scholarly journals Single-Cell RNA Sequencing Reveals Regulatory Mechanism for Trophoblast Cell-Fate Divergence in Human Peri-Implantation Embryo

2019 ◽  
Author(s):  
Bo Lv ◽  
Qin An ◽  
Qiao Zeng ◽  
Ping Lu ◽  
Xianmin Zhu ◽  
...  
Keyword(s):  
Cell Fate ◽  
Cellular Differentiation ◽  
Regulatory Mechanism ◽  
Bioinformatic Analysis ◽  
Loss Of Function ◽  
Trophoblast Differentiation ◽  
T Box ◽  
Development And Differentiation ◽  
Post Implantation

AbstractMultipotent trophoblasts undergo dynamic morphological movement and cellular differentiation after embryonic implantation to generate placenta. However, the mechanism controlling trophoblast development and differentiation during peri-implantation development remains elusive. In this study, we modeled human embryo peri-implantation development from blastocyst to early post-implantation stages by using an in vitro coculture system, and profiled the transcriptome of individual trophoblast cells from these embryos. We revealed the genetic networks regulating peri-implantation trophoblast development. While determining when trophoblast differentiation happens, our bioinformatic analysis identified T-box transcription factor 3 (TBX3) as a key regulator for the differentiation of cytotrophoblast into syncytiotrophoblast. The function of TBX3 in trophoblast differentiation is then validated by a loss-of-function experiment. In conclusion, our results provided a valuable resource to study the regulation of trophoblasts development and differentiation during human peri-implantation development.

scholarly journals LukS-PV-Regulated MicroRNA-125a-3p Promotes THP-1 Macrophages Differentiation and Apoptosis by Down-Regulating NF1 and Bcl-2

2017 ◽  
Vol 44 (3) ◽  
pp. 1093-1105 ◽  
Author(s):  
Xiao-Xi Sun ◽  
Shan-Shan Zhang ◽  
Chun-Yang Dai ◽  
Jing Peng ◽  
Qing Pan ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Cellular Differentiation ◽  
Target Genes ◽  
B Cell Lymphoma ◽  
Neurofibromatosis Type ◽  
Luciferase Reporter ◽  
Loss Of Function ◽  
Leukaemia Cell Line ◽  
Reporter Assays

Background/Aims: LukS-PV is a component of Panton-Valentine leukocidin (PVL). We have previously demonstrated that LukS-PV potently promoted differentiation and induced apoptosis in THP-1 cells. However, the precise mechanisms of these actions remain unknown. MicroRNAs (miRs) play important roles in cellular differentiation and apoptosis. This study aimed to investigate the role of miR-125a-3p in LukS-PV-regulated differentiation and apoptosis and its underlying mechanism in THP-1 cells. Methods: MicroRNA profiling analyses were conducted to determine differential miRNA expression levels in THP-1 cells treated with LukS-PV. Cell differentiation and apoptosis were measured in THP-1 cells by gain-of-function and loss-of-function experiments. Bioinformatics analysis and luciferase reporter assays were used to confirm the targets of miR-125a-3p. The effects of the miR-125a-3p targets on cellular differentiation were determined by knocking them down. Results: MiR-125a-3p was up-regulated after treating the human monocytic leukaemia cell line THP-1 with LukS-PV. In addition, miR-125a-3p positively regulated apoptosis and differentiation in THP-1 cells treated with LukS-PV. Concordantly, luciferase reporter assays confirmed that neurofibromatosis type 1 (NF1) and B-cell lymphoma 2 (Bcl-2) were direct target genes of miR-125a-3p. Moreover, NF1 knockdown in THP-1 cells significantly promoted differentiation in vitro. Finally, the extracellular signal-regulated kinase (ERK) pathway, a downstream target of NF1, was activated after NF1 knockdown. Conclusions: These findings confirm that miR-125a-3p is involved in LukS-PV-mediated cell differentiation and apoptosis in THP-1 cells.

The SCL Transcription Factor Supports Erythroid Cell Survival and Differentiation Downstream of the Erythropoietin Receptor.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 818-818
Author(s):  
Rachid Lahlil ◽  
Richard Martin ◽  
Peter D. Aplan ◽  
C. Glenn Begley ◽  
Jacqueline E. Damen ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Cell Survival ◽  
Cell Fate ◽  
Genetic Interaction ◽  
Fetal Liver ◽  
Erythropoietin Receptor ◽  
Erythroid Cell ◽  
Loss Of Function

Abstract Erythroid cell development critically depends on the SCL/Tal1 transcription factor and on erythropoietin signalling. In the present study, we have taken several approaches to show that the two genes operate within the same pathway to consolidate the erythroid lineage. Signaling through the erythropoietin receptor (EpoR) upregulates SCL protein levels in a clonal cell line (TF-1) in vitro, and in murine fetal liver cells in vivo, when Epor−/− cells were compared to those of wild type littermates at E12.5. In addition, we provide functional evidence for a linear pathway from EpoR to SCL that regulates erythropoiesis. Interfering with SCL induction or SCL function prevents the anti-apoptotic effect of Epo in TF-1 cells and conversely, ectopic SCL expression is sufficient to substitute for Epo to transiently maintain cell survival. In vivo, SCL gain of function complements the cellular defects in Epor−/− embryos to support cell survival and maturation during primitive and definitive erythropoiesis, as assessed by cellular and histological analyses of Epor−/− SCLtg embryos. Moreover, several erythroid specific genes that are decreased in Epor−/− embryos are rescued by the SCL transgene including glycophorinA, bH1 and bmaj globin, providing molecular confirmation of the functional and genetic interaction between Epor and SCL. Conversely, erythropoiesis becomes deficient in compound Epor+/−SCL+/− heterozygote mice, indicating that the genetic interaction between EpoR and SCL is synthetic. Finally, using EpoR mutants that harbour well defined signalling deficiencies, combined with gain and loss of function approaches for specific kinases, we identify MAPK as the major signal transduction pathway downstream of EpoR that upregulates SCL function, necessary for erythroid cell survival and differentiation. Taken together, our observations are consistent with the view that cytokines can influence cell fate by altering the dosage of lineage transcriptional regulators.

scholarly journals Yorkie and JNK revert syncytial muscles into myoblasts during Org-1 dependent lineage reprogramming

2019 ◽  
Author(s):  
Christoph Schaub ◽  
Marcel Rose ◽  
Manfred Frasch
Keyword(s):  
Cell Fate ◽  
Present Report ◽  
Cellular Reprogramming ◽  
List Type ◽  
T Box ◽  
Lineage Reprogramming ◽  
Kinase Cascade ◽  
Alary Muscles

SummaryLineage reprogramming has become a prominent focus in research since it was demonstrated that lineage restricted transcription factors can be used in vitro for direct reprogramming [1]. Recently, we reported that the ventral longitudinal musculature (VLM) of the adult Drosophila heart arises in vivo by direct lineage reprogramming from alary muscles (AM), a process which starts with dedifferentiation and fragmentation of syncytial alary muscles into mononucleate myoblasts. Central upstream activators of the genetic program regulating the development of VLMs from alary muscles are the T-box factor Org-1 (Drosophila Tbx1) and the LIM homeodomain factor Tup (Drosophila Islet1) [2]. However, the events downstream of Org-1 and Tup that exert dedifferentiation and fragmentation of alary muscles have been unknown. In the present report, we shed light on the initiation of this first step of transdifferentiation and show that AM lineage specific activation of Yorkie (Yki), the transcriptional co-activator of the transcription factor Scalloped (Sd), has a key role in initiating AM lineage reprogramming. An additional necessary input comes from active dJNK signaling, which contributes to the inactivation of the Hippo kinase cascade upstream of Yki and furthermore activates dJun. The synergistic activities of the Yki/Sd and dJun/dFos (AP-1) transcriptional activator complexes in the absence of Hippo activity initiate AM dedifferentiation and lead to the expression of Myc and piwi, which are crucial for different aspects of AM transdifferentiation. Our results provide new insights into the mechanisms that mediate muscle lineage plasticity during a cellular reprogramming process occurring in vivo.HighlightsDirect lineage reprogramming of alary muscles depends on Yorkie and JNKYorkie and JNK mediate reversal of syncytial muscle cell fateYki/Sd and AP-1 induce alary muscle dedifferentiation synergisticallyYki dependent Myc induces and Piwi mediates reprogramming of alary muscles

scholarly journals TMEM147 promotes  hepatocellular carcinoma progress  by inducing EGFR/MAPK activity

2020 ◽  
Author(s):  
Chengyi Sun ◽  
Jing Xu ◽  
She Tian ◽  
Yanqing Liu ◽  
Changhao Zhu ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Plasma Membrane ◽  
Mapk Pathway ◽  
Aerobic Glycolysis ◽  
Bioinformatic Analysis ◽  
Mapk Signaling ◽  
Clinicopathological Characteristics ◽  
Loss Of Function ◽  
Abnormal Glucose Metabolism

Abstract Backgroud: Hepatocellular carcinoma (HCC) is characterized by rapid early proliferation and distant metastasis and is extremely difficult to treat. Aerobic glycolysis is a hallmark of abnormal glucose metabolism in cancer cells as has been shown to be associated with tumor proliferation and metastasis; however, the mechanisms underlying aerobic glycolysis remain unclear. Methods: Immunohistochemistry (IHC) and qRT-PCR was performed to investigate the association between TMEM147 expression and the clinicopathological characteristics and prognosis of patients with HCC. Loss- and gain-of function assays were performed to investigate the role of TMEM147 in proliferation, metastasis and glycolysis in vitro and vivo. Bioinformatic analysis and rescue assay were used to demonstrated the TMEM147 interacted with EGFR and promoted its retromer-mediated recycling back to the plasma membrane.Results: we identified TMEM147 as a protein that was highly expressed and associated with poor survival in patients with HCC. Both gain- and loss-of-function studies revealed that TMEM147 acted as a key oncoprotein by promoting HCC growth, metastasis, and glycolysis via the EGFR/ MAPK signaling pathway. Mechanistically, TMEM147 interacted with EGFR and promoted its retromer-mediated recycling back to the plasma membrane, thus increasing the stability of EGFR and prolonging activation of the downstream MAPK pathway.Conclusion: Collectively, these results demonstrated the role and functional mechanism of TMEM147 in HCC, and indicated that TMEM147 may represent a prognostic biomarker and potential therapeutic target for HCC.

Regulation of mouse lens fiber cell development and differentiation by the Maf gene

Development ◽  
2000 ◽  
Vol 127 (2) ◽  
pp. 307-317 ◽  
Author(s):  
B.Z. Ring ◽  
S.P. Cordes ◽  
P.A. Overbeek ◽  
G.S. Barsh
Keyword(s):  
Cell Cycle ◽  
Gene Targeting ◽  
Leucine Zipper ◽  
Loss Of Function ◽  
Lens Fiber Cell ◽  
Related Family ◽  
Development And Differentiation ◽  
Lens Vesicle

Maf is a basic domain/leucine zipper domain protein originally identified as a proto-oncogene whose consensus target site in vitro, the T-MARE, is an extended version of an AP-1 site normally recognized by Fos and Jun. Maf and the closely related family members Neural retina leucine zipper (Nrl), L-Maf, and Krml1/MafB have been implicated in a wide variety of developmental and physiologic roles; however, mutations in vivo have been described only for Krml1/MafB, in which a loss-of-function causes abnormalities in hindbrain development due to failure to activate the Hoxa3 and Hoxb3 genes. We have used gene targeting to replace Maf coding sequences with those of lacZ, and have carried out a comprehensive analysis of embryonic expression and the homozygous mutant phenotype in the eye. Maf is expressed in the lens vesicle after invagination, and becomes highly upregulated in the equatorial zone, the site at which self-renewing anterior epithelial cells withdraw from the cell cycle and terminally differentiate into posterior fiber cells. Posterior lens cells in Maf(lacZ) mutant mice exhibit failure of elongation at embryonic day 11.5, do not express (α)A- and all of the (beta)-crystallin genes, and display inappropriately high levels of DNA synthesis. This phenotype partially overlaps with those reported for gene targeting of Prox1 and Sox1; however, expression of these genes is grossly normal, as is expression of Eya1, Eya2, Pax6, and Sox2. Recombinant Maf protein binds to T-MARE sites in the (alpha)A-, (beta)B2-, and (beta)A4-crystallin promoters but fails to bind to a point mutation in the (alpha)A-crystallin promoter that has been shown previously to be required for promoter function. Our results indicate that Maf directly activates many if not all of the (beta)-crystallin genes, and suggest a model for coordinating cell cycle withdrawal with terminal differentiation.

Abstract 442: Epigenetic Regulation of Ezh2 in Direct Human Cardiac Reprogramming

2020 ◽  
Vol 127 (Suppl_1) ◽  
Author(s):  
Yang Zhou ◽  
Yawen Tang ◽  
Lianzhong Zhao ◽  
Rui Lu ◽  
Jianyi Zhang
Keyword(s):  
Cardiovascular Disease ◽  
Cell Fate ◽  
Epigenetic Regulation ◽  
Ectopic Expression ◽  
The United States ◽  
Inhibitory Effect ◽  
Loss Of Function ◽  
Lower Efficiency

Cardiovascular disease is still the leading cause of death in the United States. Due to the limited regenerative capacity of adult hearts, the damage caused by heart injury could not be reversed and often progressed into heart failure. In need of cardiovascular disease treatment, many therapies aimed at either cell transplantation or cell regeneration have been proposed. Direct reprogramming of somatic cells into induced cardiomyocytes (iCMs) is considered to be a promising strategy for regenerative medicine. The induction of cardiomyocytes from non-myocytes has been achieved efficiently via ectopic expression of reprogramming factors both in vitro and in vivo with mice models. However, as human cells are more resistant to the reprogramming process, the generation of human iCMs (hiCMs) has been restricted by the factor that using more complex cocktails generated only functionally immature cells with lower efficiency and longer conversion time. The inefficiency of hiCMs production called for the identification and elucidation of underlying species-specific regulatory mechanisms in human, and removal of the additional epigenetic barriers which might be damping the hiCMs reprogramming. Here, we identified a human-specific epigenetic barrier, Enhancer of zesta homolog 2 (EZH2), via an unbiased loss-of-function screening. With the knockdown of EZH2, the hiCM reprogramming efficiency was significantly increased, accompanied with profound repression of collagen and extracellular matrix genes, which are related to the formation of fibrosis. Consistently, Inhibition of EZH2 catalytic activity via small molecules promotes hiCM reprogramming, suggesting that EZH2’s inhibitory effect was mediated by epigenetic regulation of histone modifications. Therefore, our study revealed a previously unrecognized regulatory mechanism of human cardiac reprogramming, which allows us to overcome the fibroblast fate barriers and ease the cardiac cell fate conversion.

scholarly journals Developmental and Transcriptional Consequences of Mutations in Drosophila TAFII60

2001 ◽  
Vol 21 (20) ◽  
pp. 6808-6819 ◽  
Author(s):  
Norikazu Aoyagi ◽  
David A. Wassarman
Keyword(s):  
Rna Polymerase Ii ◽  
Cell Fate ◽  
Transcription Initiation ◽  
Developmental Regulation ◽  
Regulation Of Gene Expression ◽  
Loss Of Function ◽  
Activator Proteins ◽  
Regulate Cell Growth

ABSTRACT In vitro, the TAFII60 component of the TFIID complex contributes to RNA polymerase II transcription initiation by serving as a coactivator that interacts with specific activator proteins and possibly as a promoter selectivity factor that interacts with the downstream promoter element. In vivo roles for TAFII60 in metazoan transcription are not as clear. Here we have investigated the developmental and transcriptional requirements for TAFII60 by analyzing four independent Drosophila melanogaster TAF II 60 mutants. Loss-of-function mutations in Drosophila TAF II 60 result in lethality, indicating that TAFII60 provides a nonredundant function in vivo. Molecular analysis of TAF II 60alleles revealed that essential TAFII60 functions are provided by two evolutionarily conserved regions located in the N-terminal half of the protein. TAFII60 is required at all stages of Drosophila development, in both germ cells and somatic cells. Expression of TAFII60 from a transgene rescued the lethality of TAF II 60mutants and exposed requirements for TAFII60 during imaginal development, spermatogenesis, and oogenesis. Phenotypes of rescued TAF II 60 mutant flies implicate TAFII60 in transcriptional mechanisms that regulate cell growth and cell fate specification and suggest that TAFII60 is a limiting component of the machinery that regulates the transcription of dosage-sensitive genes. Finally, TAFII60 plays roles in developmental regulation of gene expression that are distinct from those of other TAFIIproteins.

scholarly journals TMEM147 promotes  hepatocellular carcinoma progress  by inducing EGFR/MAPK activity

2020 ◽  
Author(s):  
Chengyi Sun ◽  
Jing Xu ◽  
She Tian ◽  
Yanqing Liu ◽  
Changhao Zhu ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Plasma Membrane ◽  
Mapk Pathway ◽  
Aerobic Glycolysis ◽  
Bioinformatic Analysis ◽  
Mapk Signaling ◽  
Clinicopathological Characteristics ◽  
Loss Of Function ◽  
Abnormal Glucose Metabolism

Abstract Backgroud: Hepatocellular carcinoma (HCC) is characterized by rapid early proliferation and distant metastasis and is extremely difficult to treat. Aerobic glycolysis is a hallmark of abnormal glucose metabolism in cancer cells as has been shown to be associated with tumor proliferation and metastasis; however, the mechanisms underlying aerobic glycolysis remain unclear. Methods: Immunohistochemistry (IHC) and qRT-PCR was performed to investigate the association between TMEM147 expression and the clinicopathological characteristics and prognosis of patients with HCC. Loss- and gain-of function assays were performed to investigate the role of TMEM147 in proliferation, metastasis and glycolysis in vitro and vivo. Bioinformatic analysis and rescue assay were used to demonstrated the TMEM147 interacted with EGFR and promoted its retromer-mediated recycling back to the plasma membrane.Results: we identified TMEM147 as a protein that was highly expressed and associated with poor survival in patients with HCC. Both gain- and loss-of-function studies revealed that TMEM147 acted as a key oncoprotein by promoting HCC growth, metastasis, and glycolysis via the EGFR/ MAPK signaling pathway. Mechanistically, TMEM147 interacted with EGFR and promoted its retromer-mediated recycling back to the plasma membrane, thus increasing the stability of EGFR and prolonging activation of the downstream MAPK pathway.Conclusion: Collectively, these results demonstrated the role and functional mechanism of TMEM147 in HCC, and indicated that TMEM147 may represent a prognostic biomarker and potential therapeutic target for HCC.

scholarly journals An in vitro stem cell model of human epiblast and yolk sac interaction

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Kirsty ML Mackinlay ◽  
Bailey AT Weatherbee ◽  
Viviane Souza Rosa ◽  
Charlotte E Handford ◽  
George Hudson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Cell Fate ◽  
Human Embryo ◽  
Cell Model ◽  
Embryonic Stem ◽  
Yolk Sac ◽  
Embryonic Cells ◽  
Post Implantation

Human embryogenesis entails complex signalling interactions between embryonic and extra-embryonic cells. However, how extra-embryonic cells direct morphogenesis within the human embryo remains largely unknown due to a lack of relevant stem cell models. Here, we have established conditions to differentiate human pluripotent stem cells (hPSCs) into yolk sac-like cells (YSLCs) that resemble the post-implantation human hypoblast molecularly and functionally. YSLCs induce the expression of pluripotency and anterior ectoderm markers in human embryonic stem cells (hESCs) at the expense of mesoderm and endoderm markers. This activity is mediated by the release of BMP and WNT signalling pathway inhibitors, and, therefore, resembles the functioning of the anterior visceral endoderm signalling centre of the mouse embryo, which establishes the anterior-posterior axis. Our results implicate the yolk sac in epiblast cell fate specification in the human embryo and propose YSLCs as a tool for studying post-implantation human embryo development in vitro.

scholarly journals PAX8-AS1 knockdown facilitates cell growth and inactivates autophagy in osteoblasts via the miR-1252-5p/GNB1 axis in osteoporosis

2021 ◽  
Author(s):  
Caiqiang Huang ◽  
Runguang Li ◽  
Changsheng Yang ◽  
Rui Ding ◽  
Qingchu Li ◽  
...  
Keyword(s):  
Antisense Rna ◽  
Bioinformatic Analysis ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Loss Of Function ◽  
Protein Subunit ◽  
Reporter Assays

AbstractOsteoporosis (OP) is the most common systematic bone disorder among elderly individuals worldwide. Long noncoding RNAs (lncRNAs) are involved in biological processes in various human diseases. It has been previously revealed that PAX8 antisense RNA 1 (PAX8-AS1) is upregulated in OP. However, its molecular mechanism in OP remains unclear. Therefore, we specifically designed this study to determine the specific role of PAX8-AS1 in OP. We first established a rat model of OP and then detected PAX8-AS1 expression in the rats with RT-qPCR. Next, to explore the biological function of PAX8-AS1 in osteoblasts, in vitro experiments, such as Cell Counting Kit-8 (CCK-8) assays, flow cytometry, western blotting and immunofluorescence (IF) staining, were conducted. Subsequently, we performed bioinformatic analysis and luciferase reporter assays to predict and identify the relationships between microRNA 1252-5p (miR-1252-5p) and both PAX8-AS1 and G protein subunit beta 1 (GNB1). Additionally, rescue assays in osteoblasts clarified the regulatory network of the PAX8-AS1/miR-1252-5p/GNB1 axis. Finally, in vivo loss-of-function studies verified the role of PAX8-AS1 in OP progression. The results illustrated that PAX8-AS1 was upregulated in the proximal tibia of OP rats. PAX8-AS1 silencing promoted the viability and inhibited the apoptosis and autophagy of osteoblasts. PAX8-AS1 interacted with miR-1252-5p. GNB1 was negatively regulated by miR-1252-5p. In addition, the impacts of PAX8-AS1 knockdown on osteoblasts were counteracted by GNB1 overexpression. PAX8-AS1 depletion suppressed OP progression by inhibiting apoptosis and autophagy in osteoblasts. In summary, PAX8-AS1 suppressed the viability and activated the autophagy of osteoblasts via the miR-1252-5p/GNB1 axis in OP.

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