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M Nilsen-Hamilton

Publications and source records attributed to M Nilsen-Hamilton.

49 records · Page 3Linked to original sources
PubMed1981

Insulin and growth factors stimulate rapid posttranslational changes in glucose transport in ovarian granulosa cells.

The glucose analogues, 3-O-methyl-D-glucose and 2-deoxy-D-glucose, have been used to characterize glucose transport and its regulation by serum and growth factors in monolayer cultures of granulosa cells obtained from bovine ovaries. Uptake of 3-O-methylglucose was shown to be independent of the Na+-gradient, independent of energy, did not show accelerated exchange, and was stereospecific. Serum withdrawal resulted in a biphasic decrease in initial rates of glucose uptake with half-times for the two phases of 50 minutes and 3 hours. Insulin could prevent the decrease in uptake rates with a half-maximum concentration of 10.0 1/8 3 nM. Insulin was shown to stimulate DNA synthesis with a concentration of half-maximum response of 28 nM. Insulin or serum stimulation of 3-O-methylglucose uptakes in serum-starved cells resulted in a two threefold increase in initial rates, with a time for half-maximum stimulation of 3 minutes. The insulin-stimulated increase was insensitive to cycloheximide and cyanide during the first 30 minutes, and this early, rapid stimulation was also produced by brain FGF (fibroblast growth factor), pituitary FGF, epidermal growth factor, calf serum, and some but not all samples of follicular fluid. Insulin also stimulated 2-deoxyglucose and a-aminoisobutyric acid uptake during the first 5 minutes of addition and these early stimulations were shown to be posttranslational changes.

Animals↗
PubMed1979

Inhibition of alpha-aminoisobutyric acid transport in membrane vesicles from mouse fibroblasts after phosphorylation by cyclic AMP-dependent protein kinase.

Cyclic AMP-dependent protein kinases from several mammalian sources inhibit Na+-dependent alpha-aminoisobutyric acid transport by membrane vesicles isolated from 3T3 cells. Evidence is provided that phosphorylation of membrane proteins by the enzyme is responsible for the inhibition. Lysis of the vesicles, or a reduction in the intravesicular volume is not the cause of reduced transport. The cyclic AMP-dependent protein kinase and its catalytic subunit phosphorylate a number of membrane proteins. Most of these proteins are phosphorylated, but to a lesser extent in the absence of protein kinase or cyclic AMP. The phosphorylated proteins remain associated with the membranes during hypotonic lysis treatments, which would be expected to release intravesicular contents and loosely associated membrane proteins. 32P-labeled bands detected on sodium dodecyl sulfate polyacrylamide gels after phosphorylation of membranes by the catalytic subunit of the cyclic AMP-dependent kinase are eliminated by treatment with either pronase or 1 N NaOH, but not by ribonuclease nor by phospholipase C. The stability of the incorporated radioactivity to hot acid and hydroxylamine relative to hot base suggests that most of the 32P from [gamma-32P]ATP is incorporated into protein phosphomonoester linkages.

Adenosine Triphosphate↗
PubMed1979

Fibroblast growth factor causes an early increase in phosphorylation of a membrane protein in quiescent 3T3 cells.

THE regulatory systems controlling cell division have not been identified, but it has been shown that growth factors such as epidermal growth factor, fibroblast growth factor (FGF), and serum initiate rapid changes in cellular metabolism, probably involving post-transcriptional control mechanisms(1-5). As phosphorylation has been shown to be an important regulatory mechanism in several metabolic pathways, we initiated experiments to determine whether factors which stimulate DNA synthesis also stimulate endogenous phosphorylation. We find mat, within 5 min of addition of FGF or serum to (32)P-labelled Swiss 3T3 cells, there is a specific increase in the phosphorylation of a membrane protein with an apparent molecular weight of 33,000. Experiments with isolated cell fractions demonstrate that the phosphorylation of this protein is stimulated by cyclic AMP. This rapid and specific response to mitogens raises the possibility that this phosphorylation might be part of the initial, cellular signal for DNA synthesis.

Journal Article↗
PubMed1979

Serum-dependent regulation of alpha-aminoisobutyric acid uptake in bovine granulosa cells.

The removal of serum from the medium of ovarian granulosa cells in exponential or confluent stages of growth results in a rapid and pronounced decrease in the rate of transport of the non-metabolizable amino acid, alpha-aminoisobutyric acid. This decrease is rapidly and completely reversed by the addition of serum. The decrease and its reversal are insensitive to inhibitors of RNA and protein syntheisis and are unaffected by a number of other metabolic inhibitors. The serum requirement cannot be replaced by peptide hormones known to stimulate cell division and secretion by these cells. These data are consistent with a model of post-translational control of AIB transport by a high-molecular-weight component of serum.

Aminoisobutyric Acids↗
PubMed1978

Transport of phosphate in membrane vesicles from mouse fibroblasts transformed by simian virus 40.

Membrane vesicles were prepared from mouse fibroblasts transformed by SV40 virus (SV3T3). Following disruption of the cells by nitrogen cavitation, the membrane vesicles were obtained by differential centrifugation. As measured by enzyme markers, they consist mainly of membrane from the plasma membrane and smooth and rough endoplasmic reticulum. The vesicles transport Pi by two separate, mediated systems: one is independent of Na+, and the other is secondary active transport driven by a Na+ gradient. Electrical and chemical energy can be provided by a Na+ gradient to drive the concentrative uptake of Pi by the vesicles, one or both forces being used to energize transport. Evidence is provided that both the electrical and chemical potentials produced by the asymmetric distribution of Na+ across the membrane of SV3T3 membrane vesicles are utilized to concentrate phosphate in the vesicles. Phosphate transport by the vesicles cannot be accounted for by a small contamination of this fraction with mitochondria (1 to 4%). The Pi transport properties of the membrane vesicles differ from those of the fraction enriched in mitochondria in the following respects: their kinetic properties, and their responses to a Na+ gradient, N-ethylmaleimide, mersalyl, and succinate/acetate. However, the membrane vesicles share some properties of Pi transport with mitochondria. Cyanide, azide, oligomycin, 2,4-dinitrophenol, and carbonyl cyanide m-cholophenylhydrazone, inhibitors of Pi transport by mitochondria, also inhibit membrane vesicle, Pi transport. The vesicles retain all the features of Pi transport by SV3T3 cells that have been examined. They provide a simplified system for a determination of the details of the mechanism of Pi transport under conditions where transport is dissociated from intracellular reactions and in the presence of a defined electrochemical driving force.

Biological Transport, Active↗
PubMed1977

Preliminary characterization of two thymus-dependent xenoantigens from mouse lymphocytes.

Preliminary characterization of two mouse thymus-dependent (T) lymphocyte xenoantigens, T25 and T200, which are selectively labelled by lactoperoxidase-catalysed iodination of T-cells, is described. Both molecules are membrane-bound glycoproteins. Fractionation of membrane vesicles prepared from BW5147 lymphoma cells by sedimentation through sucrose density gradients show that antigens T25 and T200 are in fractions enriched with plasma membrane. Moreover antigen T200 is partially degraded when viable cells are treated briefly with low concentrations of trypsin. Both molecules are efficiently solubilized in buffers containing sodium deoxycholate or Nonidet P-40, as measured by failure to sediment at 100000g for 60min. However, gel filtration on Sepharose 6B showed the presence of aggregated material in Nonidet P-40 extracts which was not found in deoxycholate-solubilized membranes. After solubilization in detergent, antigens T25 and T200 bind to, and may be specifically eluted from, columns of pea lectin--Sepharose or concanavalin A--Sepharose. Both molecules are heterogeneous when examined by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. As judged by its binding to columns of pea lectin, at least part of the heterogeneity of mouse thymocyte antigen T25 resides in its carbohydrate moiety.

Animals↗
PubMed1976

The subcellular distribution of adenylate and guanylate cyclases in murine lymphoid cells.

Membrane vesicles can be prepared from murine lymphoid cells by nitrogen cavitation and fractionated by sedimentation through nonlinear sucrose density gradients. Two subpopulations of membrane vesicles, PMI and PMII, can be distinguished on the basis of sedimentation rate. The subcellular distribution of adenylate and guanylate cyclases in these membrane subpopulations have been compared with the distribution of a number of marker enzymes. Approximately 20-30% of the total adenylate and guanylate cyclase activity is located at the top of the sucrose gradient (soluble enzyme), the remainder of the activity being distributed in the PMI and PMII fractions (membrane-bound enzyme). More than 90% of the 5'-nucleotidase and NADH oxidase activities detected in lymphoid cell homogenates are located in PMI and PMII fractions, whereas succinate cytochrome c reductase activity is detected only in the PMII fractions. In addition, beta-galactosidase activity is distributed in the soluble and PMII fractions of the sucrose density gradients. On the basis of the fractionation patterns of these various enzyme activities, it appears that PMI fractions contain vesicles of plasma membrane and endoplasmic reticulum, whereas PMII fractions contain mitochondria, lysomes, and plasma membrane vesicles. Approximately 30-40% of the adenylate and guanylate cyclase activities in PMII can be converted to a PMI-like form following dialysis and resedimentation through a second nonlinear sucrose gradient. Adenylate and guanulate cyclases can be distinguished on the basis of sensitivity to nonionic detergents.

Adenylyl Cyclases↗
PubMed1976

Uptake of alpha-aminoisobutyric acid and phosphate by membrane vesicles derived from growing and quiescent fibroblasts.

Membrane vesicles derived principally from the plasma membrane and endoplasmic reticulum of mouse 3T3 cells transformed by Simian virus 40 take up alpha-aminoisobutyric acid (AIB) and phosphate (Pi). When NaCl is added simultaneously with AIB or Pi, uptake rises two- to three-times above the equilibrium to accumulate AIB or Pi over the control value, in the presence of a Na+ gradient, is almost lost in membrane vesicles derived from benzpyrene-transformed 3T3 cells (BP3T3) arrested in the G1 phase of the cell cycle by serum starvation. When added to the membranes with NaCl and the uptake substrate, a combination of fibroblast growth factor (FGF) and epidermal growth factor EGF restores the ability of the membranes to accumulate AIB and Pi over the control value.

Aminoisobutyric Acids↗
PubMed1976

Sodium-stimulated alpha-aminoisobutyric acid transport by membrane vesicles from simian virus-transformed mouse cells.

Uptake of alpha-aminoisobutyric acid, by membrane vesicles derived principally from the plasma membrane and endoplasmic reticulum of mouse 3T3 cells transformed by simian virus 40, is stimulated by sodium chloride. Both in the presence and absence of Na+ uptake is time-dependent and osmotically sensitive. The Na+-stimulated uptake is inhibited by other amino acids. The kinetics of transport of alpha-aminoisobutyric acid are shown to be biphasic both in whole cells and in the membrane vesicles. Only the high affinity system is stimulated by sodium in the membrane vesicles. These results demonstrate that observations made on living cells correlate with observations made on isolated membrane vesicles, and indicate that these membrane vesicles have retained the cellular amino acid transport system functionally intact.

Amino Acids↗