scholarly journals Loss of AMPKalpha1 Triggers Centrosome Amplification via PLK4 Upregulation in Mouse Embryonic Fibroblasts

2020 ◽  
Vol 21 (8) ◽  
pp. 2772
Author(s):  
Qiang Zhao ◽  
Kathleen A Coughlan ◽  
Ming-Hui Zou ◽  
Ping Song
Keyword(s):  
Molecular Mechanisms ◽  
Adenosine Monophosphate ◽  
Centrosome Amplification ◽  
Wild Type ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Cellular Energy ◽  
Chromosome Missegregation ◽  
Cell Mitosis ◽  
Metaphase Chromosome Spread

Recent evidence indicates that activation of adenosine monophosphate-activated protein kinase (AMPK), a highly conserved sensor and modulator of cellular energy and redox, regulates cell mitosis. However, the underlying molecular mechanisms for AMPKα subunit regulation of chromosome segregation remain poorly understood. This study aimed to ascertain if AMPKα1 deletion contributes to chromosome missegregation by elevating Polo-like kinase 4 (PLK4) expression. Centrosome proteins and aneuploidy were monitored in cultured mouse embryonic fibroblasts (MEFs) isolated from wild type (WT, C57BL/6J) or AMPKα1 homozygous deficient (AMPKα1−/−) mice by Western blotting and metaphase chromosome spread. Deletion of AMPKα1, the predominant AMPKα isoform in immortalized MEFs, led to centrosome amplification and chromosome missegregation, as well as the consequent aneuploidy (34–66%) and micronucleus. Furthermore, AMPKα1 null cells exhibited a significant induction of PLK4. Knockdown of nuclear factor kappa B2/p52 ameliorated the PLK4 elevation in AMPKα1-deleted MEFs. Finally, PLK4 inhibition by Centrinone reversed centrosome amplification of AMPKα1-deleted MEFs. Taken together, our results suggest that AMPKα1 plays a fundamental role in the maintenance of chromosomal integrity through the control of p52-mediated transcription of PLK4, a trigger of centriole biogenesis.

NF-κB Protection against Apoptosis Induced by HEMA

2004 ◽  
Vol 83 (11) ◽  
pp. 837-842 ◽  
Author(s):  
G. Spagnuolo ◽  
C. Mauro ◽  
A. Leonardi ◽  
M. Santillo ◽  
R. Paternò ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Protective Role ◽  
Oxygen Species ◽  
Wild Type ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Primary Fibroblasts ◽  
Underlying Mechanisms ◽  
Induced Apoptosis ◽  
Dose Dependent

The cytotoxicity of dental monomers has been widely investigated, but the underlying mechanisms have not been elucidated. We studied the molecular mechanisms involved in cell death induced by HEMA. In human primary fibroblasts, HEMA induced a dose-dependent apoptosis that was confirmed by the activation of caspases-8, -9, and -3. We found an increase of reactive oxygen species (ROS) and NF-κB activation after HEMA exposure. Blocking of ROS production by anti-oxidants had no direct influence on apoptosis caused by HEMA, but inhibition of NF-κB increased the fraction of apoptotic cells. Accordingly, mouse embryonic fibroblasts (MEF) from p65−/− mice were more susceptible to HEMA-induced apoptosis than were wild-type controls. Our results indicate that exposure to HEMA triggers apoptosis and that this mechanism is not directly dependent upon redox signaling. Nevertheless, ROS induction by HEMA activates NF-κB, which exerts a protective role in counteracting apoptosis.

Differential expression of prohibitin and regulation of apoptosis in wild-type and COX-2 null mouse embryonic fibroblasts

2009 ◽  
Vol 53 (2) ◽  
pp. 157-159 ◽  
Author(s):  
Young-Hoon Kim ◽  
Hyangkyu Lee ◽  
Tae-Yoon Kim ◽  
Hyang-Ran Hwang ◽  
Sang Chul Lee
Keyword(s):  
Differential Expression ◽  
Null Mouse ◽  
Wild Type ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Cox 2

scholarly journals Caspase3-deficient cells require fibronectin for protection against autophagy-dependent death

2021 ◽  
Author(s):  
David B. Weir ◽  
Lawrence H. Boise
Keyword(s):  
Cell Death ◽  
Survival Advantage ◽  
Deficient Mouse ◽  
Wild Type ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Serum Withdrawal ◽  
Presence And Absence ◽  
Induced Apoptosis

ABSTRACTCaspases are required for execution of apoptosis. However, in their absence, signals that typically induce apoptosis can still result in cell death. Our laboratory previously demonstrated that Casp3-deficient mouse embryonic fibroblasts (MEFs) have increased fibronectin (FN) secretion, and an adhesion-dependent survival advantage compared to wild type (WT) MEFs. Here, we show that FN is required for survival of Casp3-deficient MEFs following serum withdrawal. Furthermore, when FN is silenced, serum withdrawal-induced death is caspase-independent. However, procaspase-7 is cleaved, suggesting that MOMP is taking place. Indeed, in the absence of FN, cytochrome c release is increased following serum withdrawal in Casp3-deficient MEFs. Yet death does not correspond to cytochrome c release in Casp3-deficient MEFs. This is true both in the presence and absence of FN. Additionally, caspase-independent death is inhibited by Bcl-XL overexpression. These findings suggest that Bcl-XL is not inhibiting death through regulation of Bax/Bak insertion into the mitochondria, but through a different mechanism. One such possibility is autophagy and induction of autophagy is associated with caspase-independent death in Casp3-deficient cells. Importantly, when ATG5 is ablated in Casp3-deficient cells, autophagy is blocked and death is largely inhibited. Taken together, our data indicate that Casp3-deficient cells incapable of undergoing canonical serum withdrawal-induced apoptosis, are protected from autophagy-dependent death by FN-mediated adhesion.

scholarly journals The Absence of Ser389 Phosphorylation in p53 Affects the Basal Gene Expression Level of Many p53-Dependent Genes and Alters the Biphasic Response to UV Exposure in Mouse Embryonic Fibroblasts

2008 ◽  
Vol 28 (6) ◽  
pp. 1974-1987 ◽  
Author(s):  
Wendy Bruins ◽  
Oskar Bruning ◽  
Martijs J. Jonker ◽  
Edwin Zwart ◽  
Tessa V. van der Hoeven ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Damage ◽  
Uv Irradiation ◽  
Target Genes ◽  
Late Response ◽  
Wild Type ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Endogenous Gene ◽  
Cellular Processes

ABSTRACT Phosphorylation is important in p53-mediated DNA damage responses. After UV irradiation, p53 is phosphorylated specifically at murine residue Ser389. Phosphorylation mutant p53.S389A cells and mice show reduced apoptosis and compromised tumor suppression after UV irradiation. We investigated the underlying cellular processes by time-series analysis of UV-induced gene expression responses in wild-type, p53.S389A, and p53−/− mouse embryonic fibroblasts. The absence of p53.S389 phosphorylation already causes small endogenous gene expression changes for 2,253, mostly p53-dependent, genes. These genes showed basal gene expression levels intermediate to the wild type and p53−/−, possibly to readjust the p53 network. Overall, the p53.S389A mutation lifts p53-dependent gene repression to a level similar to that of p53−/− but has lesser effect on p53-dependently induced genes. In the wild type, the response of 6,058 genes to UV irradiation was strictly biphasic. The early stress response, from 0 to 3 h, results in the activation of processes to prevent the accumulation of DNA damage in cells, whereas the late response, from 12 to 24 h, relates more to reentering the cell cycle. Although the p53.S389A UV gene response was only subtly changed, many cellular processes were significantly affected. The early response was affected the most, and many cellular processes were phase-specifically lost, gained, or altered, e.g., induction of apoptosis, cell division, and DNA repair, respectively. Altogether, p53.S389 phosphorylation seems essential for many p53 target genes and p53-dependent processes.

scholarly journals Mitochondrial LonP1 protease is implicated in the degradation of unstable Parkinson's disease-associated DJ-1/PARK 7 missense mutants

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Raúl Sánchez-Lanzas ◽  
José G. Castaño
Keyword(s):  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Mitochondrial Pathway ◽  
Mitochondrial Matrix ◽  
Wild Type ◽  
Strong Reduction ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Biochemical Fractionation ◽  
Early Onset Parkinson’S Disease

AbstractDJ-1/PARK7 mutations are linked with familial forms of early-onset Parkinson's disease (PD). We have studied the degradation of untagged DJ-1 wild type (WT) and missense mutants in mouse embryonic fibroblasts obtained from DJ-1-null mice, an approach closer to the situation in patients carrying homozygous mutations. The results showed that the mutants L10P, M26I, A107P, P158Δ, L166P, E163K, and L172Q are unstable proteins, while A39S, E64D, R98Q, A104T, D149A, A171S, K175E, and A179T are as stable as DJ-1 WT. Inhibition of proteasomal and autophagic-lysosomal pathways had little effect on their degradation. Immunofluorescence and biochemical fractionation studies indicated that M26I, A107P, P158Δ, L166P, E163K, and L172Q mutants associate with mitochondria. Silencing of mitochondrial matrix protease LonP1 produced a strong reduction of the degradation of the mitochondrial-associated DJ-1 mutants A107P, P158Δ, L166P, E163K, and L172Q but not of mutant L10P. These results demonstrated a mitochondrial pathway of degradation of those DJ-1 missense mutants implicated in PD pathogenesis.

Abstract 73: A Novel Role of E2F1 in Stress-induced Cardiac Fibrosis

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
Dauren Biyashev ◽  
Chan Boriboun ◽  
Gangjian Qin
Keyword(s):  
Cardiovascular System ◽  
Cardiac Fibrosis ◽  
Cardiovascular Health ◽  
Transmembrane Protein ◽  
Ang Ii ◽  
Tgf Beta ◽  
Wild Type ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts

Rationale: E2F1 transcription factor is best known as a cell cycle regulator. Recent reports indicate the importance of E2F1 in cardiovascular system, though its exact role is not clear. TGF-beta/Smad2,3 signaling pathway, on the other hand, has been long implicated in the regulation of cardiovascular health and numerous disease, including cardiac fibrosis. Interaction between these two major pathways has been reported in the cancer settings. Objective: To identify the possible interactions between E2F1 and TGF-beta/Smad2,3 signaling pathways in cardiovascular system and determine the functional outcome of these interactions in cardiac health. Methods and Results: E2F1-/- mice developed significantly higher degree of cardiac fibrosis than wild type mice in the Angiotensin II - induced cardiac fibrosis model. The levels of phosphorylated Smad2 and Smad3 were significantly higher in the hearts of E2F1-/- mice, as well as in mouse embryonic fibroblasts derived from E2F1-/- animals. Associated expression of collagen I was significantly increased in mouse embryonic fibroblasts derived from E2F1-/- animals, and treatment with TGF-beta resulted in higher collagen deposition compared to wild type fibroblasts. Treating animals with SB 431542, chemical inhibitor of Smad2,3 signaling, obliterated the difference in the degree of cardiac fibrosis between wild type and E2F1 knockout animals in the Ang II model. We discovered that levels of syndecan-4, heparan sulfate proteoglycan transmembrane protein implicated in fibrosis and known to interact with TGF-beta are significantly increased in both E2F1-/- fibroblasts and hearts. siRNA-mediated knockdown of syndecan-4 using siRNA resulted in decreased Smad2,3 phosphorylation in E2F1-/- MEFs. Similarly, down regulation of syndecan-4 in-vivo using morpholino lead to decreased cardiac fibrosis in E2F1-/- mice in Ang II model. Conclusions: E2F1 suppresses activation of TGF-beta/Smad 2,3 pathway. The E2F1-dependent suppression of cardiac fibrosis through TGF-beta/Smad 2,3 pathway is at least partially regulated by syndecan-4.

scholarly journals Lmna knockout mouse embryonic fibroblasts are less contractile than their wild-type counterparts

2017 ◽  
Vol 9 (8) ◽  
pp. 709-721 ◽  
Author(s):  
I. A. E. W. van Loosdregt ◽  
M. A. F. Kamps ◽  
C. W. J. Oomens ◽  
S. Loerakker ◽  
J. L. V. Broers ◽  
...  
Keyword(s):  
Knockout Mouse ◽  
Stress Fiber ◽  
Stress Generation ◽  
Actin Stress Fiber ◽  
Wild Type ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Contractile Stress ◽  
Fiber Organization

Lmna knockout causes an impaired actin stress fiber organization which results in a fivefold lower contractile stress generation.

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