Search PubMed⌕ Search

Biomedical subjects

D E Buetow

Publications and source records attributed to D E Buetow.

At least 19 recordsLinked to original sources
PubMed2000

Oral immunization of mice with transgenic tomato fruit expressing respiratory syncytial virus-F protein induces a systemic immune response.

Respiratory syncytial virus (RSV) is one of the most important pathogens of infancy and early childhood. Here a fruit-based edible subunit vaccine against RSV was developed by expressing the RSV fusion (F) protein gene in transgenic tomato plants. The F-gene was expressed in ripening tomato fruit under the control of the fruit-specific E8 promoter. Oral immunization of mice with ripe transgenic tomato fruits led to the induction of both serum and mucosal RSV-F specific antibodies. The ratio of immunoglobulin subclasses produced in response to immunization suggested that a type 1 T-helper cell immune response was preferentially induced. Serum antibodies showed an increased titer when the immunized mice were exposed to inactivated RSV antigen.

Administration, Oral↗
PubMed1991

Light-Induced Chloroplast Differentiation in Soybean Cells in Suspension Culture : Ultrastructural Changes during the Bleaching and Greening Cycles.

Suspension cultures of SB-P cells of soybean (Glycine max) provide a novel, reproducible, and readily manipulable greening system useful for inducing chloroplast differentiation. The cells are subcultured and grown heterotrophically (3% sucrose) in the dark for at least three successive 14-day periods, subcultured and grown in the dark for 7 days more, and finally placed under white light and grown photoautotrophically. Chlorophyll begins to accumulate by 1 hour of light and continues up to 12 days. The chlorophyll a:chlorophyll b ratio is 3:1. Dark-grown cells contain a small amount of total carotenoids which increase 10-fold during greening. Chloroplast differentiation is strictly light dependent, with photosynthetic pigments accumulating in the light and being lost from cells returned to the dark. In the dark, the chloroplasts dedifferentiate to amyloplasts as the organized thylakoid network is lost and starch accumulates. Under continuous light, the amyloplasts differentiate into mature chloroplasts as the organelle elongates, becomes spanned by several bands of thylakoids, and undergoes grana formation. Chloroplast differentiation in SB-P cells is similar to that in intact angiosperms developing under normal light-dark cycles.

Journal Article↗
PubMed1991

Genes for insulin-like growth factors I and II are expressed in senescent rat tissues.

Insulin-like growth factor IGF-I and IGF-II gene expression was measured for the first time in rat liver, brain and heart during the later stages of life, i.e., young adulthood (6 months) to senescence (25 months). IGF-I mRNA was detected in all three tissues at all ages. Its relative level decreased the most in the liver with advancing age. IGF-II mRNA was detected in the brain and heart but not the liver at all ages. The level of IGF-II mRNA decreased only slightly in the brain from young adulthood to senescence.

Aging↗
PubMed1990

RNA synthesis by nuclei and chromatin isolated from adult and senescent Wistar rat liver.

Nuclei isolated from senescent (22-26 months) female Wistar rat liver show a decreased RNA synthesis compared to nuclei from adult (12 months) liver. Kinetic analyses demonstrate a decreased rate (25-42%) and maximal incorporation (41-54%) of labeled uridine triphosphate into both rRNA and mRNA in senescent nuclei. Chromatin-bound RNA polymerase activity is decreased by 36%, whereas free RNA polymerase activity, i.e., not bound to chromatin, is increased by 41% in senescent nuclei, but the total bound plus free activity is the same in senescent and adult nuclei. Isolated senescent chromatin shows reduced transcriptional capacity and requires a higher temperature to initiate melting. A decreased ability of chromatin to bind RNA polymerases appears to underlie the observed decreased RNA synthesis in senescent liver nuclei.

Aging↗
PubMed1990

RNA synthesis by hepatocytes isolated from adult and senescent Wistar rat liver.

The rates of synthesis of total RNA, poly(A)+ RNA and poly(A)+ RNA are decreased by 41, 49 and 42%, respectively, in hepatocytes isolated from senescent (25 months) compared to adult (12 months) female Wistar rats. Results from transcription run-on assays indicate that an impaired ability to initiate transcription largely accounts for the decreased synthesis of RNA by the senescent hepatocytes.

Aging↗
PubMed1984

Kinetics of Accumulation of Ribulose-1,5-bisphosphate Carboxylase during Greening in Euglena gracilis: Photoregulation.

The preparation of a rabbit antibody to ribulose-1,5-bisphosphate carboxylase (RuBPCase) from Euglena gracilis and its use to quantitate RuBPCase in dark- and light-grown cells and during light-induced chloroplast development (greening) are described. Light-grown Euglena have at least 36 times more RuBPCase than dark-grown Euglena. Light is required for both the initiation and continued increase in net synthesis of RuBPCase over the dark level: brief illumination 12 hours before exposure to continuous light eliminates the lags in the accumulation and increase in activity of RuBPCase (as well as in chlorophyll accumulation); net synthesis is blocked in greening cells returned to the dark or exposed to 3-(3,4-dichlorophenyl)-1,1-dimethylurea. Streptomycin or cycloheximide prevents RuBPCase accumulation when added at the beginning of greening but only partially blocks accumulation when added after 25 hours of greening. After 24 hours of greening, the activity of RuBPCase per milligram chlorophyll continues to increase slowly while concentration of the enzyme per milligram chlorophyll remains constant. This increased activity may be due to activation of the enzyme as well as to net synthesis.

Journal Article↗
PubMed1984

Kinetics of Accumulation of Ribulose-1,5-bisphosphate Carboxylase during Greening in Euglena gracilis: Nutritional Regulation.

The accumulation of ribulose-1,5-bisphosphate carboxylase (RuBPCase) in resting Euglena gracilis strain Z during greening is photo-regulated (Freyssinet, Eichholz, Buetow 1984 Plant Physiol 75: 850-857. Greening resting cells are not photosynthetically competent for about the first 24 hours in the light. Therefore, substrates for a net synthesis of the enzyme must come from endogenous constituents. During this time, degradation of endogenous paramylum (carbohydrate) reserves provides the main source of substrates. By about 24 hours of greening, resting cells are photosynthetically competent and RuBPCase accumulation becomes highly sensitive to 3-(3,4 dichlorophenyl)-1,1-dimethylurea. Therefore, from about 24 hours of greening onward, substrates (and/or energy) for RuBPCase synthesis are provided by photosynthesis. Ethanol, a nutritional substrate ordinarily used constitutively by Euglena for growth, inhibits RuBPCase accumulation when added to the resting medium in the light. The alcohol exerts this negative regulatory effect by limiting the availability of substrates needed for a net synthesis of the enzyme.

Journal Article↗
PubMed1982

The complement of cytoplasmic tRNAs, including queuosine-containing tRNAs, in adult and senescent Wistar rat liver and their levels of aminoacylation.

Our previous studies showed that both total cytoplasmic tRNAs and aminoacyl-tRNA synthetases isolated from senescent (24-30 month) female Wistar rat liver were less capable of supporting cell-free protein synthesis than were the same fractions isolated from adult (10-13 month) rat liver. The present study investigates the molecular basis for this age-related result. No significant age-related differences were found in the extent of aminoacylation of the liver cytoplasmic tRNA population, the total tRNA synthetase activity, the rate of aminoacylation of individual tRNAs, or in the overall complement of tRNA species as detected by two-dimensional gel electrophoresis. In homologous senescent aminoacylation assays, consisting of tRNAs and tRNA synthetases from senescent animals, alanine, arginine and aspartic acid were charged to a greater extent and methionine to a lesser extent compared to homologous adult assays. In heterologous assays, adult synthetases were significantly more active than senescent synthetases when charging isoleucine, methionine, phenylalanine, proline and glutamic acid, and less active when charging alanine, aspartic acid and serine. Also, senescent synthetases charged both adult and senescent tRNAs with methionine to a lesser extent than did adult synthetases. In homologous senescent assays with queuosine-containing tRNAs, asparagine, aspartic acid and histidine were charged to a greater extent and tyrosine to a lesser extent compared to homologous adult assays. Results with queuosine-tRNAs are discussed in terms of their potential ability to lower the efficiency of translation in senescent liver.

Aging↗
PubMed1982

Polypeptide composition of thylakoid membranes: two-dimensional gel analysis during development of Euglena chloroplasts.

Two-dimensional gel electrophoresis is used to analyse the polypeptide composition of thylakoid membranes during chloroplast development in Euglena gracilis. The number of polypeptide spots resolved on the two-dimensional gels is 3-4 times the number of bands previously resolved on one-dimensional gels. Groups of multiple polypeptides with the same Mr but with differing isoelectric points are detected throughout the period of chloroplast development. The pattern of polypeptide synthesis and insertion into the forming thylakoid membranes of dark-grown Euglena exposed to light conforms with previous biochemical measurements on the assembly of the electron transport chain located in these membranes.

Chloroplasts↗
PubMed1981

Decreased protein synthesis by polysomes, tRNA and aminoacyl-tRNA synthetases isolated from senescent rat liver.

A cell-free in vitro protein-synthesizing system isolated from adult(10-13-month) and senescent(24-30-month) female Wistar rat liver is described. Optimal concentrations of polysomes, aminoacyl-tRNA synthetases, tRNA, Mg 2+, K+, and labeled amino acids are reported for this in vitro system. Both are rate and extent of protein synthesis are lower with the system from senescent liver compared to that from adult liver. Senescent polysomes, tRNA synthetases, and tRNA all contribute to this decrease. Free adult or senescent polysomes support protein synthesis to the same extent. However, compared to adult polysomes, senescent membrane-bound polysomes support less protein synthesis and account for an increased proportion of the total polysome fraction. Ribonuclease and protease activities are low in the adult and the senescent in vitro systems and do not account for the decreased protein synthesis with the latter.

Aging↗
PubMed1981

In vitro reconstruction of the mitochondrial translation system of yeast.

We have isolated the translation system from yeast mitochondria and have reconstructed it in vitro. This submitochondrial system, composed of mitochondrial ribosomes, tRNA, pH 5 fraction and mRNA, is maximally active at 10 mM Mg2+ and 100 mM KCl or NH4Cl. NH4+ is more stimulatory than K+. Added Escherichia coli tRNA gives less than half the activity obtained with added mitochondrial tRNA. Activity is enhanced with protease inhibitors but not with Ca2+, spermine, or spermidine. In contrast to heterologous translation systems, the present system produces products with molecular weights similar to those of products synthesized by yeast mitochondria in vivo and by intact yeast mitochondria in vitro. The results support the idea that the unique coding features of the mitochondrial genome require a unique translation system for accurate translation of mitochondrial mRNAs.

Cell-Free System↗
PubMed1981

Gel Electrophoresis of Chloroplast Polypeptides: Comparison of One-Dimensional and Two-Dimensional Gel Analyses of Chloroplast Polypeptides From Euglena gracilis.

Euglena chloroplast polypeptides are resolved by an adaptation of the two-dimensional gel electrophoretic technique of O'Farrell (1975 J Biol Chem 250: 4007-4021). The present results are compared with those obtained by our earlier two-dimensional gel analyses as well as those obtained by one-dimensional gel analyses. Up to 75 micrograms of Euglena chloroplast polypeptides are resolved on one-dimensional sodium dodecylsulfate linear gradient 7.5 to 15% polyacrylamide gels into 43 stained polypeptide bands compared to only 33 bands resolved on a similar gel containing only 10% polyacrylamide. In contrast, two-dimensional gel electrophoresis (isoelectric focusing for the first dimension, sodium dodecylsulfate gel electrophoresis for the second dimension) further improves the resolution of the chloroplast polypeptides and especially so when a linear gradient gel is used for the second dimension. Delipidation of Euglena chloroplasts with acetone-ether and subsequent solubilization of polypeptides with Triton X-100 followed by sonication are all necessary for successful resolution of chloroplast polypeptides on two-dimensional gels. Up to 300 micrograms of chloroplast polypeptides can be clearly resolved into 56 to 59 stainable spots by the present two-dimensional gel technique when a linear gradient gel is used for the second dimension. Thus, about 30% of the polypeptide bands on a one-dimensional gel are separated into multiple polypeptides on a two-dimensional gel. The use of two-dimensional gels to separate labeled polypeptides with subsequent detection of labeled spots by autoradiography or fluorography again improves the resolution of the chloroplast polypeptides. For example, when (35)S-labeled chloroplast polypeptides are separated by the present two-dimensional gel technique with a linear gradient polyacrylamide gel in the second dimension, autoradiography or fluorography detects over 80 individual polypeptide spots. This is about twice the number resolved by our previous analyses which used a 10% polyacrylamide gel in the second dimension. Polypeptides detected range in molecular weight from about 8.5 to about 145 kilodaltons with apparent isoelectric points from pH 4.5 to 8.0. Fluorography provides rapid detection of labeled polypeptides and is 10 times more sensitive than autoradiography.

Journal Article↗
PubMed1978

Total free amino acid levels in adult and senescent rat liver.

The total free amino acid content per gram of liver wet weight does not differ significantly between adult and senescent female rats, being 28.8 +/- 3.3 mumol in adult (13-17 months) and 26.2 +/- 2.2 mumol in senescent (23-29 months) animals. This is the first study in which these two age-groups of rats have been so compared. These results combined with a survey of the literature show that amino acid levels in rat liver do not change much, if at all, throughout the lifespan of the animal.

Aging↗