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D R Kiino

Publications and source records attributed to D R Kiino.

12 recordsLinked to original sources
PubMed1993

A cytoplasmic protein, NfrC, is required for bacteriophage N4 adsorption.

At least four genes are required for irreversible adsorption of bacteriophage N4. nfrA and nfrB have been characterized previously and encode an outer membrane protein and inner membrane protein, respectively. The nfrC gene product is characterized in detail in this study. We have mapped the nfrD locus to min 52 on the Escherichia coli linkage map. Maxicell analysis of nfrC and a null allele (nfrC2) cloned into a high-copy-number plasmid shows its gene product to be 42 kDa in size. We determined the nfrC nucleotide sequence which predicts a gene product of 42 kDa. Western blots (immunoblots) of Escherichia coli proteins after cellular fractionation show NfrC to be a cytoplasmic protein which is required for irreversible bacteriophage N4 adsorption, an event occurring at the cell surface.

Adsorption↗
PubMed1990

Increased expression of the bifunctional protein PrlF suppresses overproduction lethality associated with exported beta-galactosidase hybrid proteins in Escherichia coli.

We have cloned and determined the nucleotide sequence of the prlF gene. An open reading frame predicting a 111-amino-acid protein (Mr 12,351) with an acidic carboxy terminus was identified. The DNA sequence preceding this open reading frame revealed a putative promoter and a ribosome-binding site. The nucleotide sequence of the prlF1 mutation revealed a 7-base-pair duplication resulting in a slightly smaller predicted gene product of Mr 12,009 that lacked the acidic carboxy terminus. Maxicell analysis of prlF and prlF1 subclones identified peptides of sizes similar to those predicted by the nucleotide sequences. The prlF sequence was shown to be expressed in vivo by both maxicell analysis and construction of a prlF-lacZ fusion. Two kanamycin resistance insertions within the prlF open reading frame were introduced into the chromosome, replacing the wild-type gene. In contrast to the prlF1 mutation, these insertions had no detectable effect on cell growth or on the beta-galactosidase activity or maltose sensitivity (two sensitive indicators of hybrid protein export) conferred by the lamB-lacZ42-1 gene fusion. Overproduction of the wild-type prlF gene product from a plasmid carrying an active hybrid promoter, however, conferred a prlF1 phenotype. In addition, both the prlF1 mutation and both kanamycin resistance insertions increased the beta-galactosidase activity of a prlF-lacZ fusion. These results suggest that prlF is autoregulated and that overproduction of the prlF gene product increases the export efficiency of beta-galactosidase hybrid proteins from the cytoplasm.

Bacterial Proteins↗
PubMed1989

Genetic analysis of bacteriophage N4 adsorption.

We isolated six mutants of Escherichia coli K-12 that were defective in bacteriophage N4 adsorption. We mapped the mutations to four loci designated nfrA through nfrD (N four resistance). nfrA and nfrB were tightly linked to each other and were mapped to min 12 of the E. coli linkage map. nfrC was mapped to min 85, and nfrD was mapped between min 44 and 58. We isolated a clone carrying both nfrA and nfrB and identified its gene products through maxicell analysis of plasmid subclones. The nfrA gene product was an outer membrane protein of 96,000 apparent molecular weight, whereas nfrB encoded an inner-membrane protein of 69,500 apparent molecular weight. The nfrB1 mutation did not affect the export of the nfrA gene product to the outer membrane and did not affect the alkaline phosphatase activity of an nfrA-phoA fusion. We propose that nfrA encodes the structural receptor for N4 and that the nfrB gene product may be required for irreversible adsorption and injection of the phage genome and virion-encapsulated RNA polymerase through the inner membrane.

Alleles↗
PubMed1984

Mutation prlF1 relieves the lethality associated with export of beta-galactosidase hybrid proteins in Escherichia coli.

The 42-1 lamB-lacZ gene fusion confers a conditionally lethal, export-dependent phenotype known as maltose sensitivity. A maltose-resistant mutant showing decreased beta-galactosidase activity of the hybrid protein, designated prlF1 (protein localization), was unlinked to the lamB-lacZ fusion. This mutation mapped at 70 min on the Escherichia coli linkage map and conferred maltose resistance, a 30-fold reduction in beta-galactosidase activity, and a 30% decrease in cellular growth rate at 30 degrees C that was independent of the presence of a gene fusion. prlF1 also decreased the beta-galactosidase activity and relieved the maltose sensitivity conferred by fusions of lacZ to the gene specifying the periplasmic maltose-binding protein, malE. The decrease in beta-galactosidase activity, however, was specific for exported hybrid proteins. When export of the hybrid protein was blocked by a signal sequence mutation, prlF1 decreased the beta-galactosidase activity only 2.5-fold. Similarly, prlF1 did not affect the beta-galactosidase activity of fusions of lacZ to a gene specifying a nonexported protein, malK.

Bacteriophage lambda↗
PubMed1982

Prolactin storage in a clonal strain of rat pituitary tumor cells is cell-cycle dependent.

GH4C1 cells (CH cells) are a clonal strain of rat pituitary tumor cells which secrete prolactin. We measured intracellular prolactin at different stages of the cell cycle using flow microfluorometry. Prolactin was stained by an indirect immunocytochemical technique using fluorescein isothiocyanate (FITC)-conjugated antiserum, and DNA was stained simultaneously with propidium iodide. We found that prolactin storage in GH cells was cell-cycle dependent; prolactin storage increased as cells passed from G1 to S to G2 + M. We have shown previously that insulin and 17 beta-estradiol act synergistically to increase intracellular prolactin three- to sevenfold and slow the rate of cell growth to approximately 70% of control cells. In this study we observed that insulin and estradiol increased prolactin storage at each stage of the cell cycle but did not affect the cell-cycle distribution of the population even though cell growth was slowed. We conclude that insulin and estradiol did not increase prolactin storage by affecting the cell-cycle distribution of the population.

Animals↗
PubMed1981

Insulin and 17 beta-estradiol increase the intracellular prolactin content of GH4C1 cells.

We investigated the effects of insulin and 17 beta-estradiol in PRL-producing rat pituitary tumor cells (GH cells) in culture. Incubation with 1.8 X 10(-7) M insulin and 1 X 10(-9) M estradiol always increased intracellular PRL significantly to a mean of 290% of control levels. Insulin or estradiol alone caused smaller increases in intracellular PRL that were significant in five out of six experiments with insulin and three out of five experiments with estradiol, producing average increases of 150% and 160% of controls, respectively. Combined treatment with insulin and estradiol increased PRL secreted into the medium to 160% of controls, which was smaller than the effect on intracellular PRL. The effects of insulin and estradiol were detectable after 2-4 days of treatment and were maximal between 6-8 days. The effects were dose dependent; the half-maximal dose of insulin was about 2 X 10(-8) M and of estradiol was about 5 X 10(-11) M. Treatment with both insulin and estradiol increased cellular uptake of [3H]leucine by 45% compared to controls, incorporation of [3H]leucine into PRL by 65%, and total protein by 50%. The increased incorporation into PRL may represent an increase in PRL synthesis, but it is not enough to account for the increased intracellular hormone. Insulin and estradiol did not change the time required for most newly synthesized PRL to be processed and released from the cells, which was 60 min. Therefore, the amount of intracellular PRL was not increased as a result of an increased intracellular transit time. We conclude that insulin and estradiol increase the amount of intracellular PRL as a result of either decreasing the rate of degradation or increasing the cells' capacity to store PRL.

Animals↗
PubMed1975

Effects of extracellular cations and ouabain on catecholamine-stimulated sodium and potassium fluxes in turkey erythrocytes.

In turkey erythrocytes, potassium influx can be inhibited by several cations whose order of effectiveness is Rb greater than Cs greater than Li greater than Mg = Ca = Ba. Extracellular sodium does not alter potassium influx. Sodium influx is not altered by any of these monovalent cations but magnesium, calcium, or barium reduced sodium influx by 30 to 40%. Potassium outflux is not influenced by extracellular sodium or potassium while sodium outflux is not influenced by extracellular potassium but increases progressively with increasing extracellular sodium. Isoproterenol stimulates potassium influx only when sodium or lithium is present in the medium and catecholamine stimulation increases progressively with increasing extracellular sodium. Isoproterenol-stimulated sodium influx is enhanced by extracellular potassium, rubidium or cesium, and catecholamine stimulation increases progressively with increasing extracellular potassium. Isoproterenol inhibits potassium outflux in a solution free of sodium and potassium, and this inhibition can be abolished by adding sodium but not by adding potassium. In solutions containing both sodium and potassium, isoproterenol stimulates potassium outflux, and this stimulation increases progressively with increasing extracellular sodium or potassium. Isoproterenol-stimulated sodium outflux is not influenced by extracellular sodium or potassium. Isoproterenol-stimulated cellular cyclic adenosine 3':5'-monophosphate (cyclic AMP) is reduced slightly (25%) by removing extracellular sodium and potassium from the incubation solution, but this effect is not of sufficient magnitude to account for the effects of these cations on isoproterenol-stimulated cation transport. Ouabain does not alter the effect of extracellular sodium or potassium on isoproterenol-stimulated potassium influx; however, the glycoside potentiates (by 20 to 40%) the effects of these two cations on isoproterenol-stimulated sodium influx. Ouabain does not alter potassium outflux when the incubation solution contains both sodium and potassium; however, ouabain stimulates potassium outflux in an incubation solution free of sodium or potassium.

Animals↗
PubMed1975

Refractoriness of cation transport in turkey erythrocytes to stimulation by cyclic adenosine 3':5'-monophosphate.

In turkey erythrocytes bidirectional fluxes of sodium and potassium develop a time-dependent refractoriness to stimulation by endogenous cyclic adenosine 3':5'-monophosphate (cyclic AMP). The refractoriness of potassium influx and potassium outflux (both of which require extracellular sodium and potassium for stimulation by cyclic AMP) depends on the extracellular concentrations of sodium and potassium. In contrast, the refractoriness developed by sodium outflux (which does not require extracellular sodium or potassium for stimulation by cyclic AMP) does not depend on the extracellular concentrations of sodium or potassium. The refractoriness of these fluxes to cellular cyclic AMP reflects a decrease in the amount by which they can be maximally stimulated and appears to be proportional to the extent to which the transport system is utilized during the course of the incubation. Ouabain significantly reduces the rate at which cation transport in turkey erythrocytes becomes refractory to endogenous cyclic AMP. This effect of the glycoside is independent of the extracellular concentrations of sodium or potassium and does not correlate with how it alters the initial response of the transport systems to cyclic AMP.

Animals↗
PubMed1975

Effects of beta-adrenergic catecholamines on potassium transport in turkey erythrocytes.

Isoproterenol stimulates cellular accumulation of cyclic adenosine 3':5'-monophosphate (cyclic AMP) and produces a 2- to 4-fold increase in bidirectional potassium fluxes in turkey erythrocytes. Ouabain, which does not alter catecholamine-stimulated cellular cyclic AMP, inhibits potassium influx by 50 to 70%, does not alter potassium outflux or isoproterenol-stimulated potassium influx, but increases isoproterenol-stimulated potassium outflux. Stimulation of potassium transport by isoproterenol can be reproduced by adding cyclic AMP to the medium and is inhibited by propranolol or dichloroisoproterenol but not by phentolamine. Theophylline at concentrations which inhibit cyclic nucleotide phosphodiesterase in isolated turkey erythrocyte plasma membranes by greater than 90%, does not augment isoproterenol stimulation of cellular cyclic AMP or of potassium transport but does potentiate stimulation of potassium influx produced by adding cyclic AMP to the medium. Isoproterenol-stimulated cellular cyclic AMP increases steadily for at least 2 hours. Potassium transport, however, increases rapidly, becomes maximal after 20 to 30 min of incubation, and thereafter decreases progressively so that after 2 hours of incubation potassium fluxes are only slightly greater than for the control. Ouabain prolongs the duration of catecholamine-stimulated potassium influx and potassium outflux, reflecting its ability to relieve the refractoriness developed by turkey erythroyctes to endogenous cyclic AMP.

Adrenergic beta-Antagonists↗
PubMed1973

Ouabain binding and cation transport in human erythrocytes.

In the present studies we have explored the relation between ouabain binding and the inhibition of potassium influx in intact human erythrocytes. The rate at which bound ouabain molecules dissociate from the erythrocyte membrane is not altered by complete replacement of choline with sodium or by partial replacement with potassium. These findings indicate that the effects of these cations on ouabain binding reflect alterations in the rate of association of ouabain molecules with the erythrocyte membrane. Variations in the cation composition of the incubation solution did not alter the relation between the fraction of the glycosidebinding sites occupied by ouabain or the fraction of ouabain-sensitive potassium influx which was inhibited. That is, irrespective of the affinity of the erythrocyte membrane for ouabain molecules and irrespective of the magnitude of glycoside-sensitive potassium influx, occupation of a given fraction of the glycoside-binding sites by ouabain results in the inhibition of an equal fraction of the ouabain-sensitive potassium transport sites.

Adult↗
PubMedDate not supplied

Hormonal regulation of prolactin storage in a clonal strain of rat pituitary tumor cells.

GH4C1 cells (GH cells) are a clonal strain of rat pituitary tumor cells which secrete prolactin. GH cells have been used to study hormone secretion, but they store relatively little prolactin compared to normal prolactin-secreting cells. They are not suitable, therefore, for studying some aspects of pituitary function. We have found that the amount of prolactin GH cells store can be regulated. When GH cells were plated at 10(6) cells/well and treated for six days with 180 nM insulin or 1 nM estradiol, there was a 60 percent increase in prolactin storage compared to control cells. Insulin and estradiol in combination acted synergistically to cause a 190 percent increase in prolactin storage. In contrast, they were additive in increasing extracellular prolactin; there was a 40 percent increase in extracellular prolactin after insulin, a 20 percent increase after estradiol, and a 50 percent increase after insulin plus estradiol. The increases in prolactin storage were always greater than the increases in extracellular prolactin. The increases in prolactin storage were dose-dependent and reached maximal levels after four days of treatment with 180 nM insulin plus 1 nM estradiol. Reducing the plating density to 10(3) cells/well increased the response to insulin and estradiol to nineteenfold. Epidermal growth factor (10 nM) acted synergistically with estradiol and insulin in combination to increase prolactin storage 27-fold. The insulin- and estradiol-induced increase in extracellular prolactin was caused by a specific increase in the rate of prolactin synthesis. The fractional increase in prolactin storage above the increase in prolactin production could not be explained by an increase in prolactin synthesis, an increase in intracellular transit time, or a change in the cell-cycle distribution of the population. Hormone storage can, therefore, be regulated independently from other processes which control hormone production. The prolactin stored in response to insulin and estradiol was releasable by potassium depolarization. Following depletion of intracellular prolactin by depolarization, the cells retained their increased capacity for prolactin storage. The ability to increase prolactin storage will make GH cells a more useful system in which to study pituitary function.

Animals↗