. Loss of FoxO3 in 5xFAD mice led to exacerbated Aß pathology and synapse loss and altered local response of astrocytes and microglia in the vicinity of Aß plaques. Astrocyte-specific overexpression of FOXO3 displayed opposite effects, suggesting that FoxO3 functions cell autonomously to mediate astrocyte activity and also interacts with microglia to address Aß pathology. Our studies support a protective role of astroglial FoxO3 against brain aging and AD.


Asunto(s)
Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/metabolismo , Astrocitos/metabolismo , Proteína Forkhead Box O3/deficiencia , Metabolismo de los Lípidos/fisiología , Enfermedad de Alzheimer/patología , Animales , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Transfección
15.
Cancers (Basel) ; 13(4)2021 Feb 17.
Artículo en Inglés | MEDLINE | ID: mdl-33671345

RESUMEN

Multiple myeloma and its precursor plasma cell dyscrasias affect 3% of the elderly population in the US. Proteasome inhibitors are an essential part of several standard drug combinations used to treat this incurable cancer. These drugs interfere with the main pathway of protein degradation and lead to the accumulation of damaged proteins inside cells. Despite promising initial responses, multiple myeloma cells eventually become drug resistant in most patients. The biology behind relapsed/refractory multiple myeloma is complex and poorly understood. Several studies provide evidence that in addition to the proteasome, mitochondrial proteases can also contribute to protein quality control outside of mitochondria. We therefore hypothesized that mitochondrial proteases might counterbalance protein degradation in cancer cells treated with proteasome inhibitors. Using clinical and experimental data, we found that overexpression of the mitochondrial matrix protease LonP1 (Lon Peptidase 1) reduces the efficacy of proteasome inhibitors. Some proteasome inhibitors partially crossinhibit LonP1. However, we show that the resistance effect of LonP1 also occurs when using drugs that do not block this protease, suggesting that LonP1 can compensate for loss of proteasome activity. These results indicate that targeting both the proteasome and mitochondrial proteases such as LonP1 could be beneficial for treatment of multiple myeloma.

16.
Sci Rep ; 10(1): 13942, 2020 08 18.
Artículo en Inglés | MEDLINE | ID: mdl-32811853

RESUMEN

Transcription is regulated through a dynamic interplay of DNA-associated proteins, and the composition of gene-regulatory complexes is subject to continuous adjustments. Protein alterations include post-translational modifications and elimination of individual polypeptides. Spatially and temporally controlled protein removal is, therefore, essential for gene regulation and accounts for the short half-life of many transcription factors. The ubiquitin-proteasome system is responsible for site- and target-specific ubiquitination and protein degradation. Specificity of ubiquitination is conferred by ubiquitin ligases. Cullin-RING complexes, the largest family of ligases, require multi-unit assembly around one of seven cullin proteins. To investigate the direct role of cullins in ubiquitination of DNA-bound proteins and in gene regulation, we analyzed their subcellular locations and DNA-affinities. We found CUL4A and CUL7 to be largely excluded from the nucleus, whereas CUL4B was primarily nuclear. CUL1,2,3, and 5 showed mixed cytosolic and nuclear expression. When analyzing chromatin affinity of individual cullins, we discovered that CUL1 preferentially associated with active promoter sequences and co-localized with 23% of all DNA-associated protein degradation sites. CUL1 co-distributed with c-MYC and specifically repressed nuclear-encoded mitochondrial and splicing-associated genes. These studies underscore the relevance of spatial control in chromatin-associated protein ubiquitination and define a novel role for CUL1 in gene repression.


Asunto(s)
Cromatina/metabolismo , Proteínas Cullin/genética , Proteínas Cullin/metabolismo , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , Núcleo Celular/metabolismo , Proteínas de Unión al ADN , Genes myc , Células HeLa , Humanos , Unión Proteica , Procesamiento Proteico-Postraduccional , Proteolisis , Factores de Transcripción/metabolismo , Transcripción Genética , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitinación
17.
FEBS Lett ; 590(7): 908-23, 2016 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-26832397

RESUMEN

The recruitment of transcription factors to promoters and enhancers is a critical step in gene regulation. Many of these proteins are quickly removed from DNA after they completed their function. Metabolic genes in particular are dynamically regulated and continuously adjusted to cellular requirements. Transcription factors controlling metabolism are therefore under constant surveillance by the ubiquitin-proteasome system, which can degrade DNA-bound proteins in a site-specific manner. Several of these metabolic transcription factors are critical to cancer cells, as they promote uncontrolled growth and proliferation. This review highlights recent findings in the emerging field of nuclear proteolysis and outlines novel paradigms for cancer treatment, with an emphasis on multiple myeloma.


Asunto(s)
Núcleo Celular/metabolismo , Regulación Neoplásica de la Expresión Génica , Modelos Biológicos , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Factores de Transcripción/metabolismo , Animales , Antineoplásicos/uso terapéutico , Núcleo Celular/efectos de los fármacos , Núcleo Celular/enzimología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Mieloma Múltiple/tratamiento farmacológico , Mieloma Múltiple/enzimología , Mieloma Múltiple/metabolismo , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/genética , Neoplasias/tratamiento farmacológico , Neoplasias/enzimología , Proteínas Nucleares/antagonistas & inhibidores , Proteínas Nucleares/genética , Péptido Hidrolasas/química , Péptido Hidrolasas/genética , Péptido Hidrolasas/metabolismo , Inhibidores de Proteasas/uso terapéutico , Complejo de la Endopetidasa Proteasomal/efectos de los fármacos , Complejo de la Endopetidasa Proteasomal/genética , Complejo de la Endopetidasa Proteasomal/metabolismo , Estabilidad Proteica/efectos de los fármacos , Proteolisis/efectos de los fármacos , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/genética , Complejos de Ubiquitina-Proteína Ligasa/antagonistas & inhibidores , Complejos de Ubiquitina-Proteína Ligasa/genética , Complejos de Ubiquitina-Proteína Ligasa/metabolismo
18.
Sci Rep ; 6: 38579, 2016 12 06.
Artículo en Inglés | MEDLINE | ID: mdl-27922125

RESUMEN

Estrogen, via estrogen receptor alpha (ERα), exerts several beneficial effects on metabolism and energy homeostasis by controlling size, enzymatic activity and hormonal content of adipose tissue. The actions of estrogen on sympathetic ganglia, which are key players in the browning process, are less well known. In the present study we show that ERß influences browning of subcutaneous adipose tissue (SAT) via its actions both on sympathetic ganglia and on the SAT itself. A 3-day-treatment with a selective ERß agonist, LY3201, induced browning of SAT in 1-year-old obese WT and ERα-/- female mice. Browning was associated with increased expression of ERß in the nuclei of neurons in the sympathetic ganglia, increase in tyrosine hydroxylase in both nerve terminals in the SAT and sympathetic ganglia neurons and an increase of ß3-adrenoceptor in the SAT. LY3201 had no effect on browning in young female or male mice. In the case of young females browning was already maximal while in males there was very little expression of ERß in the SAT and very little expression of the ß3-adrenoceptor. The increase in both sympathetic tone and responsiveness of adipocytes to catecholamines reveals a novel role for ERß in controlling browning of adipose tissue.


Asunto(s)
Tejido Adiposo Pardo/metabolismo , Receptor beta de Estrógeno/agonistas , Obesidad/metabolismo , Grasa Subcutánea Abdominal/metabolismo , Tejido Adiposo Pardo/efectos de los fármacos , Tejido Adiposo Blanco/metabolismo , Factores de Edad , Animales , Benzopiranos/farmacología , Receptor alfa de Estrógeno/genética , Receptor alfa de Estrógeno/metabolismo , Receptor beta de Estrógeno/genética , Receptor beta de Estrógeno/metabolismo , Femenino , Expresión Génica , Masculino , Ratones , Ratones Noqueados , Ratones Obesos , Modelos Biológicos , Obesidad/genética , Factores Sexuales , Grasa Subcutánea Abdominal/efectos de los fármacos , Sistema Nervioso Simpático/efectos de los fármacos
19.
J Mol Med (Berl) ; 92(11): 1179-200, 2014 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-25081415

RESUMEN

UNLABELLED: The existence of a link between estrogen deprivation and osteoarthritis (OA) in postmenopausal women suggests that 17ß-estradiol (17ß-E2) may be a modulator of cartilage homeostasis. Here, we demonstrate that 17ß-E2 stimulates, via its receptor human estrogen receptor α 66 (hERα66), type II collagen expression in differentiated and dedifferentiated (reflecting the OA phenotype) articular chondrocytes. Transactivation of type II collagen gene (COL2A1) by ligand-independent transactivation domain (AF-1) of hERα66 was mediated by "GC" binding sites of the -266/-63-bp promoter, through physical interactions between ERα, Sp1/Sp3, Sox9, and p300, as demonstrated in chromatin immunoprecipitation (ChIP) and Re-Chromatin Immuno-Precipitation (Re-ChIP) assays in primary and dedifferentiated cells. 17ß-E2 and hERα66 increased the DNA-binding activities of Sp1/Sp3 and Sox-9 to both COL2A1 promoter and enhancer regions. Besides, Sp1, Sp3, and Sox-9 small interfering RNAs (siRNAs) prevented hERα66-induced transactivation of COL2A1, suggesting that these factors and their respective cis-regions are required for hERα66-mediated COL2A1 up-regulation. Our results highlight the genomic pathway by which 17ß-E2 and hERα66 modulate Sp1/Sp3 heteromer binding activity and simultaneously participate in the recruitment of the essential factors Sox-9 and p300 involved respectively in the chondrocyte-differentiated status and COL2A1 transcriptional activation. These novel findings could therefore be attractive for tissue engineering of cartilage in OA, by the fact that 17ß-E2 could promote chondrocyte redifferentiation. KEY MESSAGES: 17ß-E2 up-regulates type II collagen gene expression in articular chondrocytes. An ERα66/Sp1/Sp3/Sox-9/p300 protein complex mediates this stimulatory effect. This heteromeric complex interacts and binds to Col2a1 promoter and enhancer in vivo. Our findings highlight a new regulatory mechanism for 17ß-E2 action in chondrocytes. 17ß-E2 might be an attractive candidate for cartilage engineering applications.


Asunto(s)
Condrocitos/citología , Colágeno Tipo II/metabolismo , Estradiol/farmacología , Receptor alfa de Estrógeno/metabolismo , Factor de Transcripción SOX9/metabolismo , Factor de Transcripción Sp1/metabolismo , Factor de Transcripción Sp3/metabolismo , Animales , Sitios de Unión , Cartílago Articular/citología , Diferenciación Celular , Colágeno Tipo II/genética , Humanos , Masculino , Fenotipo , Regiones Promotoras Genéticas , ARN Interferente Pequeño/metabolismo , Conejos , Activación Transcripcional , Regulación hacia Arriba
20.
Tissue Eng Part C Methods ; 19(7): 550-67, 2013 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-23270543

RESUMEN

Cartilage healing by tissue engineering is an alternative strategy to reconstitute functional tissue after trauma or age-related degeneration. However, chondrocytes, the major player in cartilage homeostasis, do not self-regenerate efficiently and lose their phenotype during osteoarthritis. This process is called dedifferentiation and also occurs during the first expansion step of autologous chondrocyte implantation (ACI). To ensure successful ACI therapy, chondrocytes must be differentiated and capable of synthesizing hyaline cartilage matrix molecules. We therefore developed a safe procedure for redifferentiating human chondrocytes by combining appropriate physicochemical factors: hypoxic conditions, collagen scaffolds, chondrogenic factors (bone morphogenetic protein-2 [BMP-2], and insulin-like growth factor I [IGF-I]) and RNA interference targeting the COL1A1 gene. Redifferentiation of dedifferentiated chondrocytes was evaluated using gene/protein analyses to identify the chondrocyte phenotypic profile. In our conditions, under BMP-2 treatment, redifferentiated and metabolically active chondrocytes synthesized a hyaline-like cartilage matrix characterized by type IIB collagen and aggrecan molecules without any sign of hypertrophy or osteogenesis. In contrast, IGF-I increased both specific and noncharacteristic markers (collagens I and X) of chondrocytes. The specific increase in COL2A1 gene expression observed in the BMP-2 treatment was shown to involve the specific enhancer region of COL2A1 that binds the trans-activators Sox9/L-Sox5/Sox6 and Sp1, which are associated with a decrease in the trans-inhibitors of COL2A1, c-Krox, and p65 subunit of NF-kappaB. Our procedure in which BMP-2 treatment under hypoxia is associated with a COL1A1 siRNA, significantly increased the differentiation index of chondrocytes, and should offer the opportunity to develop new ACI-based therapies in humans.


Asunto(s)
Proteína Morfogenética Ósea 2/farmacología , Condrocitos/citología , Colágeno/farmacología , Matriz Extracelular/metabolismo , Cartílago Hialino/metabolismo , ARN Interferente Pequeño/metabolismo , Andamios del Tejido/química , Anciano , Anciano de 80 o más Años , Agrecanos/genética , Agrecanos/metabolismo , Animales , Bovinos , Diferenciación Celular/efectos de los fármacos , Hipoxia de la Célula/efectos de los fármacos , Hipoxia de la Célula/genética , Células Cultivadas , Condrocitos/efectos de los fármacos , Condrocitos/metabolismo , Condrocitos/ultraestructura , Condrogénesis/efectos de los fármacos , Cilios/efectos de los fármacos , Cilios/metabolismo , Colágeno/genética , Colágeno/metabolismo , ADN/metabolismo , Elementos de Facilitación Genéticos/genética , Matriz Extracelular/efectos de los fármacos , Adhesiones Focales/efectos de los fármacos , Adhesiones Focales/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Cartílago Hialino/citología , Persona de Mediana Edad , Fenotipo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factor de Transcripción SOX9/genética , Factor de Transcripción SOX9/metabolismo , Transcripción Genética/efectos de los fármacos
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