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1.
Cell Rep ; 37(6): 109987, 2021 Nov 09.
Artigo em Inglês | MEDLINE | ID: mdl-34758320

RESUMO

CENP-A (centromeric protein A), a histone H3 variant, specifies centromere identity and is essential to centromere maintenance. Little is known about how protein levels of CENP-A are controlled in mammalian cells. Here, we report that the phosphorylation of CENP-A Ser68 primes the ubiquitin-proteasome-mediated proteolysis of CENP-A during mitotic phase in human cultured cells. We identify two major polyubiquitination sites that are responsible for this phosphorylation-dependent degradation. Substituting the two residues, Lys49 and Lys124, with arginines abrogates proper CENP-A degradation and results in CENP-A mislocalization to non-centromeric regions. Furthermore, we find that DCAF11 (DDB1 and CUL4 associated factor 11/WDR23) is the E3 ligase that specifically mediates the observed polyubiquitination. Deletion of DCAF11 hampers CENP-A degradation and causes its mislocalization. We conclude that the Ser68 phosphorylation plays an important role in regulating cellular CENP-A homeostasis via DCAF11-mediated degradation to prevent ectopic localization of CENP-A during the cell cycle.

2.
Cell Prolif ; 54(11): e13133, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34585448

RESUMO

OBJECTIVES: Maternal factors that are enriched in oocytes have attracted great interest as possible key factors in somatic cell reprogramming. We found that surfeit locus protein 4 (Surf4), a maternal factor, can facilitate the generation of induced pluripotent stem cells (iPSCs) previously, but the mechanism remains elusive. MATERIALS AND METHODS: In this study, we investigated the function and mechanism of Surf4 in somatic cell reprogramming using a secondary reprogramming system. Alkaline phosphatase (AP) staining, qPCR and immunofluorescence (IF) staining of expression of related markers were used to evaluate efficiency of iPSCs derived from mouse embryonic fibroblasts. Embryoid body and teratoma formation assays were performed to evaluate the differentiation ability of the iPSC lines. RNA-seq, qPCR and western blot analysis were applied to validate the downstream targets of Surf4. RESULTS: Surf4 can significantly facilitate the generation of iPSCs in a proliferation-independent manner. When co-expressed with Oct4, Sox2, Klf4 and c-Myc (OSKM), Surf4 can activate the response to endoplasmic reticulum (ER) stress at the early stage of reprogramming. We further demonstrated that Hspa5, a major ER chaperone, and the active spliced form of Xbp1 (sXbp1), a major mediator of ER stress, can mimic the effects of Surf4 on somatic cell reprogramming. Concordantly, blocking the unfolded protein response compromises the effect of Surf4 on reprogramming. CONCLUSIONS: Surf4 promotes somatic cell reprogramming by activating the response to ER stress.


Assuntos
Reprogramação Celular/fisiologia , Estresse do Retículo Endoplasmático/fisiologia , Fibroblastos/metabolismo , Proteínas de Membrana/metabolismo , Animais , Diferenciação Celular/fisiologia , Corpos Embrioides/metabolismo , Células-Tronco Pluripotentes Induzidas/citologia , Camundongos , Fatores de Transcrição/metabolismo
3.
Br J Ophthalmol ; 2021 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-34385166

RESUMO

BACKGROUND/AIMS: Pathological myopia (PM) is a leading cause of blindness worldwide. We aimed to evaluate microvascular and chorioretinal changes in different stages of myopia with wide-field (WF) swept-source (SS) optical coherence tomography angiography (OCTA). METHODS: This prospective cross-sectional observational study included 186 eyes of 122 patients who had undergone imaging between November 2018 and October 2020. Vessel density (VD) and vessel skeletonised density (VSD) of superficial capillary plexus, deep capillary plexus and whole retina, as well as foveal avascular zone parameters, retinal thickness (RT) and choroidal thickness (CT), were calculated. RESULTS: This study evaluated 75 eyes of 48 patients with high myopia (HM), 43 eyes of 31 patients with mild to moderate myopia and 68 eyes of 53 age-matched controls. Controlling for age and the presence of systemic hypertension, we found that HM was associated with decrease in VD and VSD in all layers on 12×12 mm² scans. Furthermore, HM was associated with a VD and VSD decrease in every Early Treatment Diabetic Retinopathy Study grid, with a larger decrease temporally (ßVD=-0.39, ßVSD=-10.25, p<0.01). HM was associated with decreased RT and CT. Reduction in RT was outside the macular region, while reduction in CT was in the macular region. CONCLUSION: Using WF SS-OCTA, we identified reduction in microvasculature and structural changes associated with myopia. Decrease in VD and VSD was greater in the temporal quadrant, and reductions in RT and CT were uneven across the retina. Further work may help identify risk factors for the progression of PM and associated vision-threatening complications.

5.
Acta Ophthalmol ; 2021 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-33880864

RESUMO

PURPOSE: To report the efficacy and safety of bleb-independent penetrating canaloplasty in the management of primary angle-closure glaucoma (PACG). METHODS: This single-centre prospective interventional case series enrolled 57 eyes from 53 PACG patients with medically uncontrolled intraocular pressure (IOP) and peripheral anterior synechiae of over 270°. Penetrating canaloplasty, mainly consisted of tensioning suture-aided Schlemm's canal dilation and a trabeculectomy, was performed to create a direct communication between the anterior chamber and the Schlemm's canal. Postoperative IOP, number of glaucoma medications and procedure-related complications were evaluated. Rate of success was defined as IOP ≤ 21, ≤18 and ≤15 mmHg, and a ≥30% IOP reduction without (complete) or with/without (qualified) IOP-lowering medications. RESULTS: A total of 45 eyes had 360° catheterization successfully completed. The mean preoperative IOP was 33.9 ± 11.7 mmHg (range, 13-59.6 mmHg), on 3.2 ± 0.8 glaucoma medications (range 2-5), which was decreased to 15.4 ± 3.7 mmHg (range, 8.6-22.5) and 0.2 ± 0.6 (range, 0-3) medications at 6 months and 14.8 ± 3.5 mmHg (range, 9-24) and 0.1 ± 0.3 (range, 0-1) medications at 12 months postoperatively. Complete success at 12 months were achieved in 78.9% (95% CI: 0.65-0.93), 71.1% (0.56-0.86) and 50.0% (0.33-0.67) eyes at IOP ≤ 21, ≤18 and ≤15 mmHg, respectively. Transient IOP elevation (>30 mmHg, 26.7%) and hyphema (11.1%) were the most common early surgical complications. CONCLUSION: Penetrating canaloplasty in PACG appeared to have good efficacy and safety profiles in this pilot study. Further studies are justified.

6.
Protein Cell ; 12(6): 455-474, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33886094

RESUMO

N6-methyladenosine (m6A) on chromosome-associated regulatory RNAs (carRNAs), including repeat RNAs, plays important roles in tuning the chromatin state and transcription, but the intrinsic mechanism remains unclear. Here, we report that YTHDC1 plays indispensable roles in the self-renewal and differentiation potency of mouse embryonic stem cells (ESCs), which highly depends on the m6A-binding ability. Ythdc1 is required for sufficient rRNA synthesis and repression of the 2-cell (2C) transcriptional program in ESCs, which recapitulates the transcriptome regulation by the LINE1 scaffold. Detailed analyses revealed that YTHDC1 recognizes m6A on LINE1 RNAs in the nucleus and regulates the formation of the LINE1-NCL partnership and the chromatin recruitment of KAP1. Moreover, the establishment of H3K9me3 on 2C-related retrotransposons is interrupted in Ythdc1-depleted ESCs and inner cell mass (ICM) cells, which consequently increases the transcriptional activities. Our study reveals a role of m6A in regulating the RNA scaffold, providing a new model for the RNA-chromatin cross-talk.

7.
Ophthalmology ; 128(9): 1312-1324, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-33647282

RESUMO

PURPOSE: To investigate the association among widefield swept-source (SS) OCT angiography (OCTA) metrics and systemic parameters and vitreous hemorrhage (VH) occurrence in eyes with proliferative diabetic retinopathy (PDR). DESIGN: Prospective, observational study. PARTICIPANTS: Fifty-five eyes from 45 adults with PDR, with no history of VH, followed up for at least 3 months. METHODS: All patients underwent widefield SS OCTA (Montage 15 × 15 mm and high-definition (HD)-51 line scan) imaging. Images were evaluated independently by 2 graders for quantitative and qualitative widefield SS OCTA metrics defined a priori. Systemic and ocular parameters and widefield SS OCTA metrics were screened using least absolute shrinkage and selection operator and logistic or Cox regression for variable selection. Firth's bias-reduced logistic regression models (outcome, occurrence of VH) and Cox regression models (outcome, time to occurrence of VH) were used to identify parameters associated with VH occurrence. MAIN OUTCOME MEASURES: Occurrence of VH. RESULTS: Over a median follow-up of 363 days (range, 28-710 days), 13 of 55 PDR eyes (24%) demonstrated VH during the follow-up period. Presence of extensive neovascularizations (odds ratio, 8.05; 95% confidence interval [CI], 1.43-58.56; P = 0.02), defined as neovascularizations with total area of more than 4 disc diameters, and forward neovascularizations (odds ratio, 5.42; 95% CI, 1.26-35.16; P = 0.02) that traversed the posterior hyaloid face into the vitreous were associated with the occurrence of VH. The presence of flat neovascularizations (odds ratio, 0.25; 95% CI, 0.04-1.01; P = 0.05) confined to the posterior hyaloid face was associated with a lower risk of VH with borderline significance. Similarly, presence of extensive neovascularizations (hazard ratio, 18.24; 95% CI, 3.51-119.47; P < 0.001) and forward neovascularizations (hazard ratio, 9.60; 95% CI, 2.07-68.08; P = 0.002) was associated significantly with time to development of VH. CONCLUSIONS: Widefield SS OCTA is useful for evaluating neovascularizations and their relationship with the vitreous. The presence of forward and extensive neovascularizations was associated with the occurrence of VH in patients with PDR. Larger samples and longer follow-up are needed to verify the risk factors and imaging biomarkers for diabetic VH.


Assuntos
Retinopatia Diabética/diagnóstico , Angiofluoresceinografia , Tomografia de Coerência Óptica , Hemorragia Vítrea/diagnóstico , Adulto , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Neovascularização Retiniana/diagnóstico , Fatores de Tempo , Acuidade Visual/fisiologia
8.
Stem Cell Reports ; 16(3): 458-469, 2021 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-33636112

RESUMO

A small subgroup of embryonic stem cells (ESCs) exhibit molecular features similar to those of two-cell embryos (2C). However, it remains elusive whether 2C-like cells and 2C embryos share similar epigenetic features. Here, we map the genome-wide profiles of histone H3K4me3 and H3K27me3 in 2C-like cells. We found that the majority of genes in 2C-like cells inherit their histone status from ESCs. Among the genes showing a switch in their histone methylation status during 2C-like transitions, only a small number acquire 2C-embryo epigenetic signatures. In contrast, broad H3K4me3 domains display extensive loss in 2C-like cells. Most of the differentially expressed genes display decreased H3K4me3 and H3K27me3 levels in 2C-like cells, whereas de novo H3K4me3 deposition is closely linked with the expression levels of upregulated 2C-specific genes. Taken together, our study reveals the unique epigenetic profiles of 2C-like cells, facilitating the further exploration of totipotency in the future.

9.
Br J Ophthalmol ; 105(4): 577-581, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-32591347

RESUMO

AIMS: To compare widefield swept-source optical coherence tomography angiography (WF SS-OCTA) with ultra-widefield colour fundus photography (UWF CFP) and fluorescein angiography (UWF FA) for detecting diabetic retinopathy (DR) lesions. METHODS: This prospective, observational study was conducted at Massachusetts Eye and Ear from December 2018 to October 2019. Proliferative DR, non-proliferative DR and diabetic patients with no DR were included. All patients were imaged with a WF SS-OCTA using a Montage 15×15 mm scan. UWF CFP and UWF FA were taken by a 200°, single capture retinal imaging system. Images were independently evaluated for the presence or absence of DR lesions including microaneurysms (MAs), intraretinal microvascular abnormalities (IRMAs), neovascularisation elsewhere (NVE), neovascularisation of the optic disc (NVD) and non-perfusion areas (NPAs). All statistical analyses were performed using SPSS V.25.0. RESULTS: One hundred and fifty-two eyes of 101 participants were included in the study. When compared with UWF CFP, WF SS-OCTA was found to be superior in detecting IRMAs (p<0.001) and NVE/NVD (p=0.007). The detection rates of MAs, IRMAs, NVE/NVD and NPAs in WF SS-OCTA were comparable with UWF FA images (p>0.05). Furthermore, when we compared WF SS-OCTA plus UWF CFP with UWF FA, the detection rates of MAs, IRMAs, NVE/NVD and NPAs were identical (p>0.005). Agreement (κ=0.916) between OCTA and FA in classifying DR was excellent. CONCLUSION: WF SS-OCTA is useful for identification of DR lesions. WF SS-OCTA plus UWF CFP may offer a less invasive alternative to FA for DR diagnosis.


Assuntos
Retinopatia Diabética/diagnóstico , Angiofluoresceinografia/métodos , Disco Óptico/patologia , Vasos Retinianos/patologia , Tomografia de Coerência Óptica/métodos , Acuidade Visual , Feminino , Fundo de Olho , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos
10.
Cell Stem Cell ; 28(4): 732-747.e9, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33357405

RESUMO

Telomeres play vital roles in ensuring chromosome stability and are thus closely linked with the onset of aging and human disease. Telomeres undergo extensive lengthening during early embryogenesis. However, the detailed molecular mechanism of telomere resetting in early embryos remains unknown. Here, we show that Dcaf11 (Ddb1- and Cul4-associated factor 11) participates in telomere elongation in early embryos and 2-cell-like embryonic stem cells (ESCs). The deletion of Dcaf11 in embryos and ESCs leads to reduced telomere sister-chromatid exchange (T-SCE) and impairs telomere lengthening. Importantly, Dcaf11-deficient mice exhibit gradual telomere erosion with successive generations, and hematopoietic stem cell (HSC) activity is also greatly compromised. Mechanistically, Dcaf11 targets Kap1 (KRAB-associated protein 1) for ubiquitination-mediated degradation, leading to the activation of Zscan4 downstream enhancer and the removal of heterochromatic H3K9me3 at telomere/subtelomere regions. Our study therefore demonstrates that Dcaf11 plays important roles in telomere elongation in early embryos and ESCs through activating Zscan4.


Assuntos
Homeostase do Telômero , Telômero , Animais , Células-Tronco Embrionárias , Camundongos
11.
Cell Death Dis ; 11(12): 1040, 2020 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-33288747

RESUMO

Diabetic nephropathy (DN) as a global health concern is closely related to inflammation and oxidation. Isoliquiritigenin (ISL), a natural flavonoid compound, has been demonstrated to inhibit inflammation in macrophages. Herein, we investigated the effect of ISL in protecting against the injury in STZ-induced type 1 DN and in high glucose-induced NRK-52E cells. In this study, it was revealed that the administration of ISL not only ameliorated renal fibrosis and apoptosis, but also induced the deterioration of renal function in diabetic mice. Mediated by MAPKs and Nrf-2 signaling pathways, respectively, upstream inflammatory response and oxidative stress were neutralized by ISL in vitro and in vivo. Moreover, as further revealed by the results of molecular docking, sirtuin 1 (SIRT1) binds to ISL directly, and the involvement of SIRT1 in ISL-mediated renoprotective effects was confirmed by studies using in vitro models of SIRT1 overexpression and knockdown. In summary, by reducing inflammation and oxidative stress, ISL has a significant pharmacological effect on the deterioration of DN. The benefits of ISL are associated with the direct binding to SIRT1, the inhibition of MAPK activation, and the induction of Nrf-2 signaling, suggesting the potential of ISL for DN treatment.


Assuntos
Chalconas/uso terapêutico , Hiperglicemia/tratamento farmacológico , Inflamação/complicações , Rim/lesões , Estresse Oxidativo , Sirtuína 1/metabolismo , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Chalconas/química , Chalconas/farmacologia , Fibrose , Rim/efeitos dos fármacos , Rim/patologia , Camundongos Endogâmicos C57BL , Modelos Moleculares , Oxirredução , Estresse Oxidativo/efeitos dos fármacos , Ratos
12.
Br J Ophthalmol ; 2020 Dec 21.
Artigo em Inglês | MEDLINE | ID: mdl-33355148

RESUMO

AIMS: To compare the efficacy of diabetic retinal neovascularisation (NV) detection using the widefield swept-source optical coherence tomography angiography (WF SS-OCTA) vitreoretinal interface (VRI) Angio slab and SS-OCT VRI Structure slab. METHODS: A prospective, observational study was performed at Massachusetts Eye and Ear from January 2019 to June 2020. Patients with proliferative diabetic retinopathy (PDR), patients with non-proliferative diabetic retinopathy and patients with diabetes but without diabetic retinopathy were included. All patients were imaged with WF SS-OCTA using the 12×12 mm Angio scan protocol centred on the fovea and optic disc. The en-face SS-OCTA VRI Angio slab and SS-OCT VRI Structure slab were evaluated for the presence or absence of NV. SS-OCTA B-scan was used to classify NV according to cross-sectional morphology (forward, tabletop or flat). All statistical analyses were performed using SPSS V.26.0. RESULTS: One hundred and forty-two eyes of 89 participants were included in the study. VRI Angio detected NV at higher rates compared with VRI Structure (p<0.05). Combining VRI Angio and Structure improved detection rates compared with VRI Angio alone (p<0.05). Due to segmentation errors of the internal limiting membrane, NV with flat morphological classification had lower rates of detection on VRI Angio compared with NV with forward and tabletop morphology (p<0.05). CONCLUSIONS: WF SS-OCTA 12×12 mm VRI Angio and SS-OCT VRI Structure imaging centred on the fovea and optic disc detected NV with high sensitivity and low false positives. The VRI slab may be useful to diagnose and monitor PDR in clinical practice.

13.
Exp Eye Res ; 199: 108146, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32726604

RESUMO

PURPOSE: To examine the protective effects of Isoliquiritigenin (ISL) in angiotensin II (ANG II)-induced inflammation and fibrosis on Human Tenon's capsule Fibroblasts (HTFs) and Mouse Peritoneal Macrophages (MPMs). This study also investigated the potential mechanism of action of ISL. METHOD: Methyl-thiazolyl tetrazolium (MTT) assay was used to test ISL toxicity. An ELISA and an RT-qPCR assay detected the inflammatory cytokines (TNF-α, IL-6, COX-2, and ICAM-1). A Western blot investigated the expression levels of inflammation-related signals [nuclear factor-κB (NF-κB), peroxisome proliferator-activated receptor γ (PPARγ)], and fibrogenesis, including fibronectin and alpha-smooth muscle actin (α-SMA)]. Protein expressions of α-SMA were measured by immunofluorescence. RESULTS: Pre-treatment with ISL (10 or 20 µM) dose-dependently decreased the mRNA levels of TNF-α, IL-6, ICAM-1, and COX-2 induced by ANG II (1 µg/ml) in both MPMs and HTFs. ANG II remarkably increased the amount of P65 in the nuclei and decreased the amount of P65 in the cytoplasm. Additionally, ANG II reduced PPARγ expression levels in a time-dependent manner. Furthermore, these effects which were induced by ISL were remarkably neutralized by ISL pre-treatment. Finally, ANG II markedly elevated the expression of fibronectin and α-SMA. CONCLUSION: ISL could alleviate ANG II-induced fibrogenesis by inhibiting the NF-κB/PPARγ inflammatory pathway. In addition, ISL may be a potential agent for the treatment of conjunctival fibrosis. Most importantly, the NF-κB/PPARγ signaling pathway could be an effective therapeutic target for the prevention and treatment of conjunctival fibrosis after glaucoma surgery.


Assuntos
Angiotensina II/efeitos adversos , Chalconas/farmacologia , Conjuntivite/prevenção & controle , NF-kappa B/genética , PPAR gama/genética , Cápsula de Tenon/metabolismo , Aldeído Redutase , Animais , Western Blotting , Células Cultivadas , Túnica Conjuntiva/efeitos dos fármacos , Túnica Conjuntiva/metabolismo , Túnica Conjuntiva/patologia , Conjuntivite/metabolismo , Conjuntivite/patologia , Inibidores Enzimáticos/farmacologia , Ensaio de Imunoadsorção Enzimática , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , NF-kappa B/metabolismo , PPAR gama/metabolismo , RNA Mensageiro/genética , Transdução de Sinais/efeitos dos fármacos , Cápsula de Tenon/efeitos dos fármacos , Cápsula de Tenon/patologia , Trabeculectomia/efeitos adversos , Vasoconstritores/efeitos adversos
14.
Ophthalmic Surg Lasers Imaging Retina ; 51(7): 407-412, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32706899

RESUMO

Herein, the authors describe an initial case report of widefield swept-source optical coherence tomography angiography (SS-OCTA) in Vogt-Koyanagi-Harada (VKH) disease. When compared to fluorescent angiography, indocyanine green angiography, and enhanced-depth OCT - upon which the revised criteria for VHK are based - widefield SS-OCTA enables detection of vitreous inflammation, noninvasive identification of characteristic areas of flow void at the level of choriocapillaris in the acute phase and may be a novel valuable tool not only for noninvasive diagnosis and monitoring of disease progression, persistence, resolution, and recurrence to guide therapy in VKH disease in the future. [Ophthalmic Surg Lasers Imaging Retina. 2020;51:407-412.].


Assuntos
Corioide/patologia , Angiofluoresceinografia/métodos , Tomografia de Coerência Óptica/métodos , Síndrome Uveomeningoencefálica/diagnóstico , Adulto , Feminino , Fundo de Olho , Humanos
15.
J Genet Genomics ; 47(3): 123-130, 2020 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-32305172

RESUMO

Embryonic stem cells possess fascinating capacity of self-renewal and developmental potential, leading to significant progress in understanding the molecular basis of pluripotency, disease modeling, and reprogramming technology. Recently, 2-cell-like embryonic stem cells (ESCs) and expanded potential stem cells or extended pluripotent stem cells (EPSCs) generated from early-cleavage embryos display some features of totipotent embryos. These cell lines provide valuable in vitro models to study underlying principles of totipotency, cell plasticity, and lineage segregation. In this review, we summarize the current progress in this filed and highlight the application potentials of these cells in the future.


Assuntos
Diferenciação Celular/genética , Reprogramação Celular/genética , Células-Tronco Embrionárias/citologia , Células-Tronco Totipotentes/citologia , Linhagem da Célula/genética , Plasticidade Celular/genética , Autorrenovação Celular/genética , Humanos
16.
EMBO J ; 37(18)2018 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-30111536

RESUMO

Mammalian oocytes and zygotes have the unique ability to reprogram a somatic cell nucleus into a totipotent state. SUV39H1/2-mediated histone H3 lysine-9 trimethylation (H3K9me3) is a major barrier to efficient reprogramming. How SUV39H1/2 activities are regulated in early embryos and during generation of induced pluripotent stem cells (iPSCs) remains unclear. Since expression of the CRL4 E3 ubiquitin ligase in oocytes is crucial for female fertility, we analyzed putative CRL4 adaptors (DCAFs) and identified DCAF13 as a novel CRL4 adaptor that is essential for preimplantation embryonic development. Dcaf13 is expressed from eight-cell to morula stages in both murine and human embryos, and Dcaf13 knockout in mice causes preimplantation-stage mortality. Dcaf13 knockout embryos are arrested at the eight- to sixteen-cell stage before compaction, and this arrest is accompanied by high levels of H3K9me3. Mechanistically, CRL4-DCAF13 targets SUV39H1 for polyubiquitination and proteasomal degradation and therefore facilitates H3K9me3 removal and zygotic gene expression. Taken together, CRL4-DCAF13-mediated SUV39H1 degradation is an essential step for progressive genome reprogramming during preimplantation embryonic development.


Assuntos
Blastocisto/metabolismo , Desenvolvimento Embrionário , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Pluripotentes Induzidas/metabolismo , Metiltransferases/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas Repressoras/metabolismo , Animais , Blastocisto/citologia , Estabilidade Enzimática , Histonas/genética , Histonas/metabolismo , Células-Tronco Pluripotentes Induzidas/citologia , Metiltransferases/genética , Camundongos , Camundongos Endogâmicos ICR , Camundongos Knockout , Oócitos/citologia , Oócitos/metabolismo , Proteólise , Proteínas de Ligação a RNA/genética , Proteínas Repressoras/genética , Complexos Ubiquitina-Proteína Ligase/genética , Complexos Ubiquitina-Proteína Ligase/metabolismo , Ubiquitinação/genética
17.
Nat Cell Biol ; 20(5): 620-631, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29686265

RESUMO

H3K9me3-dependent heterochromatin is a major barrier of cell fate changes that must be reprogrammed after fertilization. However, the molecular details of these events are lacking in early embryos. Here, we map the genome-wide distribution of H3K9me3 modifications in mouse early embryos. We find that H3K9me3 exhibits distinct dynamic features in promoters and long terminal repeats (LTRs). Both parental genomes undergo large-scale H3K9me3 reestablishment after fertilization, and the imbalance in parental H3K9me3 signals lasts until blastocyst. The rebuilding of H3K9me3 on LTRs is involved in silencing their active transcription triggered by DNA demethylation. We identify that Chaf1a is essential for the establishment of H3K9me3 on LTRs and subsequent transcriptional repression. Finally, we find that lineage-specific H3K9me3 is established in post-implantation embryos. In summary, our data demonstrate that H3K9me3-dependent heterochromatin undergoes dramatic reprogramming during early embryonic development and provide valuable resources for further exploration of the epigenetic mechanism in early embryos.


Assuntos
Blastocisto/metabolismo , Reprogramação Celular , Montagem e Desmontagem da Cromatina , Metilação de DNA , Epigênese Genética , Heterocromatina/metabolismo , Histonas/metabolismo , Animais , Linhagem Celular , Fator 1 de Modelagem da Cromatina/genética , Fator 1 de Modelagem da Cromatina/metabolismo , Técnicas de Cultura Embrionária , Implantação do Embrião , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Idade Gestacional , Heterocromatina/genética , Histonas/genética , Masculino , Metilação , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco Embrionárias Murinas/metabolismo , Regiões Promotoras Genéticas , Sequências Repetidas Terminais
18.
Stem Cell Reports ; 10(2): 477-493, 2018 02 13.
Artigo em Inglês | MEDLINE | ID: mdl-29396184

RESUMO

Androgenetic haploid embryonic stem cells (AG-haESCs) hold great promise for exploring gene functions and generating gene-edited semi-cloned (SC) mice. However, the high incidence of self-diploidization and low efficiency of SC mouse production are major obstacles preventing widespread use of these cells. Moreover, although SC mice generation could be greatly improved by knocking out the differentially methylated regions of two imprinted genes, 50% of the SC mice did not survive into adulthood. Here, we found that the genome-wide DNA methylation level in AG-haESCs is extremely low. Subsequently, downregulation of both de novo methyltransferase Dnmt3b and other methylation-related genes was determined to be responsible for DNA hypomethylation. We further demonstrated that ectopic expression of Dnmt3b in AG-haESCs could effectively improve DNA methylation level, and the high incidence of self-diploidization could be markedly rescued. More importantly, the developmental potential of SC embryos was improved, and most SC mice could survive into adulthood.


Assuntos
DNA (Citosina-5-)-Metiltransferases/genética , Metilação de DNA/genética , Diploide , Células-Tronco Embrionárias Murinas/citologia , Animais , Sobrevivência Celular/genética , Clonagem de Organismos , Feminino , Edição de Genes , Regulação da Expressão Gênica no Desenvolvimento , Haploidia , Camundongos , Camundongos Knockout
19.
Biochem Biophys Res Commun ; 496(2): 245-252, 2018 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-29180018

RESUMO

Sepsis, one of the most fatal diseases worldwide, often leads to multiple organ failure, mainly due to uncontrolled inflammatory responses. Despite accumulating knowledge obtained in recent years, effective drugs to treat sepsis in the clinic are still urgently needed. Isoliquiritigenin (ISL), a chalcone compound, has been reported to exert anti-inflammatory properties. However, little is known about the effects of ISL on sepsis and its related complications. In this study, we investigated the potential protective effects of ISL on lipopolysaccharide (LPS)-induced injuries and identified the mechanisms underlying these effects. ISL inhibited inflammatory cytokine expression in mouse primary peritoneal macrophages (MPMs) exposed to LPS. In an acute lung injury (ALI) mouse model, ISL prevented LPS-induced structural damage and inflammatory cell infiltration. Additionally, pretreatment with ISL attenuated sepsis-induced lung and liver injury, accompanied by a reduction in inflammatory responses. Moreover, these protective effects were mediated by the nuclear factor kappa B (NF-κB) pathway-mediated inhibition of inflammatory responses in vitro and in vivo. Our study suggests that ISL may be a potential therapeutic agent for sepsis-induced injuries.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Anti-Inflamatórios/farmacologia , Chalconas/farmacologia , Pulmão/efeitos dos fármacos , Sepse/tratamento farmacológico , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/imunologia , Lesão Pulmonar Aguda/patologia , Animais , Citocinas/genética , Citocinas/imunologia , Modelos Animais de Doenças , Regulação da Expressão Gênica , Células Hep G2 , Humanos , Lipopolissacarídeos , Fígado/efeitos dos fármacos , Fígado/imunologia , Fígado/patologia , Pulmão/imunologia , Pulmão/patologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/imunologia , Macrófagos Peritoneais/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/genética , NF-kappa B/imunologia , Cultura Primária de Células , Sepse/induzido quimicamente , Sepse/imunologia , Sepse/patologia
20.
Oncotarget ; 8(48): 84276-84284, 2017 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-29137422

RESUMO

Aerobic glycolysis is one of the most important common characteristics in both cancer cells and stem cells. Metabolism switch has been discovered as an important early event in the process of reprogramming somatic cells to induced pluripotent stem cells (iPSCs). As a rate limiting kinase in glycolysis, Pkm2 has been reported playing critical roles in many tumors, yet its role in stem cells and iPSCs induction is poorly defined. In the present study, we showed that Pkm2 is a predominant pyruvate kinase in embryonic stem cells (ESCs), and its expression increases many pluripotent genes. During somatic cell reprogramming, up-regulation of Pkm2 can be observed and over-expression of Pkm2 can facilitate iPSCs induction, while Pkm1 or a mutant form of Pkm2 (Pkm2K422R) showed no enhancement role in iPSCs induction. Therefore, our data demonstrated that Pkm2 enhances the pluripotency maintenance in ESCs and promotes the pluripotency acquisition during somatic cell reprogramming.

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