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1.
Biochem Biophys Res Commun ; 696: 149422, 2024 02 12.
Artículo en Inglés | MEDLINE | ID: mdl-38183795

RESUMEN

Identification and functional analysis of key genes regulated by the circadian clock system will provide a comprehensive understanding of the underlying mechanisms through which circadian clock disruption impairs the health of living organisms. The initial phase involved bioinformatics analysis, drawing insights from three RNA-seq datasets (GSE184303, GSE114400, and GSE199061) derived from wild-type mouse liver tissues, which encompassed six distinct time points across a day. As expected, 536 overlapping genes exhibiting rhythmic expression patterns were identified. By intersecting these genes with differentially expressed genes (DEGs) originating from liver RNA-seq data at two representative time points (circadian time, CT: CT2 and CT14) in global Bmal1 knockout mice (Bmal1-/-), hepatocyte-specific Bmal1 knockout mice (L-Bmal1-/-), and their corresponding control groups, 80 genes potentially regulated by BMAL1 (referred to as BMAL1-regulated genes, BRGs) were identified. These genes were significantly enriched in glycolipid metabolism, immune response, and tumorigenesis pathways. Eight BRGs (Nr1d1, Cry1, Gys2, Homer2, Serpina6, Slc2a2, Nmrk1, and Upp2) were selected to validate their expression patterns in both control and L-Bmal1-/- mice livers over 24 h. Real-time quantitative polymerase chain reaction results demonstrated a comprehensive loss of rhythmic expression patterns in the eight selected BRGs in L-Bmal1-/- mice, in contrast to the discernible rhythmic patterns observed in the livers of control mice. Additionally, significant reductions in the expression levels of these selected BRGs, excluding Cry1, were also observed in L-Bmal1-/- mice livers. Chromatin immunoprecipitation (ChIP)-seq (GSE13505 and GSE39860) and JASPAR analyses validated the rhythmic binding of BMAL1 to the promoter and intron regions of these genes. Moreover, the progression of conditions, from basic steatosis to non-alcoholic fatty liver disease, and eventual malignancy, demonstrated a continuous gradual decline in Bmal1 transcripts in the human liver. Combining the aforementioned BRGs with DEGs derived from human liver cancer datasets identified Gys2 and Upp2 as potential node genes bridging the circadian clock system and hepatocellular carcinoma (HCC). In addition, CCK8 and wound healing assays demonstrated that the overexpression of human GYS2 and UPP2 proteins inhibited the proliferation and migration of HepG2 cells, accompanied by elevated expression of p53, a tumor suppressor protein. In summary, this study systematically identified rhythmic genes in the mouse liver, and a subset of circadian genes potentially regulated by BMAL1. Two circadian genes, Gys2 and Upp2, have been proposed and validated as potential candidates for advancing the prevention and treatment of HCC.


Asunto(s)
Carcinoma Hepatocelular , Relojes Circadianos , Neoplasias Hepáticas , Animales , Humanos , Ratones , Factores de Transcripción ARNTL/genética , Factores de Transcripción ARNTL/metabolismo , Carcinoma Hepatocelular/patología , Relojes Circadianos/genética , Ritmo Circadiano/genética , Proteínas CLOCK/genética , Regulación de la Expresión Génica , Proteínas de Andamiaje Homer/metabolismo , Hígado/metabolismo , Neoplasias Hepáticas/patología , Ratones Noqueados , Uridina Fosforilasa/metabolismo , Glucógeno Sintasa/metabolismo
2.
ACS Nano ; 18(6): 4822-4839, 2024 Feb 13.
Artículo en Inglés | MEDLINE | ID: mdl-38285698

RESUMEN

Efficiently delivering exogenous materials into primary neurons and neural stem cells (NSCs) has long been a challenge in neurobiology. Existing methods have struggled with complex protocols, unreliable reproducibility, high immunogenicity, and cytotoxicity, causing a huge conundrum and hindering in-depth analyses. Here, we establish a cutting-edge method for transfecting primary neurons and NSCs, named teleofection, by a two-step process to enhance the formation of biocompatible calcium phosphate (CaP) nanoparticles. Teleofection enables both nucleic acid and protein transfection into primary neurons and NSCs, eliminating the need for specialized skills and equipment. It can easily fine-tune transfection efficiency by adjusting the incubation time and nanoparticle quantity, catering to various experimental requirements. Teleofection's versatility allows for the delivery of different cargos into the same cell culture, whether simultaneously or sequentially. This flexibility proves invaluable for long-term studies, enabling the monitoring of neural development and synapse plasticity. Moreover, teleofection ensures the consistent and robust expression of delivered genes, facilitating molecular and biochemical investigations. Teleofection represents a significant advancement in neurobiology, which has promise to transcend the limitations of current gene delivery methods. It offers a user-friendly, cost-effective, and reproducible approach for researchers, potentially revolutionizing our understanding of brain function and development.


Asunto(s)
Nanopartículas , Células-Madre Neurales , Ácidos Nucleicos , Ácidos Nucleicos/metabolismo , Reproducibilidad de los Resultados , Células-Madre Neurales/metabolismo , Nanopartículas/química , Transfección , Fosfatos de Calcio/química
3.
Front Cell Dev Biol ; 11: 1075215, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36910151

RESUMEN

Long-term maintenance of synaptic connections is important for brain function, which depends on varying proteostatic regulations to govern the functional integrity of neuronal proteomes. Proteostasis supports an interconnection of pathways that regulates the fate of proteins from synthesis to degradation. Defects in proteostatic signaling are associated with age-related functional decline and neurodegenerative diseases. Recent studies have advanced our knowledge of how cells have evolved distinct mechanisms to safely control protein homeostasis during synthesis, folding and degradation, and in different subcellular organelles and compartments. Neurodegeneration occurs when these protein quality controls are compromised by accumulated pathogenic proteins or aging to an irreversible state. Consequently, several therapeutic strategies, such as targeting the unfolded protein response and autophagy pathways, have been developed to reduce the burden of misfolded proteins and proved useful in animal models. Here, we present a brief overview of the molecular mechanisms involved in maintaining proteostatic networks, along with some examples linking dysregulated proteostasis to neuronal diseases.

4.
Trends Genet ; 38(10): 991-995, 2022 10.
Artículo en Inglés | MEDLINE | ID: mdl-35637074

RESUMEN

Polyploidization and polyploidy reversal (depolyploidization) are crucial pathways to conversely alter genomic contents in organisms. Understanding the mechanisms switching between polyploidization and polyploidy reversal should broaden our knowledge of the generation of pathological polyploidy and pave a new path to prevent related diseases.


Asunto(s)
Mitosis , Poliploidía , Humanos
5.
Mol Psychiatry ; 27(8): 3343-3354, 2022 08.
Artículo en Inglés | MEDLINE | ID: mdl-35491410

RESUMEN

Immune dysregulation plays a key role in the pathogenesis of autism. Changes occurring at the systemic level, from brain inflammation to disturbed innate/adaptive immune in the periphery, are frequently observed in patients with autism; however, the intrinsic mechanisms behind them remain elusive. We hypothesize a common etiology may lie in progenitors of different types underlying widespread immune dysregulation. By single-cell RNA sequencing (sc-RNA seq), we trace the developmental origins of immune dysregulation in a mouse model of idiopathic autism. It is found that both in aorta-gonad-mesonephros (AGM) and yolk sac (YS) progenitors, the dysregulation of HDAC1-mediated epigenetic machinery alters definitive hematopoiesis during embryogenesis and downregulates the expression of the AP-1 complex for microglia development. Subsequently, these changes result in the dysregulation of the immune system, leading to gut dysbiosis and hyperactive microglia in the brain. We further confirm that dysregulated immune profiles are associated with specific microbiota composition, which may serve as a biomarker to identify autism of immune-dysregulated subtypes. Our findings elucidate a shared mechanism for the origin of immune dysregulation from the brain to the gut in autism and provide new insight to dissecting the heterogeneity of autism, as well as the therapeutic potential of targeting immune-dysregulated autism subtypes.


Asunto(s)
Trastorno Autístico , Ratones , Animales , Trastorno Autístico/genética , Mesonefro , Saco Vitelino/fisiología , Gónadas , Epigénesis Genética/genética , Modelos Animales de Enfermedad
6.
Theriogenology ; 174: 9-19, 2021 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-34416563

RESUMEN

The circadian clock system plays an important role in regulating testosterone synthesis in mammals. Male Bmal1-/- mice are infertile with low serum testosterone levels and decreased expression of testicular steroidogenic genes, suggesting that circadian clock genes regulate testosterone biosynthesis by activating steroidogenic gene transcription. However, whether the circadian clock regulates testosterone production via other genes remains unknown. Using Bmal1-/- mice and their wild-type (WT) siblings, we aimed to identify additional genes by which the circadian clock regulates testosterone synthesis. WT and Bmal1-/- mouse testes sections had similar normal morphologies, although there was a decrease in testicular spermatozoa in the Bmal1-/- mice. Low serum testosterone levels were detected in the Bmal1-/- mice. RNA sequencing identified 37 and 48 genes that were differentially expressed between WT and Bmal1-/- mouse testes at circadian time (CT2 and CT14), respectively. The cholesterol metabolism pathway was significantly enriched in the KEGG pathway analysis, and there was lower expression of three apolipoprotein genes (Apoa1, Apoa2, and Apoc3) at CT2 in the testes of Bmal1-/- mice than in those of WT mice. These decreases in Apoa1, Apoa2, and Apoc3 expression were verified by quantitative polymerase chain reaction analysis, which also revealed downregulation of the expression of the circadian clock (Per2, Dbp, and Nr1d1) and steroidogenic (StAR, Cyp11a1, and Hsd17b3) genes. The expression of circadian clock genes was relatively stable in WT mice over a 20-h period, whereas there was clear circadian rhythmic expression of Apoa1, Apoa2, Apoc3, StAR, Cyp11a1, Hsd3b2, and Hsd17b3. Bmal1-/- mice showed severely reduced expression of testicular circadian clock genes at three time points (CT4, CT12, and CT20), and a reduction in mRNA expression levels of Apo (Apoa1, Apoa2, and Apoc3) and steroidogenic (StAR, Cyp11a1, Hsd3b2, and Hsd17b3) genes. Oil Red O staining showed decreased lipid aggregation in the Leydig cells of Bmal1-/- mouse testes. Considering the vital role of Apo genes in high-density lipoprotein formation and cholesterol transport, the present data suggest that the circadian clock system regulates testosterone production by orchestrating the rhythmic expression of Apo genes. These data extend our understanding of the role of the circadian clock in regulating testosterone production in mammals.


Asunto(s)
Relojes Circadianos , Testosterona , Animales , Apolipoproteínas , Proteínas CLOCK/genética , Ritmo Circadiano , Expresión Génica , Regulación de la Expresión Génica , Masculino , Ratones , Testículo
7.
Neuropsychopharmacology ; 46(9): 1669-1679, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-33941859

RESUMEN

Susceptibility or resilience to posttraumatic stress disorder (PTSD) depends on one's ability to appropriately adjust synaptic plasticity for coping with the traumatic experience. Activity-regulated mRNA translation synthesizes plasticity-related proteins to support long-term synaptic changes and memory. Hence, cytoplasmic polyadenylation element-binding protein 3-knockout (CPEB3-KO) mice, showing dysregulated translation-associated synaptic rigidity, may be susceptible to PTSD-like behavior. Here, using a context-dependent auditory fear conditioning and extinction paradigm, we found that CPEB3-KO mice exhibited traumatic intensity-dependent PTSD-like fear memory. A genome-wide screen of CPEB3-bound transcripts revealed that Nr3c1, encoding glucocorticoid receptor (GR), was translationally suppressed by CPEB3. Thus, CPEB3-KO neurons with elevated GR expression exhibited increased corticosterone-induced calcium influx and decreased mRNA and protein levels of brain-derived neurotrophic factor (Bdnf). Moreover, the reduced expression of BDNF was associated with increased GR level during fear extinction in CPEB3-KO hippocampi. Intracerebroventricular delivery of BDNF before extinction training mitigated spontaneous fear intrusion in CPEB3-KO mice during extinction recall. Analysis of two GEO datasets revealed decreased transcriptomic expression of CPEB3 but not NR3C1 in peripheral blood mononuclear cells of humans with PTSD. Collectively, this study reveals that CPEB3, as a potential PTSD-risk gene, downregulates Nr3c1 translation to maintain proper GR-BDNF signaling for fear extinction.


Asunto(s)
Miedo , Trastornos por Estrés Postraumático , Animales , Extinción Psicológica , Leucocitos Mononucleares , Ratones , Ratones Noqueados , Biosíntesis de Proteínas , Proteínas de Unión al ARN , Receptores de Glucocorticoides/genética
8.
Am J Physiol Endocrinol Metab ; 320(4): E747-E759, 2021 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-33554778

RESUMEN

Prostaglandin G/H synthase 2 (PTGS2) is a rate-limiting enzyme in prostaglandin synthesis. The present study assessed the role of the uterine circadian clock on Ptgs2 transcription in response to steroid hormones during early pregnancy. We demonstrated that the core clock genes (Bmal1, Per2, Nr1d1, and Dbp), Vegf, and Ptgs2, and their encoded proteins, have rhythmic expression in the mouse uterus from days 3.5 to 4.5 (D3.5-4.5) of pregnancy. Progesterone (P4) treatment of cultured uterus endometrial stromal cells (UESCs) isolated from mPer2Luciferase reporter gene knock-in mice on D4 induced a phase shift in PER2::LUCIFERASE oscillations. This P4-induced phase shift of PER2::LUCIFERASE oscillations was significantly attenuated by the P4 antagonist RU486. Additionally, the amplitude of PER2::LUCIFERASE oscillations was increased by estradiol (E2) treatment in the presence of P4. Consistently, the mRNA levels of clock genes (Bmal1 and Per2), Vegf, and Ptgs2 were markedly increased by E2 treatment of UESCs in the presence of P4. Treatment with E2 also promoted prostaglandin E2 (PGE2) synthesis by UESCs. Depletion of Bmal1 in UESCs by small-interfering RNA (siRNA) decreased the transcript levels of clock genes (Nr1d1 and Dbp), Vegf, and Ptgs2 compared with nonsilencing siRNA treatment. Bmal1 knockdown also inhibited PGE2 synthesis. Moreover, the mRNA expression levels of clock genes (Nr1d1 and Dbp), Vegf, and Ptgs2, and their respective proteins were significantly decreased in the uterus of Bmal1-/- mice. Thus, these data suggest that Bmal1 in mice promotes PGE2 synthesis by upregulating Ptgs2 in response to increases in E2 on D4 of pregnancy.NEW & NOTEWORTHY Rhythmic expression of Bmal1 and Ptgs2 was observed in the uterus isolated from D3.5-4.5 of pregnant mice. E2 increased the expression of Bmal1 and Ptg2 in UESCs isolated from mice on D4. The expression of Ptgs2 was significantly decreased in Bmal1-siRNA treated UESCs. Bmal1 knockdown also inhibited PGE2 synthesis. Thus, these data suggest that Bmal1 in mice promotes PGE2 synthesis by upregulating Ptgs2 in response to increases in E2 on D4 of pregnancy.


Asunto(s)
Factores de Transcripción ARNTL/fisiología , Ciclooxigenasa 2/genética , Dinoprostona/biosíntesis , Estradiol/sangre , Factores de Transcripción ARNTL/genética , Animales , Células Cultivadas , Ciclooxigenasa 2/metabolismo , Estradiol/farmacología , Femenino , Edad Gestacional , Masculino , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Embarazo , Progesterona/farmacología , Activación Transcripcional/efectos de los fármacos , Útero/efectos de los fármacos , Útero/metabolismo
9.
J Cell Physiol ; 236(9): 6706-6725, 2021 09.
Artículo en Inglés | MEDLINE | ID: mdl-33598947

RESUMEN

Testosterone is produced by Leydig cells (LCs) and undergoes diurnal changes in serum levels in rats, mice, and humans, but little is known in goats. The present study revealed that goat serum testosterone levels displayed diurnal rhythmic changes (peak time at ZT11.2). Immunohistochemical staining showed that BMAL1, a circadian clock protein, is highly expressed in goat LCs. ELISA revealed that both hCG (0-5 IU/ml) and 22R-OH-cholesterol (0-30 µM) addition stimulated testosterone synthesis in primary goat LCs in a dose-dependent manner. Treating goat LCs with hCG (5 IU/ml) significantly increased intracellular cAMP levels. Additionally, real-time quantitative polymerase chain reaction (PCR) analysis revealed that the circadian clock (BMAL1, PER1, PER2, DBP, and NR1D1) and steroidogenesis-related genes (SF1, NUR77, StAR, HSD3B2, CYP17A1, CYP11A1, and HSD17B3) showed rhythmic expression patterns in goat LCs following dexamethasone synchronization. Several Bmal1-Luc circadian oscillations were clearly observed in dexamethasone-treated goat LCs transfected with the pLV6-Bmal1-Luc plasmid. BMAL1 knockdown significantly downregulated mRNA levels of PER2, NR1D1, DBP, StAR, HSD3B2, SF1, NUR77, and GATA4, and dramatically decreased StAR and HSD3B2 protein levels and testosterone production. In contrast, BMAL1 overexpression significantly increased the mRNA and protein expression levels of StAR and HSD17B3 and enhanced testosterone production. Reporter assays revealed that goat BMAL1, or in combination with mouse CLOCK, activated goat HSD17B3 transcription in vitro. These data indicate that BMAL1 contributes to testosterone production by regulating transcription of steroidogenesis-related genes in goat LCs, providing a basis for further exploring the underlying mechanism by which the circadian clock regulates ruminant reproductive capability.


Asunto(s)
Factores de Transcripción ARNTL/genética , Relojes Circadianos/genética , Regulación de la Expresión Génica , Cabras/genética , Células Intersticiales del Testículo/metabolismo , Fosfoproteínas/genética , Testosterona/biosíntesis , Transcripción Genética , 17-Hidroxiesteroide Deshidrogenasas/metabolismo , Factores de Transcripción ARNTL/metabolismo , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Relojes Circadianos/efectos de los fármacos , Dexametasona/farmacología , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Cabras/sangre , Humanos , Hidroxicolesteroles/metabolismo , Células Intersticiales del Testículo/efectos de los fármacos , Masculino , Modelos Biológicos , Testosterona/sangre , Transcripción Genética/efectos de los fármacos
10.
Nat Commun ; 12(1): 645, 2021 01 28.
Artículo en Inglés | MEDLINE | ID: mdl-33510150

RESUMEN

Hepatocellular carcinoma (HCC) is the most predominant primary malignancy in the liver. Genotoxic and genetic models have revealed that HCC cells are derived from hepatocytes, but where the critical region for tumor foci emergence is and how this transformation occurs are still unclear. Here, hyperpolyploidization of hepatocytes around the centrilobular (CL) region is demonstrated to be closely linked with the development of HCC cells after diethylnitrosamine treatment. We identify the CL region as a dominant lobule for accumulation of hyperpolyploid hepatocytes and preneoplastic tumor foci formation. We also demonstrate that upregulation of Aurkb plays a critical role in promoting hyperpolyploidization. Increase of AURKB phosphorylation is detected on the midbody during cytokinesis, causing abscission failure and hyperpolyploidization. Pharmacological inhibition of AURKB dramatically reduces nucleus size and tumor foci number surrounding the CL region in diethylnitrosamine-treated liver. Our work reveals an intimate molecular link between pathological hyperpolyploidy of CL hepatocytes and transformation into HCC cells.


Asunto(s)
Carcinoma Hepatocelular/genética , Transformación Celular Neoplásica/genética , Hepatocitos/metabolismo , Neoplasias Hepáticas/genética , Hígado/metabolismo , Poliploidía , Lesiones Precancerosas/genética , Animales , Carcinoma Hepatocelular/inducido químicamente , Carcinoma Hepatocelular/metabolismo , Transformación Celular Neoplásica/inducido químicamente , Transformación Celular Neoplásica/metabolismo , Células Cultivadas , Dietilnitrosamina/toxicidad , Femenino , Hepatocitos/efectos de los fármacos , Humanos , Hígado/efectos de los fármacos , Hígado/patología , Neoplasias Hepáticas/inducido químicamente , Neoplasias Hepáticas/metabolismo , Masculino , Ratones Endogámicos C57BL , Ratones Endogámicos ICR , Ratones Endogámicos NOD , Ratones Noqueados , Ratones SCID , Microscopía Confocal , Lesiones Precancerosas/inducido químicamente , Lesiones Precancerosas/metabolismo
11.
J Cell Physiol ; 236(3): 2023-2035, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-32730662

RESUMEN

The downregulation of melatonin receptor 1A (MTNR1A) is associated with a range of pathological conditions, including membranous nephropathy. Knowledge of the mechanism underlying MTNR1A expression has been limited to the transcriptional regulation level. Here, RNA interference screening in human kidney cells revealed that heterogeneous nuclear ribonucleoprotein L (hnRNPL) upregulated MTNR1A RNA post-transcriptionally. hnRNPL knockdown or overexpression led to increased or decreased levels of cyclic adenosine monophosphate-responsive element-binding protein phosphorylation, respectively. Molecular studies showed that cytoplasmic hnRNPL exerts a stabilizing effect on the MTNR1A transcript through CA-repeat elements in its coding region. Further studies revealed that the interaction between hnRNPL and MTNR1A serves to protect MNTR1A RNA degradation by the exosome component 10 protein. MTNR1A, but not hnRNPL, displays a diurnal rhythm in mouse kidneys. Enhanced levels of MTNR1A recorded at midnight correlated with robust binding activity between cytoplasmic hnRNPL and the MTNR1A transcript. Both hnRNPL and MTNR1A were decreased in the cytoplasm of tubular epithelial cells from experimental membranous nephropathy kidneys, supporting their clinical relevance. Collectively, our data identified cytoplasmic hnRNPL as a novel player in the upregulation of MTNR1A expression in renal tubular epithelial cells, and as a potential therapeutic target.


Asunto(s)
Citoplasma/metabolismo , Ribonucleoproteína Heterogénea-Nuclear Grupo L/metabolismo , Túbulos Renales/metabolismo , Receptor de Melatonina MT1/genética , Animales , Línea Celular , Ritmo Circadiano/genética , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Células Epiteliales/metabolismo , Exorribonucleasas/metabolismo , Complejo Multienzimático de Ribonucleasas del Exosoma/metabolismo , Glomerulonefritis Membranosa/genética , Glomerulonefritis Membranosa/patología , Humanos , Túbulos Renales/patología , Ratones Endogámicos BALB C , Modelos Biológicos , Sistemas de Lectura Abierta/genética , Fosforilación , Estabilidad del ARN/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptor de Melatonina MT1/metabolismo , Secuencias Repetitivas de Ácidos Nucleicos/genética , Regulación hacia Arriba/genética
12.
J Biomed Sci ; 27(1): 52, 2020 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-32295602

RESUMEN

BACKGROUND: Alveologenesis is the final stage of lung development to form air-exchanging units between alveoli and blood vessels. Genetic susceptibility or hyperoxic stress to perturb this complicated process can cause abnormal enlargement of alveoli and lead to bronchopulmonary dysplasia (BPD)-associated emphysema. Platelet-derived growth factor receptor α (PDGFRα) signaling is crucial for alveolar myofibroblast (MYF) proliferation and its deficiency is associated with risk of BPD, but posttranscriptional mechanisms regulating PDGFRα synthesis during lung development remain largely unexplored. Cytoplasmic polyadenylation element-binding protein 2 (CPEB2) is a sequence-specific RNA-binding protein and translational regulator. Because CPEB2-knockout (KO) mice showed emphysematous phenotypes, we investigated how CPEB2-controlled translation affects pulmonary development and function. METHODS: Respiratory and pulmonary functions were measured by whole-body and invasive plethysmography. Histological staining and immunohistochemistry were used to analyze morphology, proliferation, apoptosis and cell densities from postnatal to adult lungs. Western blotting, RNA-immunoprecipitation, reporter assay, primary MYF culture and ectopic expression rescue were performed to demonstrate the role of CPEB2 in PDGFRα mRNA translation and MYF proliferation. RESULTS: Adult CPEB2-KO mice showed emphysema-like dysfunction. The alveolar structure in CPEB2-deficient lungs appeared normal at birth but became simplified through the alveolar stage of lung development. In CPEB2-null mice, we found reduced proliferation of MYF progenitors during alveolarization, abnormal deposition of elastin and failure of alveolar septum formation, thereby leading to enlarged pulmonary alveoli. We identified that CPEB2 promoted PDGFRα mRNA translation in MYF progenitors and this positive regulation could be disrupted by H2O2, a hyperoxia-mimetic treatment. Moreover, decreased proliferating ability in KO MYFs due to insufficient PDGFRα expression was rescued by ectopic expression of CPEB2, suggesting an important role of CPEB2 in upregulating PDGFRα signaling for pulmonary alveologenesis. CONCLUSIONS: CPEB2-controlled translation, in part through promoting PDGFRα expression, is indispensable for lung development and function. Since defective pulmonary PDGFR signaling is a key feature of human BPD, CPEB2 may be a risk factor for BPD.


Asunto(s)
Proliferación Celular , Miofibroblastos/fisiología , Factor de Crecimiento Derivado de Plaquetas/genética , Biosíntesis de Proteínas , Alveolos Pulmonares/crecimiento & desarrollo , Proteínas de Unión al ARN/genética , Animales , Ratones , Ratones Noqueados , Factor de Crecimiento Derivado de Plaquetas/metabolismo , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/metabolismo
13.
Int J Mol Sci ; 20(2)2019 Jan 13.
Artículo en Inglés | MEDLINE | ID: mdl-30642126

RESUMEN

Industrialized society-caused dysregular human behaviors and activities such as overworking, excessive dietary intake, and sleep deprivation lead to perturbations in the metabolism and the development of metabolic syndrome. Non-alcoholic fatty liver disease (NAFLD), the most common chronic liver disease worldwide, affects around 30% and 25% of people in Western and Asian countries, respectively, which leads to numerous medical costs annually. Insulin resistance is the major hallmark of NAFLD and is crucial in the pathogenesis and for the progression from NAFLD to non-alcoholic steatohepatitis (NASH). Excessive dietary intake of saturated fats and carbohydrate-enriched foods contributes to both insulin resistance and NAFLD. Once NAFLD is established, insulin resistance can promote the progression to the more severe state of liver endangerment like NASH. Here, we review current and potential studies for understanding the complexity between insulin-regulated glycolytic and lipogenic homeostasis and the underlying causes of NAFLD. We discuss how disruption of the insulin signal is associated with various metabolic disorders of glucoses and lipids that constitute both the metabolic syndrome and NAFLD.


Asunto(s)
Glucosa/metabolismo , Resistencia a la Insulina , Metabolismo de los Lípidos , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Progresión de la Enfermedad , Glucólisis , Homeostasis , Humanos , Lipogénesis , Enfermedad del Hígado Graso no Alcohólico/epidemiología
14.
Front Mol Neurosci ; 11: 428, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30534052

RESUMEN

Members of the ribonuclease A (RNase A) superfamily regulate various physiological processes. RNase A, the best-studied member of the RNase A superfamily, is widely expressed in different tissues, including brains. We unexpectedly found that RNase A can trigger proliferation of neuronal progenitor cells (NPC) both in vitro and in vivo. RNase A treatment induced cell proliferation in dissociated neuronal cultures and increased cell mass in neurosphere cultures. BrdU (5-Bromo-2'-Deoxyuridine) labeling confirmed the effect of RNase A on cell proliferation. Those dividing cells were Nestin- and SOX2-positive, suggesting that RNase A triggers NPC proliferation. The proliferation inhibitor Ara-C completely suppressed the effect of RNase A on NPC counts, further supporting that RNase A increases NPC number mainly by promoting proliferation. Moreover, we found that RNase A treatment increased ERK phosphorylation and blockade of the ERK pathway inhibited the effect of RNase A on NPC proliferation. Intracerebroventricular injection of RNase A into mouse brain increased the population of 5-ethynyl-2'-deoxyuridine (EdU) or BrdU-labeled cells in the subventricular zone. Those RNase A-induced NPCs were able to migrate into other brain areas, including hippocampus, amygdala, cortex, striatum, and thalamus. In conclusion, our study shows that RNase A promotes proliferation of NPCs via an ERK-dependent pathway and further diversifies the physiological functions of the RNase A family.

15.
Nat Commun ; 8(1): 2238, 2017 12 21.
Artículo en Inglés | MEDLINE | ID: mdl-29269828

RESUMEN

Liver metabolism undergoes robust circadian oscillations in gene expression and enzymatic activity essential for liver homeostasis, but whether the circadian clock controls homeostatic self-renewal of hepatocytes is unknown. Here we show that hepatocyte polyploidization is markedly accelerated around the central vein, the site of permanent cell self-renewal, in mice deficient in circadian Period genes. In these mice, a massive accumulation of hyperpolyploid mononuclear and binuclear hepatocytes occurs due to impaired mitogen-activated protein kinase phosphatase 1 (Mkp1)-mediated circadian modulation of the extracellular signal-regulated kinase (Erk1/2) activity. Time-lapse imaging of hepatocytes suggests that the reduced activity of Erk1/2 in the midbody during cytokinesis results in abscission failure, leading to polyploidization. Manipulation of Mkp1 phosphatase activity is sufficient to change the ploidy level of hepatocytes. These data provide clear evidence that the Period genes not only orchestrate dynamic changes in metabolic activity, but also regulate homeostatic self-renewal of hepatocytes through Mkp1-Erk1/2 signaling pathway.


Asunto(s)
Fosfatasa 1 de Especificidad Dual/metabolismo , Hepatocitos/metabolismo , Hígado/metabolismo , Sistema de Señalización de MAP Quinasas/fisiología , Proteínas Circadianas Period/genética , Poliploidía , Animales , Relojes Circadianos/genética , Hepatocitos/citología , Hepatocitos/patología , Hígado/citología , Hígado/patología , Ratones , Ratones Noqueados , Microscopía , Imagen de Lapso de Tiempo
16.
Cell Rep ; 21(8): 2264-2276, 2017 Nov 21.
Artículo en Inglés | MEDLINE | ID: mdl-29166615

RESUMEN

Mammalian olfactory bulbs (OBs) require continuous replenishment of interneurons (mainly granule cells [GCs]) to support local circuits throughout life. Two spatiotemporally distinct waves of postnatal neurogenesis contribute to expanding and maintaining the GC pool. Although neonate-born GCs have a higher survival rate than adult-born GCs, the molecular mechanism underlying this survival remains unclear. Here, we find that cytoplasmic polyadenylation element-binding protein 4 (CPEB4) acts as a survival factor exclusively for early postnatal GCs. In mice, during the first 2 postnatal weeks, olfactory experience initiated CPEB4-activated c-Fos mRNA translation. In CPEB4-knockout mice, c-FOS insufficiency reduced neurotrophic signaling to impair GC survival and cause OB hypoplasia. Both cyclic AMP responsive element binding protein (CREB)-dependent transcription and CPEB4-promoted translation support c-FOS expression early postnatal OBs but disengage in adult OBs. Activity-related c-FOS synthesis and GC survival are thus developmentally controlled by distinct molecular mechanisms to govern OB growth.


Asunto(s)
Gránulos Citoplasmáticos/metabolismo , Neuronas/metabolismo , Bulbo Olfatorio/metabolismo , Proteínas Proto-Oncogénicas c-fos/metabolismo , Proteínas de Unión al ARN/metabolismo , Animales , Ratones , Ratones Noqueados , Neurogénesis/fisiología , Bulbo Olfatorio/citología , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/genética
17.
Nat Commun ; 7: 10583, 2016 Feb 17.
Artículo en Inglés | MEDLINE | ID: mdl-26882873

RESUMEN

G-protein-coupled receptors (GPCRs) participate in a broad range of physiological functions. A priority for fundamental and clinical research, therefore, is to decipher the function of over 140 remaining orphan GPCRs. The suprachiasmatic nucleus (SCN), the brain's circadian pacemaker, governs daily rhythms in behaviour and physiology. Here we launch the SCN orphan GPCR project to (i) search for murine orphan GPCRs with enriched expression in the SCN, (ii) generate mutant animals deficient in candidate GPCRs, and (iii) analyse the impact on circadian rhythms. We thereby identify Gpr176 as an SCN-enriched orphan GPCR that sets the pace of circadian behaviour. Gpr176 is expressed in a circadian manner by SCN neurons, and molecular characterization reveals that it represses cAMP signalling in an agonist-independent manner. Gpr176 acts independently of, and in parallel to, the Vipr2 GPCR, not through the canonical Gi, but via the unique G-protein subclass Gz.


Asunto(s)
Ritmo Circadiano , Receptores Acoplados a Proteínas G/metabolismo , Animales , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas/metabolismo , Receptores Acoplados a Proteínas G/genética , Transducción de Señal , Núcleo Supraquiasmático/metabolismo
18.
Front Cell Neurosci ; 8: 367, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-25404896

RESUMEN

Cytoplasmic polyadenylation element binding protein 3 (CPEB3) is a sequence-specific RNA-binding protein that confines the strength of glutamatergic synapses by translationally downregulating the expression of multiple plasticity-related proteins (PRPs), including the N-methyl-D-aspartate receptor (NMDAR) and the postsynaptic density protein 95 (PSD95). CPEB3 knockout (KO) mice exhibit hippocampus-dependent abnormalities related not only to long-term spatial memory but also to the short-term acquisition and extinction of contextual fear memory. In this study, we identified a specific form of NMDAR-dependent synaptic depotentiation (DPT) that is impaired in the adult CPEB3 KO hippocampus. In parallel, cultured KO neurons also exhibited delayed morphological and biochemical responses under NMDA-induced chemical long-term depression (c-LTD). The c-LTD defects in the KO neurons include elevated activation of calcium/calmodulin-dependent protein kinase II alpha subunit (CaMKIIα), increased Ser831 phosphorylation of GluA1 and slow degradation of PSD95 and GluA1. Because transient pharmacological suppression of CaMKIIα activity during the DPT-initiating phase successfully reversed the LTP in the KO hippocampus, DPT and c-LTD in the two different systems shared common molecular defects due to the absence of CPEB3. Together, our results suggest that CPEB3 deficiency imbalances NMDAR-activated CaMKIIα signaling, which consequently fails to depress synaptic strength under certain stimulation conditions.

19.
J Neurosci ; 33(43): 17008-22, 2013 Oct 23.
Artículo en Inglés | MEDLINE | ID: mdl-24155305

RESUMEN

Long-term memory requires activity-dependent synthesis of plasticity-related proteins (PRPs) to strengthen synaptic efficacy and consequently consolidate memory. Cytoplasmic polyadenylation element binding protein (CPEB)3 is a sequence-specific RNA-binding protein that regulates translation of several PRP RNAs in neurons. To understand whether CPEB3 plays a part in learning and memory, we generated CPEB3 knock-out (KO) mice and found that the null mice exhibited enhanced hippocampus-dependent, short-term fear memory in the contextual fear conditioning test and long-term spatial memory in the Morris water maze. The basal synaptic transmission of Schaffer collateral-CA1 neurons was normal but long-term depression evoked by paired-pulse low-frequency stimulation was modestly facilitated in the juvenile KO mice. Molecular and cellular characterizations revealed several molecules in regulating plasticity of glutamatergic synapses are translationally elevated in the CPEB3 KO neurons, including the scaffolding protein PSD95 and the NMDA receptors along with the known CPEB3 target, GluA1. Together, CPEB3 functions as a negative regulator to confine the strength of glutamatergic synapses by downregulating the expression of multiple PRPs and plays a role underlying certain forms of hippocampus-dependent memories.


Asunto(s)
Guanilato-Quinasas/metabolismo , Hipocampo/fisiología , Proteínas de la Membrana/metabolismo , Memoria a Corto Plazo , Proteínas de Unión al ARN/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Animales , Condicionamiento Clásico , Homólogo 4 de la Proteína Discs Large , Miedo , Guanilato-Quinasas/genética , Hipocampo/citología , Hipocampo/metabolismo , Depresión Sináptica a Largo Plazo , Aprendizaje por Laberinto , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas/metabolismo , Neuronas/fisiología , Proteínas de Unión al ARN/genética , Receptores AMPA/genética , Receptores AMPA/metabolismo , Receptores de N-Metil-D-Aspartato/genética , Sinapsis/fisiología , Transmisión Sináptica
20.
Nucleic Acids Res ; 40(17): 8484-98, 2012 Sep 01.
Artículo en Inglés | MEDLINE | ID: mdl-22730302

RESUMEN

Cytoplasmic polyadenylation element-binding protein (CPEB)3 is a nucleocytoplasm-shuttling RNA-binding protein and predominantly resides in the cytoplasm where it represses target RNA translation. When translocated into the nucleus, CPEB3 binds to Stat5b and downregulates Stat5b-dependent transcription. In neurons, the activation of N-methyl-d-aspartate receptors (NMDARs) accumulates CPEB3 in the nucleus and redistributes CPEB3 in the nucleocytoplasmic compartments to control gene expression. Nonetheless, it is unclear which karyopherin drives the nuclear import of CPEB3 and which transport direction is most affected by NMDA stimulation to increase the nuclear pool of CPEB3. Here, we have identified that the karyopherins, IPO5 and CRM1, facilitate CPEB3 translocation by binding to RRM1 and a leucine-containing motif of CPEB3, respectively. NMDAR signaling increases RanBP1 expression and reduces the level of cytoplasmic GTP-bound Ran. These changes enhance CPEB3-IPO5 interaction, which consequently accelerates the nuclear import of CPEB3. This study uncovers a novel NMDA-regulated import pathway to facilitate the nuclear translocation of CPEB3.


Asunto(s)
Núcleo Celular/metabolismo , Proteínas de Unión al ARN/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , beta Carioferinas/metabolismo , Transporte Activo de Núcleo Celular , Animales , Células Cultivadas , Células HeLa , Humanos , Ratones , Neuronas/metabolismo , Señales de Exportación Nuclear , Señales de Localización Nuclear , Dominios y Motivos de Interacción de Proteínas , ARN Interferente Pequeño , Proteínas de Unión al ARN/química , Proteínas de Unión al ARN/genética , Ratas , Ribonucleósido Difosfato Reductasa , Transducción de Señal , Proteínas Supresoras de Tumor/metabolismo , beta Carioferinas/antagonistas & inhibidores , Proteína de Unión al GTP ran/metabolismo
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