Regulation, mechanisms and proposed function of ferritin translocation to cell nuclei

2002 ◽  
Vol 115 (10) ◽  
pp. 2165-2177
Author(s):  
Khristy J. Thompson ◽  
Michael G. Fried ◽  
Zheng Ye ◽  
Phillip Boyer ◽  
James R. Connor
Keyword(s):  
Cell Culture ◽  
Oxidative Damage ◽  
Biological Significance ◽  
Nuclear Pore ◽  
Cell Nuclei ◽  
Iron Storage ◽  
Cellular Iron ◽  
Human Astrocytoma ◽  
Culture Models

Ferritin is traditionally considered a cytoplasmic iron-storage protein,but recent reports indicate that it is also found in cell nuclei. Nuclear ferritin has been proposed to be involved in both the protection of DNA and the exacerbation of iron-induced oxidative damage to DNA. We demonstrate that H-rich ferritin is present in the nucleus of human astrocytoma tumor cells. To study the mechanism and regulation of ferritin translocation to the nucleus,we developed a cell culture model using SW1088 human astrocytoma cells. Changes in cellular iron levels, cytokine treatments and hydrogen peroxide exposure affected the distribution of ferritin between the cytosol and the nucleus. Ferritin enters the nucleus via active transport through the nuclear pore and does not require NLS-bearing cytosolic factors for transport. Furthermore, H-rich ferritin is preferred over L-rich ferritin for uptake into the nucleus. Whole cell crosslinking studies revealed that ferritin is associated with DNA. Ferritin protected DNA from iron-induced oxidative damage in both in vitro and in cell culture models. These results strongly suggest a novel role for ferritin in nuclear protection. This work should lead to novel characterization of ferritin functions in the context of genomic stability and may have unparalleled biological significance in terms of the accessibility of metals to DNA. The knowledge generated as a result of these studies will also improve our understanding of iron-induced damage of nuclear constituents.

scholarly journals Turnover of the Active Fraction of IRS1 Involves Raptor-mTOR- and S6K1-Dependent Serine Phosphorylation in Cell Culture Models of Tuberous Sclerosis

2006 ◽  
Vol 26 (17) ◽  
pp. 6425-6434 ◽  
Author(s):  
O. Jameel Shah ◽  
Tony Hunter
Keyword(s):  
Cell Culture ◽  
Tuberous Sclerosis ◽  
High Speed ◽  
Serine Phosphorylation ◽  
Feedback Signal ◽  
Insulin Receptor Substrates ◽  
Igf I ◽  
Culture Models ◽  
Cell Culture Models

ABSTRACT The TSC1-TSC2/Rheb/Raptor-mTOR/S6K1 cell growth cassette has recently been shown to regulate cell autonomous insulin and insulin-like growth factor I (IGF-I) sensitivity by transducing a negative feedback signal that targets insulin receptor substrates 1 and 2 (IRS1 and -2). Using two cell culture models of the familial hamartoma syndrome, tuberous sclerosis, we show here that Raptor-mTOR and S6K1 are required for phosphorylation of IRS1 at a subset of serine residues frequently associated with insulin resistance, including S307, S312, S527, S616, and S636 (of human IRS1). Using loss- and gain-of-function S6K1 constructs, we demonstrate a requirement for the catalytic activity of S6K1 in both direct and indirect regulation of IRS1 serine phosphorylation. S6K1 phosphorylates IRS1 in vitro on multiple residues showing strong preference for RXRXXS/T over S/T,P sites. IRS1 is preferentially depleted from the high-speed pellet fraction in TSC1/2-deficient mouse embryo fibroblasts or in HEK293/293T cells overexpressing Rheb. These studies suggest that, through serine phosphorylation, Raptor-mTOR and S6K1 cell autonomously promote the depletion of IRS1 from specific intracellular pools in pathological states of insulin and IGF-I resistance and thus potentially in lesions associated with tuberous sclerosis.

scholarly journals Conjunctival melanoma and electrochemotherapy: preliminary results using 2D and 3D cell culture models in vitro

2018 ◽  
Vol 97 (4) ◽  
pp. e632-e640 ◽  
Author(s):  
Miltiadis Fiorentzis ◽  
Periklis Katopodis ◽  
Helen Kalirai ◽  
Berthold Seitz ◽  
Arne Viestenz ◽  
...  
Keyword(s):  
Cell Culture ◽  
3D Cell Culture ◽  
Conjunctival Melanoma ◽  
Preliminary Results ◽  
Culture Models ◽  
2D And 3D ◽  
Cell Culture Models

scholarly journals Applying phasor approach analysis of multiphoton FLIM measurements to probe the metabolic activity of three-dimensional in vitro cell culture models

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Pirmin H. Lakner ◽  
Michael G. Monaghan ◽  
Yvonne Möller ◽  
Monilola A. Olayioye ◽  
Katja Schenke-Layland
Keyword(s):  
Cell Culture ◽  
Metabolic Activity ◽  
Three Dimensional ◽  
In Vitro Cell Culture ◽  
Culture Models ◽  
Cell Culture Models

scholarly journals In vitro models of the blood–brain barrier: An overview of commonly used brain endothelial cell culture models and guidelines for their use

2016 ◽  
Vol 36 (5) ◽  
pp. 862-890 ◽  
Author(s):  
Hans C Helms ◽  
N Joan Abbott ◽  
Malgorzata Burek ◽  
Romeo Cecchelli ◽  
Pierre-Olivier Couraud ◽  
...  
Keyword(s):  
Cell Culture ◽  
Endothelial Cell ◽  
Blood Brain Barrier ◽  
Brain Parenchyma ◽  
Brain Barrier ◽  
Blood Brain ◽  
Culture Models ◽  
Cell Culture Models ◽  
The Brain

The endothelial cells lining the brain capillaries separate the blood from the brain parenchyma. The endothelial monolayer of the brain capillaries serves both as a crucial interface for exchange of nutrients, gases, and metabolites between blood and brain, and as a barrier for neurotoxic components of plasma and xenobiotics. This “blood-brain barrier” function is a major hindrance for drug uptake into the brain parenchyma. Cell culture models, based on either primary cells or immortalized brain endothelial cell lines, have been developed, in order to facilitate in vitro studies of drug transport to the brain and studies of endothelial cell biology and pathophysiology. In this review, we aim to give an overview of established in vitro blood–brain barrier models with a focus on their validation regarding a set of well-established blood–brain barrier characteristics. As an ideal cell culture model of the blood–brain barrier is yet to be developed, we also aim to give an overview of the advantages and drawbacks of the different models described.

In Vitro Human Lung Cell Culture Models to Study the Toxic Potential of Nanoparticles

Nanotoxicity ◽  
2009 ◽  
pp. 379-395 ◽  
Author(s):  
Fabian Blank ◽  
Peter Gehr ◽  
Barbara Rothen-Rutishauser
Keyword(s):  
Cell Culture ◽  
Human Lung ◽  
Culture Models ◽  
Toxic Potential ◽  
Cell Culture Models

scholarly journals A Pilot Study To Establish an In Vitro Model To Study Premature Intestinal Epithelium and Gut Microbiota Interactions

mSphere ◽  
2021 ◽  
Author(s):  
Justin Gibbons ◽  
Ji Youn Yoo ◽  
Tina Mutka ◽  
Maureen Groer ◽  
Thao T. B. Ho
Keyword(s):  
Cell Culture ◽  
In Vitro Model ◽  
Bacterial Flora ◽  
Mucosal Barrier ◽  
Clinical Observations ◽  
Culture Models ◽  
Vitro Model ◽  
Cell Culture Models

The gut bacterial flora influences the development of the immune system and long-term health outcomes in preterm infants. Studies of the mechanistic interactions between the gut bacteria and mucosal barrier are limited to clinical observations, animal models, and in vitro cell culture models for this vulnerable population.

scholarly journals Emerging Mechanisms of Cellular Iron Transport and Trafficking

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. SCI-2-SCI-2
Author(s):  
Caroline Philpott ◽  
Moon-Suhn Ryu
Keyword(s):  
Iron Deficiency ◽  
Mammalian Cells ◽  
Conflicts Of Interest ◽  
Friedreich Ataxia ◽  
Iron Storage ◽  
Mononuclear Iron ◽  
Early Stages ◽  
Cellular Iron ◽  
Dinuclear Iron

Abstract Iron is an essential nutrient required by every cell in the human body, yet it can also be a potent cellular toxin. Iron is essential because enzymes that require iron co-factors (namely, heme, iron-sulfur clusters, mono- and dinuclear iron centers) are involved in virtually every major metabolic process in the cell. Iron deficiency continues to be the most common nutritional deficiency in the world, while iron overload is a feature of an increasing number of human diseases, including genetic disorders such as hereditary hemochromatosis, thalassemias, and Friedreich ataxia, as well as chronic inflammatory diseases of the liver, such as hepatitis C and steatohepatitis. The biochemical mechanisms by which iron causes toxicity are not completely known. Previously, we identified Poly rC-binding protein 1 (Pcbp1) as a protein that directly binds and delivers iron to ferritin, the major iron storage protein in mammalian cells. Pcbp1 and its paralog Pcbp2 are multifunctional adaptors that bind cellular RNA, DNA, proteins, and iron, altering the fate of their binding partners. Ferritin is a large polypeptide, containing 24 subunits of H- and L-chains assembled into a hollow sphere. Reduced (ferrous) iron enters the sphere through pores formed between the subunits and is oxidized on the interior surface to form nanocrystals of ferric oxyhydroxides. Pcbp1 is the first example of an iron chaperone- a protein that specifically binds iron ions and delivers them to target proteins, such as ferritin, through direct protein-protein interactions. In cultured cells, Pcbp1 is also required for efficient iron delivery to non-heme enzymes in addition to ferritin. These enzymes include the mononuclear iron-containing prolyl hydroxylases that regulate HIF and the dinuclear iron-containing deoxyhypusine hydroxylase, which is required for the modification of lysine to hypusine. In the adult human, 70% of total body iron is present in circulating erythrocytes, which are produced by the bone marrow at a rate of 2 million reticulocytes per second. We examined the roles of Pcbp1 and Ncoa4 in this extraordinary flux of iron through the erythron. Ncoa4 is the cargo receptor that recruits ferritin into the autophagosome for degradation in the lysosome. Using an in vitro model of erythroid differentiation, we showed that depletion of Pcbp1 or Ncoa4 impeded trafficking of iron through ferritin, which impaired the synthesis of heme and hemoglobin. Mice with tamoxifen-induced Pcbp1 deficiency exhibited a microcytic anemia typical of iron deficiency, with activation of compensatory erythropoiesis. The role of ferritin in erythropoiesis has been controversial, but our studies indicate that iron flux through ferritin is an obligatory process in the early stages of erythroid terminal differentiation. Our in vitro studies of erythrocytes revealed that the interactions of Pcbp1 and Ncoa4 with ferritin changed during differentiation, with maximal binding of Pcbp1 at early stages and Ncoa4 at late stages. These binding activities are regulated by cellular iron in mechanistically distinct ways. Ongoing studies of Pcbp1 deficiency in other murine tissues suggest the importance of iron chaperones in maintaining the bioavailable pool of iron in mammalian cells. Disclosures No relevant conflicts of interest to declare.

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