Activity and protein localization of multiple glutamate transporters in gestation day 14 vs. day 20 rat placenta

1998 ◽  
Vol 274 (3) ◽  
pp. C603-C614 ◽  
Author(s):  
James C. Matthews ◽  
Mark J. Beveridge ◽  
Marc S. Malandro ◽  
Jeffrey D. Rothstein ◽  
Martha Campbell-Thompson ◽  
...  
Keyword(s):  
Steady State ◽  
Fetal Development ◽  
Cellular Localization ◽  
Glutamate Uptake ◽  
Protein Localization ◽  
Basal Membrane ◽  
Mrna Levels ◽  
Transport Activity ◽  
Chorioallantoic Placenta ◽  
Apical Membranes

Concentrative absorption of glutamate by the developing placenta is critical for proper fetal development. The expression of GLAST1, GLT1, EAAC1, and EAAT4, known to be capable ofd-aspartate-inhibitable and Na+-coupled glutamate transport (system [Formula: see text]), was evaluated in day 14 vs. day 20 rat chorioallantoic placenta. Steady-state mRNA levels were greater at day 20 for all transporters. Immunohistochemistry determined that the expression of GLAST1, GLT1, and EAAC1 was greater throughout the day 20 placenta and was asymmetric with respect to cellular localization. EAAT4 protein was not detected. System[Formula: see text] activity was responsible for most of the Na+-dependent glutamate uptake and was greater in day 20 than in day 14apical and basal membrane subdomains of the labyrinth syncytiotrophoblast. Greater quantities of EAAC1 and GLAST1 protein were identified on day 20, and quantities were greater in basal than in apical membranes. GLT1 expression, unchanged in apical membranes, was decreased in basal membranes. These data correlate transporter mRNA and protein content with transport activity and demonstrate an increasing capacity for glutamate absorption by the developing placenta.

Placental Transfer of Calcium and Strontium in the Rat and Rabbit

1957 ◽  
Vol 189 (1) ◽  
pp. 91-97 ◽  
Author(s):  
R. H. Wasserman ◽  
C. L. Comar ◽  
M. M. Nold ◽  
F. W. Lengemann
Keyword(s):  
Steady State ◽  
Urinary Excretion ◽  
Fetal Development ◽  
Placental Transfer ◽  
Maternal Diet ◽  
Late Pregnancy ◽  
Placental Barrier ◽  
Tracer Techniques ◽  
Comparative Metabolism ◽  
Differential Movement

The comparative metabolism of calcium and strontium during fetal development was investigated in rats and rabbits using double tracer techniques. In general, the placental transfer from dam to fetus of strontium was about one-half that of calcium; the site of discrimination was the placental barrier. The major discrimination occurred in movement of Ca* and Sr* from dam to fetus, with little or no differential movement from fetus to dam. Under steady state conditions in the rat the relative Sr*/Ca* ratios in the fetus, maternal skeleton and diet were 0.17, 0.28 and 1, respectively. The over-all discrimination of 0.17 between fetus and diet resulted from absorption (0.42), urinary excretion (0.63) and placental transfer (0.65). In the rat it was estimated that 92% of the fetal calcium had originated from the maternal diet. In the rabbit during late pregnancy, it was determined that about 24 mg of calcium/fetus/day moved across the placenta as compared with a need of about 13 mg for fetal development.

scholarly journals Molecular Mechanisms of ZC3H12C/Reg-3 Biological Activity and Its Involvement in Psoriasis Pathology

2021 ◽  
Vol 22 (14) ◽  
pp. 7311
Author(s):  
Mateusz Wawro ◽  
Jakub Kochan ◽  
Weronika Sowinska ◽  
Aleksandra Solecka ◽  
Karolina Wawro ◽  
...  
Keyword(s):  
Family Member ◽  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Inflammatory Diseases ◽  
Association Studies ◽  
Psoriasis Patient ◽  
Genome Wide Association Studies ◽  
Mrna Levels ◽  
Transcript Levels

The members of the ZC3H12/MCPIP/Regnase family of RNases have emerged as important regulators of inflammation. In contrast to Regnase-1, -2 and -4, a thorough characterization of Regnase-3 (Reg-3) has not yet been explored. Here we demonstrate that Reg-3 differs from other family members in terms of NYN/PIN domain features, cellular localization pattern and substrate specificity. Together with Reg-1, the most comprehensively characterized family member, Reg-3 shared IL-6, IER-3 and Reg-1 mRNAs, but not IL-1β mRNA, as substrates. In addition, Reg-3 was found to be the only family member which regulates transcript levels of TNF, a cytokine implicated in chronic inflammatory diseases including psoriasis. Previous meta-analysis of genome-wide association studies revealed Reg-3 to be among new psoriasis susceptibility loci. Here we demonstrate that Reg-3 transcript levels are increased in psoriasis patient skin tissue and in an experimental model of psoriasis, supporting the immunomodulatory role of Reg-3 in psoriasis, possibly through degradation of mRNA for TNF and other factors such as Reg-1. On the other hand, Reg-1 was found to destabilize Reg-3 transcripts, suggesting reciprocal regulation between Reg-3 and Reg-1 in the skin. We found that either Reg-1 or Reg-3 were expressed in human keratinocytes in vitro. However, in contrast to robustly upregulated Reg-1 mRNA levels, Reg-3 expression was not affected in the epidermis of psoriasis patients. Taken together, these data suggest that epidermal levels of Reg-3 are negatively regulated by Reg-1 in psoriasis, and that Reg-1 and Reg-3 are both involved in psoriasis pathophysiology through controlling, at least in part different transcripts.

scholarly journals Genomic structure of murine methylmalonyl-CoA mutase: evidence for genetic and epigenetic mechanisms determining enzyme activity

1993 ◽  
Vol 296 (3) ◽  
pp. 663-670 ◽  
Author(s):  
M F Wilkemeyer ◽  
E R Andrews ◽  
F D Ledley
Keyword(s):  
Enzyme Activity ◽  
Steady State ◽  
Transcription Initiation ◽  
Cultured Cells ◽  
Genomic Structure ◽  
Genetic Regulation ◽  
Regulatory Elements ◽  
Transcription Unit ◽  
Mrna Levels ◽  
Sequence Motifs

Methylmalonyl-CoA mutase (MCM) is a nuclear-encoded mitochondrial matrix enzyme. We have reported characterization of murine MCM and cloning of a murine MCM cDNA and now describe the murine Mut locus, its promoter and evidence for tissue-specific variation in MCM mRNA, enzyme and holo-enzyme levels. The Mut locus spans 30 kb and contains 13 exons constituting a unique transcription unit. A B1 repeat element was found in the 3′ untranslated region (exon 13). The transcription initiation site was identified and upstream sequences were shown to direct expression of a reporter gene in cultured cells. The promoter contains sequence motifs characteristic of: (1) TATA-less housekeeping promoters; (2) enhancer elements purportedly involved in co-ordinating expression of nuclear-encoded mitochondrial proteins; and (3) regulatory elements including CCAAT boxes, cyclic AMP-response elements and potential AP-2-binding sites. Northern blots demonstrate a greater than 10-fold variation in steady-state mRNA levels, which correlate with tissue levels of enzyme activity. However, the ratio of holoenzyme to total enzyme varies among different tissues, and there is no correlation between steady-state mRNA levels and holoenzyme activity. These results suggest that, although there may be regulation of MCM activity at the level of mRNA, the significance of genetic regulation is unclear owning to the presence of epigenetic regulation of holoenzyme formation.

Influence of environmental enrichment on steady-state mRNA levels for EAAC1, AMPA1 and NMDA2A receptor subunits in rat hippocampus

2007 ◽  
Vol 1174 ◽  
pp. 18-27 ◽  
Author(s):  
Josefine Andin ◽  
Martin Hallbeck ◽  
Abdul H. Mohammed ◽  
Jan Marcusson
Keyword(s):  
Steady State ◽  
Environmental Enrichment ◽  
Rat Hippocampus ◽  
Mrna Levels

Quantitative assessment of ground squirrel mRNA levels in multiple stages of hibernation

2002 ◽  
Vol 10 (2) ◽  
pp. 93-102 ◽  
Author(s):  
L. Elaine Epperson ◽  
Sandra L. Martin
Keyword(s):  
Steady State ◽  
Core Body Temperature ◽  
Ground Squirrels ◽  
Apolipoprotein A1 ◽  
Mrna Levels ◽  
Cdna Subtraction ◽  
Global Issues ◽  
Α2 Macroglobulin ◽  
Steady State Levels ◽  
Molecular Components

Hibernators in torpor dramatically reduce their metabolic, respiratory, and heart rates and core body temperature. These extreme physiological conditions are frequently and rapidly reversed during the winter hibernation season via endogenous mechanisms. This phenotype must derive from regulated expression of the hibernator’s genome; to identify its molecular components, a cDNA subtraction was used to enrich for seasonally upregulated mRNAs in liver of golden-mantled ground squirrels. The relative steady-state levels for seven mRNAs identified by this screen, plus five others, were measured and analyzed for seasonal and stage-specific differences using kinetic RT-PCR. Four mRNAs show seasonal upregulation in which all five winter stages differ significantly from and are higher than summer (α2-macroglobulin, apolipoprotein A1, cathepsin H, and thyroxine-binding globulin). One of these mRNAs, α2-macroglobulin, varies during the winter stages with significantly lower levels at late torpor. None of the 12 mRNAs increased during torpor. The implications for these newly recognized upregulated mRNAs for hibernation as well as more global issues of maintaining steady-state levels of mRNA during torpor are discussed.

scholarly journals Triosephosphate isomerase deficiency: consequences of an inherited mutation at mRNA, protein and metabolic levels

2005 ◽  
Vol 392 (3) ◽  
pp. 675-683 ◽  
Author(s):  
Judit Oláh ◽  
Ferenc Orosz ◽  
László G. Puskás ◽  
László Hackler ◽  
Margit Horányi ◽  
...  
Keyword(s):  
Steady State ◽  
Triosephosphate Isomerase ◽  
Specific Activity ◽  
Energy State ◽  
Dihydroxyacetone Phosphate ◽  
Peptidase Activity ◽  
Mrna Levels ◽  
Regulatory Enzymes ◽  
Computational Data ◽  
Fructose 1,6 Bisphosphate

Triosephosphate isomerase (TPI) deficiency is a unique glycolytic enzymopathy coupled with neurodegeneration. Two Hungarian compound heterozygote brothers inherited the same TPI mutations (F240L and E145Stop), but only the younger one suffers from neurodegeneration. In the present study, we determined the kinetic parameters of key glycolytic enzymes including the mutant TPI for rational modelling of erythrocyte glycolysis. We found that a low TPI activity in the mutant cells (lower than predicted from the protein level and specific activity of the purified recombinant enzyme) is coupled with an increase in the activities of glycolytic kinases. The modelling rendered it possible to establish the steady-state flux of the glycolysis and metabolite concentrations, which was not possible experimentally due to the inactivation of the mutant TPI and other enzymes during the pre-steady state. Our results showed that the flux was 2.5-fold higher and the concentration of DHAP (dihydroxyacetone phosphate) and fructose 1,6-bisphosphate increased 40- and 5-fold respectively in the erythrocytes of the patient compared with the control. Although the rapid equilibration of triosephosphates is not achieved, the energy state of the cells is not ‘sick’ due to the activation of key regulatory enzymes. In lymphocytes of the two brothers, the TPI activity was also lower (20%) than that of controls; however, the remaining activity was high enough to maintain the rapid equilibration of triosephosphates; consequently, no accumulation of DHAP occurs, as judged by our experimental and computational data. Interestingly, we found significant differences in the mRNA levels of the brothers for TPI and some other, apparently unrelated, proteins. One of them is the prolyl oligopeptidase, the activity decrease of which has been reported in well-characterized neurodegenerative diseases. We found that the peptidase activity of the affected brother was reduced by 30% compared with that of his neurologically intact brother.

Cyclic AMP analogs and retinoic acid influence the expression of retinoic acid receptor alpha, beta, and gamma mRNAs in F9 teratocarcinoma cells

1990 ◽  
Vol 10 (1) ◽  
pp. 391-396
Author(s):  
L Hu ◽  
L J Gudas
Keyword(s):  
Retinoic Acid ◽  
Steady State ◽  
Cyclic Amp ◽  
Cellular Response ◽  
Retinoic Acid Receptor ◽  
Mrna Level ◽  
Mrna Levels ◽  
Protein Synthesis Inhibitors ◽  
Receptor Alpha ◽  
F9 Teratocarcinoma

Retinoic acid (RA) receptor alpha (RAR alpha) and RAR gamma steady-state mRNA levels remained relatively constant over time after the addition of RA to F9 teratocarcinoma stem cells. In contrast, the steady-state RAR beta mRNA level started to increase within 12 h after the addition of RA and reached a 20-fold-higher level by 48 h. This RA-associated RAR beta mRNA increase was not prevented by protein synthesis inhibitors but was prevented by the addition of cyclic AMP analogs. In the presence of RA, cyclic AMP analogs also greatly reduced the RAR alpha and RAR gamma mRNA levels, even though cyclic AMP analogs alone did not alter these mRNA levels. The addition of either RA or RA plus cyclic AMP analogs did not result in changes in the three RAR mRNA half-lives. These results suggest that agents which elevate the internal cyclic AMP concentration may also affect the cellular response to RA by altering the expression of the RARs.

Codon replacement in the PGK1 gene of Saccharomyces cerevisiae: experimental approach to study the role of biased codon usage in gene expression

1987 ◽  
Vol 7 (8) ◽  
pp. 2914-2924
Author(s):  
A Hoekema ◽  
R A Kastelein ◽  
M Vasser ◽  
H A de Boer
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Steady State ◽  
Codon Usage ◽  
Experimental Approach ◽  
Mrna Translation ◽  
Yeast Cells ◽  
Expression Level ◽  
Start Codon ◽  
Mrna Levels

The coding sequences of genes in the yeast Saccharomyces cerevisiae show a preference for 25 of the 61 possible coding triplets. The degree of this biased codon usage in each gene is positively correlated to its expression level. Highly expressed genes use these 25 major codons almost exclusively. As an experimental approach to studying biased codon usage and its possible role in modulating gene expression, systematic codon replacements were carried out in the highly expressed PGK1 gene. The expression of phosphoglycerate kinase (PGK) was studied both on a high-copy-number plasmid and as a single copy gene integrated into the chromosome. Replacing an increasing number (up to 39% of all codons) of major codons with synonymous minor ones at the 5' end of the coding sequence caused a dramatic decline of the expression level. The PGK protein levels dropped 10-fold. The steady-state mRNA levels also declined, but to a lesser extent (threefold). Our data indicate that this reduction in mRNA levels was due to destabilization caused by impaired translation elongation at the minor codons. By preventing translation of the PGK mRNAs by the introduction of a stop codon 3' and adjacent to the start codon, the steady-state mRNA levels decreased dramatically. We conclude that efficient mRNA translation is required for maintaining mRNA stability in S. cerevisiae. These findings have important implications for the study of the expression of heterologous genes in yeast cells.

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