Investigation of Myelinogenesis in Vitro: Transient Expression of 3,5,3’-Triiodothyronine Nuclear Receptors in Secondary Cultures of Pure Rat Oligodendrocytes

1989 ◽  
pp. 113-130 ◽  
Author(s):  
L. L. Sarliève ◽  
F. Besnard ◽  
G. Labourdette ◽  
B. Yusta ◽  
A. Pascual ◽  
...  
Keyword(s):  
Transient Expression ◽  
Nuclear Receptors

Transient expression of 3,5,3′-triiodothyronine nuclear receptors in rat oligodendrocytes: In vivo and in vitro immunocytochemical studies

1994 ◽  
Vol 37 (3) ◽  
pp. 313-323 ◽  
Author(s):  
F. Besnard ◽  
M. Luo ◽  
M. Miehe ◽  
J. H. Dusssault ◽  
J. Puymirat ◽  
...  
Keyword(s):  
Transient Expression ◽  
Nuclear Receptors ◽  
Immunocytochemical Studies

Mouse models to study von Willebrand factor structure-function relationships in vivo

2009 ◽  
Vol 29 (01) ◽  
pp. 17-20 ◽  
Author(s):  
I. Marx ◽  
I. Badirou ◽  
R. Pendu ◽  
O. Christophe ◽  
C. V. Denis
Keyword(s):  
Structure Function ◽  
Transient Expression ◽  
Von Willebrand Factor ◽  
Large Size ◽  
Mouse Hepatocytes ◽  
Function Relationship ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryVon Willebrand factor (VWF) structure-function relationship has been studied only through in vitro approaches. The VWF-deficient mouse model has been extremely useful to examine the in vivo function of VWF but does not allow a more subtle analysis of the relative importance of its different domains. However, considering the large size of VWF and its capacity to interact with various ligands in order to support platelet adhesion and aggregation, the necessity to evaluate independently these interactions appeared increasingly crucial. A recently developed technique, known as hydrodynamic injection, which allows transient expression of a transgene by mouse hepatocytes, proved very useful in this regard. Indeed, transient expression of various VWF mutants in VWF-deficient mice contributed to improve our knowledge about the role of VWF interaction with subendothelial collagens and with platelets receptors in VWF roles in haemostasis and thrombosis. These findings can provide new leads in the development of anti-thrombotic therapies.

scholarly journals Induction of the Cellular E2F-1 Promoter by the Adenovirus E4-6/7 Protein

2000 ◽  
Vol 74 (5) ◽  
pp. 2084-2093 ◽  
Author(s):  
Joel Schaley ◽  
Robert J. O'Connor ◽  
Laura J. Taylor ◽  
Dafna Bar-Sagi ◽  
Patrick Hearing
Keyword(s):  
Dna Binding ◽  
Binding Site ◽  
Transient Expression ◽  
Binding Sites ◽  
Promoter Region ◽  
Fluorescent Protein ◽  
Protein Accumulation ◽  
Promoter Regions ◽  
Single Binding Site

ABSTRACT The adenovirus type 5 (Ad5) E4-6/7 protein interacts directly with different members of the E2F family and mediates the cooperative and stable binding of E2F to a unique pair of binding sites in the Ad5 E2a promoter region. This induction of E2F DNA binding activity strongly correlates with increased E2a transcription when analyzed using virus infection and transient expression assays. Here we show that while different adenovirus isolates express an E4-6/7 protein that is capable of induction of E2F dimerization and stable DNA binding to the Ad5 E2a promoter region, not all of these viruses carry the inverted E2F binding site targets in their E2a promoter regions. The Ad12 and Ad40 E2a promoter regions bind E2F via a single binding site. However, these promoters bind adenovirus-induced (dimerized) E2F very weakly. The Ad3 E2a promoter region binds E2F very poorly, even via a single binding site. A possible explanation of these results is that the Ad E4-6/7 protein evolved to induce cellular gene expression. Consistent with this notion, we show that infection with different adenovirus isolates induces the binding of E2F to an inverted configuration of binding sites present in the cellular E2F-1 promoter. Transient expression of the E4-6/7 protein alone in uninfected cells is sufficient to induce transactivation of the E2F-1 promoter linked to chloramphenicol acetyltransferase or green fluorescent protein reporter genes. Further, expression of the E4-6/7 protein in the context of adenovirus infection induces E2F-1 protein accumulation. Thus, the induction of E2F binding to the E2F-1 promoter by the E4-6/7 protein observed in vitro correlates with transactivation of E2F-1 promoter activity in vivo. These results suggest that adenovirus has evolved two distinct mechanisms to induce the expression of the E2F-1 gene. The E1A proteins displace repressors of E2F activity (the Rb family members) and thus relieve E2F-1 promoter repression; the E4-6/7 protein complements this function by stably recruiting active E2F to the E2F-1 promoter to transactivate expression.

scholarly journals Activating Signal Cointegrator 2 Belongs to a Novel Steady-State Complex That Contains a Subset of Trithorax Group Proteins

2003 ◽  
Vol 23 (1) ◽  
pp. 140-149 ◽  
Author(s):  
Young-Hwa Goo ◽  
Young Chang Sohn ◽  
Dae-Hwan Kim ◽  
Seung-Whan Kim ◽  
Min-Jung Kang ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Steady State ◽  
Nuclear Receptors ◽  
Transcriptional Activation ◽  
Retinoic Acid Receptor ◽  
Dependent Manner ◽  
Trithorax Group ◽  
Trithorax Group Proteins

ABSTRACT Many transcription coactivators interact with nuclear receptors in a ligand- and C-terminal transactivation function (AF2)-dependent manner. These include activating signal cointegrator 2 (ASC-2), a recently isolated transcriptional coactivator molecule, which is amplified in human cancers and stimulates transactivation by nuclear receptors and numerous other transcription factors. In this report, we show that ASC-2 belongs to a steady-state complex of approximately 2 MDa (ASC-2 complex [ASCOM]) in HeLa nuclei. ASCOM contains retinoblastoma-binding protein RBQ-3, α/β-tubulins, and trithorax group proteins ALR-1, ALR-2, HALR, and ASH2. In particular, ALR-1/2 and HALR contain a highly conserved 130- to 140-amino-acid motif termed the SET domain, which was recently implicated in histone H3 lysine-specific methylation activities. Indeed, recombinant ALR-1, HALR, and immunopurified ASCOM exhibit very weak but specific H3-lysine 4 methylation activities in vitro, and transactivation by retinoic acid receptor appears to involve ligand-dependent recruitment of ASCOM and subsequent transient H3-lysine 4 methylation of the promoter region in vivo. Thus, ASCOM may represent a distinct coactivator complex of nuclear receptors. Further characterization of ASCOM will lead to a better understanding of how nuclear receptors and other transcription factors mediate transcriptional activation.

The phylogenetically invariant ACAGAGA and AGC sequences of U6 small nuclear RNA are more tolerant of mutation in human cells than in Saccharomyces cerevisiae

1993 ◽  
Vol 13 (9) ◽  
pp. 5377-5382
Author(s):  
B Datta ◽  
A M Weiner
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transient Expression ◽  
Human Cells ◽  
Mrna Splicing ◽  
Small Nuclear Rna ◽  
Transient Expression Assay ◽  
U6 Snrna ◽  
Nuclear Rna

U6 small nuclear RNA (snRNA) is the most highly conserved of the five spliceosomal snRNAs that participate in nuclear mRNA splicing. The proposal that U6 snRNA plays a key catalytic role in splicing [D. Brow and C. Guthrie, Nature (London) 337:14-15, 1989] is supported by the phylogenetic conservation of U6, the sensitivity of U6 to mutation, cross-linking of U6 to the vicinity of the 5' splice site, and genetic evidence for extensive base pairing between U2 and U6 snRNAs. We chose to mutate the phylogenetically invariant 41-ACAGAGA-47 and 53-AGC-55 sequences of human U6 because certain point mutations within the homologous regions of Saccharomyces cerevisiae U6 selectively block the first or second step of mRNA splicing. We found that both sequences are more tolerant to mutation in human cells (assayed by transient expression in vivo) than in S. cerevisiae (assayed by effects on growth or in vitro splicing). These differences may reflect different rate-limiting steps in the particular assays used or differential reliance on redundant RNA-RNA or RNA-protein interactions. The ability of mutations in U6 nucleotides A-45 and A-53 to selectively block step 2 of splicing in S. cerevisiae had previously been construed as evidence that these residues might participate directly in the second chemical step of splicing; an indirect, structural role seems more likely because the equivalent mutations have no obvious phenotype in the human transient expression assay.

scholarly journals Production of hepatitis B virus in vitro by transient expression of cloned HBV DNA in a hepatoma cell line.

1987 ◽  
Vol 6 (3) ◽  
pp. 675-680 ◽  
Author(s):  
C.M. Chang ◽  
K.S. Jeng ◽  
C.P. Hu ◽  
S.J. Lo ◽  
T.S. Su ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Cell Line ◽  
Transient Expression ◽  
Hepatoma Cell ◽  
Hbv Dna ◽  
Hepatoma Cell Line ◽  
B Virus

scholarly journals Liver X Receptor Nuclear Receptors Are Transcriptional Regulators of Dendritic Cell Chemotaxis

2018 ◽  
Vol 38 (10) ◽  
Author(s):  
Susana Beceiro ◽  
Attila Pap ◽  
Zsolt Czimmerer ◽  
Tamer Sallam ◽  
Jose A. Guillén ◽  
...  
Keyword(s):  
Nuclear Receptors ◽  
Low Density Lipoprotein ◽  
Myeloid Cells ◽  
Density Lipoprotein ◽  
Loss Of Function ◽  
Transcriptional Responses ◽  
Cd38 Expression ◽  
The Impact

ABSTRACTThe liver X receptors (LXRs) are ligand-activated nuclear receptors with established roles in the maintenance of lipid homeostasis in multiple tissues. LXRs exert additional biological functions as negative regulators of inflammation, particularly in macrophages. However, the transcriptional responses controlled by LXRs in other myeloid cells, such as dendritic cells (DCs), are still poorly understood. Here we used gain- and loss-of-function models to characterize the impact of LXR deficiency on DC activation programs. Our results identified an LXR-dependent pathway that is important for DC chemotaxis. LXR-deficient mature DCs are defective in stimulus-induced migrationin vitroandin vivo. Mechanistically, we show that LXRs facilitate DC chemotactic signaling by regulating the expression of CD38, an ectoenzyme important for leukocyte trafficking. Pharmacological or genetic inactivation of CD38 activity abolished the LXR-dependent induction of DC chemotaxis. Using the low-density lipoprotein receptor-deficient (LDLR−/−) LDLR−/−mouse model of atherosclerosis, we also demonstrated that hematopoietic CD38 expression is important for the accumulation of lipid-laden myeloid cells in lesions, suggesting that CD38 is a key factor in leukocyte migration during atherogenesis. Collectively, our results demonstrate that LXRs are required for the efficient emigration of DCs in response to chemotactic signals during inflammation.

β-Adrenergic receptors, voltage-operated Ca 2+-channels, nuclear triiodothyronine receptors and triiodothyronine concentration in pig myocardium after long-term low-dose amiodarone treatment

1993 ◽  
Vol 129 (4) ◽  
pp. 337-347 ◽  
Author(s):  
Liv Bjørn-Hansen Gøtzsche
Keyword(s):  
Cardiac Surgery ◽  
Nuclear Receptors ◽  
Adrenergic Receptors ◽  
Low Dose ◽  
Binding Capacity ◽  
Drug Concentrations ◽  
Cardiac Strain ◽  
Amiodarone Treatment ◽  
Β Adrenergic Receptors

Similar features during chronic amiodarone treatment and hypothyroidism suggest that amiodarone induces a state of "triiodothyronine (T3)-resistance" or "cardiac hypothyroidism", which may predispose the heart to pump failure under conditions with severe strain, such as recovery after cardiac surgery. Disagreements exist as to how amiodarone, and possibly its main metabolite desethylamiodarone, act upon the various receptor systems in the heart. The aim of the present study was to elucidate whether chronic amiodarone treatment leads to a functional reduction in the number of myocardial nuclear T3 receptors, the myocardial tissue T3 concentration and the number of β-receptors and voltage-operated Ca2+-channels. Finally, special attention was drawn to any changes that could contribute to explain previous reports on reduced haemodynamic reserve in animals exposed to severe cardiac strain, such as cardiac surgery. Pigs (72±2 kg) were assigned randomly to amiodarone treatment (20 mg·kg−1·day−1 for 30±1 days, N = 8); controls received no medical treatment (N = 6). The left ventricle was evaluated for β-adrenergic receptors, voltage-operated Ca2+-channels, T3 nuclear receptors and tissue T3 concentration. Maximum binding capacity for β-receptors and Ca2+-channels was reduced in amiodarone-treated pigs (by 38%, p<0.05, and by 52%, p<0.01) and correlated with tissue drug concentrations for both receptor types (p<0.05). No changes were observed concerning nuclear T3 receptors. In vitro competition studies revealed that amiodarone, but not desethylamiodarone, possessed binding properties to Ca2+-channels, whereas neither of the compounds bound to β-receptors. Desethylamiodarone, but not amiodarone, competitively inhibited T3 binding to its nuclear receptors. Myocardial T3 was undetectable (<0.05 nmol/kg wet wt) in amiodarone-treated pigs. From our observations we suggest that the active metabolite desethylamiodarone, rather than the parent drug, is mainly responsible for the observed local hypothyroid-like effects during amiodarone treatment. The observed changes after treatment with low-dose amiodarone in pigs are likely to have biological implications. Functionally, the changes may imply reduced cardiac reserve during conditions of extraordinary strain.

scholarly journals Thyroid Hormone Signaling in Embryonic Stem Cells: Crosstalk with the Retinoic Acid Pathway

2020 ◽  
Vol 21 (23) ◽  
pp. 8945
Author(s):  
Mercedes Fernández ◽  
Micaela Pannella ◽  
Vito Antonio Baldassarro ◽  
Alessandra Flagelli ◽  
Giuseppe Alastra ◽  
...  
Keyword(s):  
Stem Cells ◽  
Retinoic Acid ◽  
Embryonic Stem Cells ◽  
Thyroid Hormone ◽  
Nuclear Receptors ◽  
Embryonic Stem ◽  
Embryoid Bodies ◽  
Postnatal Life ◽  
Gene And Protein Expression

While the role of thyroid hormones (THs) during fetal and postnatal life is well-established, their role at preimplantation and during blastocyst development remains unclear. In this study, we used an embryonic stem cell line isolated from rat (RESC) to study the effects of THs and retinoic acid (RA) on early embryonic development during the pre-implantation stage. The results showed that THs play an important role in the differentiation/maturation processes of cells obtained from embryoid bodies (EB), with thyroid hormone nuclear receptors (TR) (TRα and TRβ), metabolic enzymes (deiodinases 1, 2, 3) and membrane transporters (Monocarboxylate transporters -MCT- 8 and 10) being expressed throughout in vitro differentiation until the Embryoid body (EB) stage. Moreover, thyroid hormone receptor antagonist TR (1-850) impaired RA-induced neuroectodermal lineage specification. This effect was significantly higher when cells were treated with retinoic acid (RA) to induce neuroectodermal lineage, studied through the gene and protein expression of nestin, an undifferentiated progenitor marker from the neuroectoderm lineage, as established by nestin mRNA and protein regulation. These results demonstrate the contribution of the two nuclear receptors, TR and RA, to the process of neuroectoderm maturation of the in vitro model embryonic stem cells obtained from rat.

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