Biomedical subjects

F L Crane

Publications and source records attributed to F L Crane.

At least 37 records · Page 2Linked to original sources

PubMed1994

Iron at the cell surface controls DNA synthesis in CCl 39 cells.

Treatment of CCl 39 cells with the impermeable iron II chelator bathophenanthroline disulfonate (BPS) inhibits both DNA synthesis and transplasma membrane electron transport. The inhibition persists when the BPS is removed, and the extract from 10(6) cells contains up to 1.28 nmoles iron II chelated to BPS. The BPS iron II chelate itself is not inhibitory. Both DNA synthesis and electron transport are restored by addition of microM iron II or iron III compounds to extracted cells. Other impermeable chelators for iron II give similar inhibition, whereas the iron III-specific Tiron or copper-specific bathocuproine sulfonate do not inhibit. The inhibition differs from the permeable iron III chelator inhibition of ribonucleotide reductase, because inhibition of DNA synthesis by the permeable chelators is reversed when chelator is removed. The response to growth factors also differs, with no impermeable chelator inhibition on 10% fetal calf serum contrasting to inhibition by permeable chelators. DNA synthesis with both activation of tyrosine kinase with EGF plus insulin or by thrombin or ceruloplasmin led to protein kinase C activation as inhibited by the impermeable chelators. It is proposed that an iron available on the cell surface is required for DNA synthesis and plasma membrane electron transport.

Animals↗

PubMed1994

Iron reverses impermeable chelator inhibition of DNA synthesis in CCl 39 cells.

Treatment of Chinese hamster lung fibroblasts (CCl 39 cells) with the impermeable iron(II) chelator bathophenanthroline disulfonate (BPS) inhibits DNA synthesis when cell growth is initiated with growth factors including epidermal growth factor plus insulin, thrombin, or ceruloplasmin, but not with 10% fetal calf serum. The BPS treatment inhibits transplasma membrane electron transport. The treatment leads to release of iron from the cells as determined by BPS iron(II) complex formation over 90 min. Growth factor stimulation of DNA synthesis and electron transport are restored by addition of di- or trivalent iron to the cells in the form of ferric ammonium citrate, ferrous ammonium sulfate, or diferric transferrin. The effect with BPS differs from the inhibition of growth by hydroxyurea, which acts on the ribonucleotide reductase, or diethylenetriaminepentaacetic acid, which is another impermeable chelating agent, in that these agents inhibit growth in 10% fetal calf serum. The BPS effect is consistent with removal of iron from a site on the cell surface that controls DNA synthesis.

Animals↗

PubMed1994

Bombesin stimulates transplasma-membrane electron transport by Swiss 3T3 cells.

Bombesin, a mitogenic neuropeptide, stimulates transplasmalemma reduction of diferric transferrin or ferricyanide by Swiss 3T3 cells. The stimulation of diferric transferrin reduction occurs in the range of bombesin concentrations that stimulate proliferation of Swiss 3T3 cells. Diferric transferrin reduction by the 3T3 cells is accompanied by increased proton release from the cells and bombesin increases the differic transferrin-stimulated proton release twofold. Insulin increases the diferric transferrin reductase response and increases growth stimulation with bombesin. The effect of bombesin on the transmembrane electron transport is a new aspect of its effect on the plasma membrane in addition to increase in phosphatidylinositol turnover and protein kinase c activation. The electron transport can provide an independent mechanism of activation of the Na+/H+ exchange or it can change the redox state of pyridine nucleotide in the cytoplasm.

3T3 Cells↗

PubMed1994

Coenzyme Q10, plasma membrane oxidase and growth control.

The plasma membrane of eukaryotic cells contains an NADH oxidase which can transfer electrons across the membrane. This oxidase is controlled by hormones, growth factors and other ligands which bind to receptors in the plasma membrane. Oncogenes also affect activity of the oxidase. Natural serum components such as diferric transferrin and ceruloplasmin which stimulate proliferation also stimulate membrane oxidase activity. Additional growth factors can be required to complement the proliferative effect. Electron transport across the plasma membrane can be measured by the reduction of impermeable electron acceptors, such as ferricyanide, which also stimulate cell growth. The oxidants activate growth-related signals such as cytosolic alkalinization and calcium mobilization. Antiproliferative agents such as adriamycin and retinoic acid inhibit the plasma membrane electron transport. Flavin, Coenzyme Q and an iron chelate on the cell surface are apparent electron carriers for the transmembrane electron transport. Coenzyme Q10 stimulates cell growth, and Coenzyme Q analogs such as capsaicin and chloroquine reversibly inhibit both growth and transmembrane electron transport. Addition of iron salts to the depleted cells restores activity and growth. The ligand-activated oxidase in the plasma membrane introduces a new basis for control of signal transduction in cells. The redox state of the quinone in the oxidase is proposed to control tyrosine kinase either by generation of H2O2 or redox-induced conformational change.

Animals↗

PubMed1994

Iron chelators hydroxyurea and bathophenanthroline disulfonate inhibit DNA synthesis by different pathways.

We previously showed that thrombin-stimulated DNA synthesis in CCL 39 cells was inhibited by hydroxyurea (HU) and bathophenanthroline disulfonate (BPS) (Proc. Natl. Acad. Sci. USA, in press). A clear difference exists between these two inhibitors. Inhibition mediated by HU was immediate and must be present in the culture medium. BPS was equally effective when it was present in the medium or after preincubation, but it required at least 12 h to achieve maximal effect. The permeable form 1,10 phenanthroline had the same inhibitory effect in short-term incubations that BPS. Moreover, 1,10 phenanthroline was cytotoxic in long-term incubations indicating that the site of BPS inhibition was outside the cell. Further, long-term incubations with HU did not affect the ability of the cell to reinitiate DNA synthesis after removal of the chelator.

Animals↗

PubMed1993

Ha-ras stimulates the transplasma membrane oxidoreductase activity of C3H10T1/2 cells.

The ras family of oncogenes, as well as the transplasma membrane electron transport system, have been implicated in cellular growth control. Here we present evidence that the Ha-ras oncoprotein increases the activity of the transplasma membrane electron transport system in the mouse embryonic fibroblast cell line C3H10T1/2. Both cytochrome c and ferricyanide are reduced at a faster rate by C3H10T1/2 cells which are expressing the Ha-ras oncogene. In addition, the Ha-ras transformed cells extrude protons faster in response to stimulation by external oxidants than their normal counterparts. These results suggest that transformation events initiated by the expression of the Ha-ras oncoprotein, p21, in C3H10T1/2 cells have a controlling effect on the transplasma membrane electron transport system.

Animals↗

PubMed1993

NADH oxidase activity of rat liver plasma membrane activated by guanine nucleotides.

The activity of a hormone- and growth-factor-stimulated NADH oxidase of the rat liver plasma membrane responds to guanine nucleotides, but in a manner that differs from that of the classic trimeric and low-molecular-mass monomeric G-proteins. In the absence of added bivalent ions, both GTP and GDP as well as guanosine 5'-[gamma-thio]triphosphate (GTP[gamma-S]) but not guanosine 5'[beta-thio]diphosphate (GDP[beta-S]) stimulate the activity over the range 1 microM to 100 microM. Other di- and tri-nucleotides also stimulate, but only at concentrations of 100 microM or higher. Added bivalent ions are not required either for NADH oxidation or guanine nucleotide stimulation. Bivalent ions (Mg2+ > Mn2+ > or = Ca2+) alone stimulate only slightly at low concentrations and then inhibit at high concentrations. The inhibitions are augmented by GDP or GTP [gamma-S] but not by GTP. Although the activity is the same, or less, in the presence of 0.5 mM MgCl2, GTP at 1-100 nM and other nucleotides at 0.1 mM or 1 mM still stimulate in its presence. The NADH oxidase is activated by mastoparan but aluminum fluoride is weakly inhibitory. Cholera and pertussis toxins elicit only marginal responses. Both the Mg2+ and the GDP and GTP[gamma-S] inhibitions (but not the GTP stimulations) shift to higher concentrations when the membrane preparations are first solubilized with Triton X-100. The results suggest a role for guanine nucleotides in the regulation of plasma membrane NADH oxidase, but with properties that differ from those of either trimeric or the low-molecular-mass G proteins thus far described.

Animals↗

PubMed1993

The essential functions of coenzyme Q.

The essential role of coenzyme Q in biological energy transduction is well established. Coenzyme Q is a unique carrier for two-electron transfer within the lipid phase of the mitochondrial membrane. The function is essential for proton-based energy coupling. The sites of entry and exit of electrons into the quinone are at specific quinone-binding sites which are constructed to allow only two-electron transfer and thus prevent damaging free radical formation by direct reaction of oxygen with the semiquinone. Failure of proper function with diminished energy supply can be related to insufficient quinone, modification of lipid fluidity, or lipid protein interaction and damage or poisoning in binding sites. Supplementation with coenzyme Q can act by reversal of deficiency or decreased mobility, or by overcoming binding site modification. Coenzyme Q has also been shown to increase antioxidant protection in membranes. New sites for coenzyme Q function in Golgi and plasma membrane show evidence for a role in growth control and secretion-related membrane flow.

Animals↗

PubMed1993

The effect of boron on plasma membrane electron transport and associated proton secretion by cultured carrot cells.

Plasma membrane electron transport reactions and associated proton secretion were studied in boron-deficient carrot cells. It was found that the hormone-sensitive plasma membrane NADH oxidase was inhibited by boron deficiency and that under such conditions activity could be restored by exogenous boric acid with or without 2,4-dichlorophenoxy acetic acid. Gramicidin, a channel-forming protonophore, further stimulated NADH oxidase by carrot cells. Proton secretion, associated with plasma membrane H(+)-ATPase, was also affected by boron deficiency, but not as severely as ferricyanide-generated proton secretion, reflecting plasma membrane electron transport. The addition of 1 mM boric acid and 1 microM 2,4-dichlorophenoxy acetic acid to carrot cells fully restored the H+ secretion in presence of ferricyanide. The effect of boron deficiency in cultured carrot cells can, therefore, be directly associated with cell growth through its effect on the plasma membrane NADH oxidase and H+ secretion. Ferricyanide provides a probe which activates transmembrane electron transport that is only coupled to proton release when boron is present.

2,4-Dichlorophenoxyacetic Acid↗

PubMed1992

Requirement for coenzyme Q in plasma membrane electron transport.

Coenzyme Q is required in the electron transport system of rat hepatocyte and human erythrocyte plasma membranes. Extraction of coenzyme Q from the membrane decreases NADH dehydrogenase and NADH:oxygen oxidoreductase activity. Addition of coenzyme Q to the extracted membrane restores the activity. Partial restoration of activity is also found with alpha-tocopherylquinone, but not with vitamin K1. Analogs of coenzyme Q inhibit NADH dehydrogenase and oxidase activity and the inhibition is reversed by added coenzyme Q. Ferricyanide reduction by transmembrane electron transport from HeLa cells is inhibited by coenzyme Q analogs and restored with added coenzyme Q10. Reduction of external ferricyanide and diferric transferrin by HeLa cells is accompanied by proton release from the cells. Inhibition of the reduction by coenzyme Q analogs also inhibits the proton release, and coenzyme Q10 restores the proton release activity. Trans-plasma membrane electron transport stimulates growth of serum-deficient cells, and added coenzyme Q10 increases growth of HeLa (human adenocarcinoma) and BALB/3T3 (mouse fibroblast) cells. The evidence is consistent with a function for coenzyme Q in a trans-plasma membrane electron transport system which influences cell growth.

Cell Division↗

PubMed1992

Stimulation of serum-free cell proliferation by coenzyme Q.

Coenzyme Q added to culture media stimulates the growth of HeLa and Balb/3T3 cells in serum free conditions. The stimulation by coenzyme Q is additive to the stimulation by ferricyanide, an impermeable electron acceptor for the transplasma membrane electron transport. alpha Tocopherylquinone can also stimulate cell growth, but vitamin K1 is inactive or inhibitory. The response to coenzyme Q and ferricyanide is enhanced with insulin. A contribution to plasma membrane NADH oxidation or modification of the membrane quinone redox balance can be a basis for the growth stimulation.

3T3 Cells↗

PubMed1992

Ceruloplasmin stimulates NADH oxidation of pig liver plasma membrane.

NADH oxidation by pig liver plasma membranes is stimulated by ceruloplasmin (CUP) reaching a maximal value at 50 U/ml of CUP. NADH oxidation activated by CUP is proportional to the amount of protein. Concanavalin A (Con A) which recognizes the glucidic residues of the CUP required for binding to the receptor inhibits the NADH oxidation in a dose-responsive manner. Both adriamycin and bathophenantroline disulfonate (BPS), previously reported as transplasma membrane electron transport inhibitors, also inhibit the CUP-stimulated NADH oxidation of pig liver plasma membranes. Our results show a clear interaction between CUP and the NADH oxidase of plasma membrane, which supports an oxidative role for CUP in its growth effect.

Animals↗

PubMed1992

Inhibition of transplasma membrane electron transport by transferrin-adriamycin conjugates.

Transplasma membrane electron transport from HeLa cells, measured by reduction of ferricyanide or diferric transferrin in the presence of bathophenanthroline disulfonate, is inhibited by low concentrations of adriamycin and adriamycin conjugated to diferric transferrin. Inhibition with the conjugate is observed at one-tenth the concentration required for adriamycin inhibition. The inhibitory action of the conjugate appears to be at the plasma membrane since (a) the conjugate does not transfer adriamycin to the nucleus, (b) the inhibition is observed within three minutes of addition to cells, and (c) the inhibition is observed with NADH dehydrogenase and oxidase activities of isolated plasma membranes. Cytostatic effects of the compounds on HeLa cells show the same concentration dependence as for enzyme inhibition. The adriamycin-ferric transferrin conjugate provides a more effective tool for inhibition of the plasma membrane electron transport than is given by the free drug.

Animals↗

PubMed1992

A growth factor- and hormone-stimulated NADH oxidase from rat liver plasma membrane.

NADH oxidase activity (electron transfer from NADH to molecular oxygen) of plasma membranes purified from rat liver was characterized by a cyanide-insensitive rate of 1 to 5 nmol/min per mg protein. The activity was stimulated by growth factors (diferric transferrin and epidermal growth factor) and hormones (insulin and pituitary extract) 2- to 3-fold. In contrast, NADH oxidase was inhibited up to 80% by several agents known to inhibit growth or induce differentiation (retinoic acid, calcitriol, and the monosialoganglioside, GM3). The growth factor-responsive NADH oxidase of isolated plasma membranes was not inhibited by common inhibitors of oxidoreductases of endoplasmic reticulum or mitochondria. As well, NADH oxidase of the plasma membrane was stimulated by concentrations of detergents which strongly inhibited mitochondrial NADH oxidases and by lysolipids or fatty acids. Growth factor-responsive NADH oxidase, however, was inhibited greater than 90% by chloroquine and quinone analogues. Addition of coenzyme Q10 stimulated the activity and partially reversed the analogue inhibition. The pH optimum for NADH oxidase was 7.0 both in the absence and presence of growth factors. The Km for NADH was 5 microM and was increased in the presence of growth factors. The stoichiometry of the electron transfer reaction from NADH to oxygen was 2 to 1, indicating a 2 electron transfer. NADH oxidase was separated from NADH-ferricyanide reductase, also present at the plasma membrane, by ion exchange chromatography. Taken together, the evidence suggests that NADH oxidase of the plasma membrane is a unique oxidoreductase and may be important to the regulation of cell growth.

Animals↗

PubMed1992

Destruction of vitamin K1 of cultured carrot cells by ultraviolet radiation and its effect on plasma membrane electron transport reactions.

The effect of ultraviolet radiation on plasma membrane electron transport reactions was studied in cultured carrot cells. It was found that a 90 min treatment inhibited transmembrane hexacyanoferrate reduction greater than 50%. Extraction of lipophilic quinones from irradiated cells showed that vitamin K1 and coenzyme Q were totally destroyed, while control unirradiated cells showed the presence of 0.4 mumole vitamin K1 g dry wt.-1. The addition of exogenous vitamin K1 in concentrations of 1-10 microM partially restored plasma membrane electron transport with impermeable hexacyanoferrate as the electron acceptor. Total restoration of activity was given by growing irradiated cells in vitamin K1 supplemented growth media for 6 days. This shows that vitamin K1 may function as a member of the transplasma membrane electron transport chain in cultured carrot cells.

Cell Membrane↗

PubMed1991

Ceruloplasmin stimulates thymidine incorporation by CCL-39 cells in the absence of serum or growth factors.

The incorporation of tritiated thymidine into CCL-39 cells grown in the absence of fetal calf serum or other growth factors is greatly increased by low concentrations of ceruloplasmin. The stimulation is greater than observed with serum or thrombin. Addition of serum decreases the thymidine incorporation with ceruloplasmin to the level with serum alone. As with serum, the response to ceruloplasmin is high at both 20% and 1% oxygen, which is consistent with the action of ceruloplasmin as an oxidant with a high affinity for oxygen. Since transplasma membrane electron transport increases cell growth and thymidine incorporation, ceruloplasmin may act as a terminal oxidase for ferrous iron or ascorbate to stimulate transplasma membrane electron transport. The four electron transfer from ceruloplasmin to oxygen to form water will prevent peroxide formation at the cell surface. Alternatively, superoxide formation inside the cell or membrane could employ the superoxide dismutase function of ceruloplasmin to produce peroxide. Either mechanism would be consistent with the previously described stimulation of growth by external oxidants.

3T3 Cells↗

PubMed1991

Chloroquine-sensitive transplasmalemma electron transport in Tetrahymena pyriformis: a hypothesis for control of parasite protozoa through transmembrane redox.

Plasma membrane electron transport was studied in a protozoan cell, Tetrahymena pyriformis, by assaying transmembrane ferricyanide reduction and the reduction of iron compounds. The rates of ferricyanide reduction varied between 0.5 and 2.5 mumol/g dry wt. per min, with a pH optimum at 7.0-7.5. Other active non-permeable electron acceptors, with redox potentials from +360 to -125 mV, were cytochrome c, hexaammine ruthenium chloride, ferric-EDTA, ammonium ferric citrate, and indigo di-, tri- and tetrasulfonates. It was found that Tetrahymena cells can reduce external electron acceptors with redox potentials at pH 7.0 down to -125 mV. Ferricyanide stimulates ciliary action. Transmembrane ferricyanide reduction by Tetrahymena was not inhibited by such mitochondrial inhibitors as antimycin A, 2-n-heptyl-4-hydroxyquinoline N-oxide, or potassium cyanide, but it responded to inhibitors of glycolysis. Transmembrane ferricyanide reduction by Tetrahymena appears to involve a plasma membrane electron transport chain similar to those of other animal cells. As in other cells, the transmembrane electron transport is associated with proton release which may be involved in internal pH control. The transmembrane redox system differs from that of mammalian cells in a 20-fold greater sensitivity to chloroquine and quinacrine. The Tetrahymena ferricyanide reduction is also inhibited by chlorpromazine and suramin. Sensitivity to these drugs indicates that the transplasma membrane electron transport and associated proton pumping may be a target for drugs used against malaria, Trypanosomes and other protozoa.

Animals↗

PubMed1991

Inhibition of transplasma membrane electron transport by monoclonal antibodies to the transferrin receptor.

Reduction of iron in diferric transferrin is inhibited by monoclonal antibodies to the transferrin receptor which bind at sites other than the high affinity transferrin binding site. These antibodies include B3/25, GB16 and GB22. Two antibodies which bind at the high affinity site for transferrin, 42/6 and GB18, do not inhibit iron reduction by transplasma membrane electron transport. The results are consistent with the proposal that differric transferrin reduction or stimulation of transmembrane NADH oxidase activity involves a site different from the high affinity diferric transferrin binding site. A synergistic action of antibodies with epitopes at the tight binding site involved in iron uptake and the antibodies which inhibit electron transport, B3/25 and GB16, can explain the increased inhibition of growth observed when both 42/6 and B3/25 are added to proliferating cells.

Animals↗