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Y Hod

Publications and source records attributed to Y Hod.

30 records · Page 2Linked to original sources
PubMed1989

Nutritional and hormonal regulation of mRNA abundance for arginine biosynthetic enzymes in kidney.

Argininosuccinate synthetase and argininosuccinate lyase catalyze the synthesis of arginine from citrulline in kidney and also serve as components of the urea cycle in liver of ureotelic animals. Dietary and hormonal regulation of mRNAs encoding these enzymes have been well studied in liver but not in kidney. Messenger RNAs for these enzymes are localized within the renal cortex. Starvation and extreme variations in dietary protein content (0% vs 60% casein) produced 2.6- to 3.5-fold increases in mRNA abundance for these two enzymes in rat kidney. Argininosuccinate lyase mRNA was not induced by dibutyryl cAMP, dexamethasone, or a combination of the two agents. In contrast, argininosuccinate synthetase mRNA was induced 2-fold by dibutyryl cAMP but was unresponsive to dexamethasone. Thus, diet and hormones regulate levels of these mRNAs in rat kidney, but the responses are both qualitatively and quantitatively distinct from the responses previously reported for rat liver.

Animals↗
PubMed1988

Cyclic AMP stabilizes the mRNA for phosphoenolpyruvate carboxykinase (GTP) against degradation.

It is now well established that cAMP induces the transcription rate of the gene for phosphoenolpyruvate carboxykinase (GTP) (EC 4.1.1.32) and that this induction is dependent on a nucleotide domain located within the promoter-regulatory region of the gene (Short, J. M., Wynshaw-Boris, A., Short, H. P., and Hanson, R. W. (1986) J. Biol. Chem. 261, 9721-9726). We report here that cAMP also stabilizes phosphoenolpyruvate carboxykinase mRNA against degradation. Using two independent experimental approaches, we show that the half-life of the mRNA for phosphoenolpyruvate carboxykinase is extended when FTO-2B rat hepatoma cells are exposed to dibutyryl cyclic AMP (Bt2cAMP). In the first experiment, the rate of decay of phosphoenolpyruvate carboxykinase mRNA was determined in cells incubated in the presence of insulin, which has been shown to block the transcription rate of the gene for the enzyme. Under these conditions, the half-life of phosphoenolpyruvate carboxykinase mRNA was 30 min. However, in cells incubated in the presence of Bt2cAMP, the mRNA decayed with a half-life of 150 min. In the other experiment, mRNA stability was measured under steady state conditions, utilizing a "pulse-chase" approach. The apparent half-life of phosphoenolpyruvate carboxykinase mRNA increased from 40 min to over 250 min in Bt2cAMP-treated cells. No significant change in the stability of total cellular RNA was noted. Other experiments have shown that the transcription rate of the gene for phosphoenolpyruvate carboxykinase peaks within the first 20 min after exposing the cells to Bt2cAMP and then levels off, while the abundance of the mRNA reaches a maximum at about 90 min and remains at this level thereafter. Thus, the long term effect of cAMP on the expression of the gene coding for phosphoenolpyruvate carboxykinase occurs at least in part, through an alteration in the degradation rate of the mRNA for this enzyme.

Algorithms↗
PubMed1986

Comparison of fiber optical and video monitor stimulators in normals and multiple sclerosis patients.

A comparison of VEP findings using a fiber optical stimulator with three color combinations (black/red, black/green and red/green) and a conventional video monitor stimulator in black/white was performed in 3 groups of subjects: a group with definite lesions in the visual pathway, a group with suspected lesions and a control group. No significant correlations of P100 latency were found in the normative group, probably because of the small range of their values. All correlations were significant in the two patient groups, except for the red/green stimuli in the definite group. In general, the red/green combination was inferior to other color combinations in eliciting VEPs. The patient groups with definite, as well as suspected, lesions indicated no benefit from the color stimuli, as compared with black/white. The major advantage of the fiber optical stimulator is its simplicity, the lack of stimulus artifacts and the accessibility to the patient's bed side, thanks to its small size. Stimulators with higher illuminance and improved pattern element shape should narrow the still unacceptable normative variability of the wave forms recorded.

Adolescent↗
PubMed1986

Nucleotide sequence of the mRNA encoding the cytosolic form of phosphoenolpyruvate carboxykinase (GTP) from the chicken.

We have determined the sequence of the mRNA encoding cytosolic phosphoenolpyruvate carboxykinase (GTP) [GTP:oxaloacetate carboxy-lyase (transphosphorylating), EC 4.1.1.32] from the chicken and have deduced the primary structure of the protein. The message for the enzyme is 2762 bases long and encodes a protein of 622 amino acids with a molecular mass of 69,522 daltons. The 5' untranslated region is 246 nucleotides long and contains two nonfunctional AUG initiator codons. The 3' untranslated sequence is 649 bases long and contains multiple polyadenylylation signals. There are regions of dyad symmetry and an A + U-rich region within the 3' translated and untranslated sequences of the message. Such regions are also present in the mRNA for the enzyme from the rat and may be of functional significance. Conserved regions of the enzyme, that may interact with substrates, were identified by comparing the amino acid sequence of phosphoenolpyruvate carboxykinase with that of other proteins that use guanine nucleotides and phosphoenolpyruvate as substrates.

Amino Acid Sequence↗
PubMed1984

Induction by cAMP of the mRNA encoding the cytosolic form of phosphoenolpyruvate carboxykinase (GTP) from the chicken. Identification and characterization of a cDNA clone for the enzyme.

Previous work from our laboratory (Hod, Y., Utter, M. F., and Hanson, R. W. (1982) J. Biol. Chem. 257, 13787-13794) has demonstrated that chicken kidney contains both mitochondrial and cytosolic forms of phosphoenolpyruvate carboxykinase (GTP) (EC 4.1.1.32) and that the two forms are distinct proteins. Using poly(A+) RNA from chicken kidney, a double-stranded cDNA library was constructed. DNA clones containing sequences complementary to the mRNA for the cytosolic form of phosphoenolpyruvate carboxykinase were initially identified by colony hybridization with 32P-labeled cDNA transcribed from an RNA fraction enriched for the enzyme mRNA. The identity of plasmids containing phosphoenolpyruvate carboxykinase cDNA was confirmed by hybrid-selected translation. Mature mRNA for cytosolic phosphoenolpyruvate carboxykinase of the chicken is 2.8 kilobases in length, similar to that previously noted for mRNA coding for the same enzyme in the rat. The cDNA for the chicken enzyme hybridizes with several restriction fragments of the corresponding cDNA for the rat cytosolic phosphoenolpyruvate carboxykinase, indicating conservation of nucleotide sequences during evolution. Wide spread conservation of sequence homology is also demonstrated by the hybridization of the cDNA for the rat phosphoenolpyruvate carboxykinase with a 2.8-kilobase RNA from the livers of a variety of vertebrates including amphibian, avian, and primate species. Specific mRNA coding for the cytosolic form of phosphoenolpyruvate carboxykinase was present in chicken kidney but absent from the liver, even in animals starved for 48 h. However, the administration of cAMP to normal fed chickens caused a rapid induction of phosphoenolpyruvate carboxykinase mRNA. These findings suggest that the gene for the cytosolic enzyme in chicken liver can be expressed if the proper hormonal stimuli are present.

Animals↗
PubMed1984

The gene encoding the cytosolic form of phosphoenolpyruvate carboxykinase (GTP) from the chicken.

The gene for cytosolic phosphoenolpyruvate carboxykinase (GTP) (EC 4.1.1.32) from the chicken was isolated from a recombinant library containing the chicken genome in phage lambda Charon 4A. The isolated clone, lambda PCK1cc, contains the complete gene for the enzyme as well as both 5' and 3' flanking sequences. The gene is approximately 8 kilobases in length divided into 8 exons, as demonstrated by restriction endonuclease mapping and DNA-RNA heteroduplex analysis. Southern blotting of chicken chromosomal DNA digested with various restriction enzymes shows a pattern predicted from the restriction map of lambda PCK1cc. The phosphoenolpyruvate carboxykinase gene is present as a single copy in the haploid chicken genome. The 5' region of the gene was defined by S1 nuclease mapping and by sequencing. Two mRNA species with discrete 5' ends were observed using S1 nuclease mapping. The ratio between the amounts of these multiple forms of mRNA is the same in chicken kidney and liver and is not affected by induction of the enzyme mRNA by cAMP. Examination of sequence homologies with the gene for rat cytosolic phosphoenolpyruvate carboxykinase indicates a putative control region contained in flanking sequences at the 5' end of the gene.

Animals↗
PubMed1982

The mitochondrial and cytosolic forms of avian phosphoenolpyruvate carboxykinase (GTP) are encoded by different messenger RNAs.

Previous work from our laboratory (Watford, M., Hod, Y., Chiao, Y. B., Utter M. F., and Hanson R. W. (1981) J. Biol. Chem. 256, 10023-10027) indicated that in the chicken, hepatic phosphoenolpyruvate carboxykinase is in the mitochondria, whereas kidney contains both a mitochondrial and cytosolic form of the enzyme. In the present study the two forms of phosphoenolpyruvate carboxykinase were purified and shown to be distinct proteins which differ in size, charge, and immunochemical properties. Using a cell-free protein synthesis system, we demonstrate that the cytosolic isozyme is encoded only by kidney mRNA and that its translation form has similar properties to that of the mature protein. On the other hand, the mitochondrial enzyme is encoded by liver and kidney mRNA and synthesized as a protein about 2000 daltons larger than the mature form. The putative precursor of the mitochondrial phosphoenolpyruvate carboxykinase is processed to a mature size by isolated, respiring mitochondria. We further show that the mRNA species for the cytosolic and mitochondrial forms are separable and are about 3 and 4 kilobases, respectively. It is concluded that the two forms of phosphoenolpyruvate carboxykinase of the chicken are encoded by distinct mRNA species.

Animals↗
PubMed1976

Relationship of the pool of intracellular valine to protein synthesis and degradation in cultured cells.

To explore the role of the pool of intracellular free valine in the processes of protein synthesis and protein degradation, cultured hepatoma (HTC) cells were incubated in media containing varying concentrations of L-valine, under conditions of constant rates of protein synthesis and protein breakdown, and at steady state levels of intracellular valine specific radioactivities. Two types of experiments were compared: in the first (designated "incorporation experiment"), unlabeled cells were exposed to [3H]valine for a short period of time. In the second (termed "reincorporation experiment"), cells were prelabled with [3H]valine and then incubated for a brief period with media containing different concentrations of unlabeled valine; reincorporation of [3H]valine was calculated by the difference between the release of [3H]valine from labeled cellular proteins at low valine concentrations, and the maximal rate of the release at high valine concentrations. In both types of experiments, the rates of [3H]valine incorporation or reincorporation were compared with the respective specific radioactivities of free intracellular valine. In the incorporation experiment, the rates of [3H]valine incorporation into protein calculated by the intracellular specific radioactivities were not constant, but showed an upward deviation at low valine concentrations. This is in agreement with the results of Mortimore, G.E., Woodside, K.H., and Henry, J.E. ((1972) J. Biol. Chem. 247, 2776-2784) in the perfused rat liver. By contrast, in the reincorporation experiment, the calculated rates of [3H]valine reincorporation based on intracellular specific radioactivities were constant throughout the range of valine concentrations. The constant value of calculated valine reincorporation was lower by 30 to 50% than the calculated rate of valine incorporation at high valine concentrations. The following model is proposed to explain these results. There is one common pool of free intracellular valine, but there are two sites where valyl-tRNA can be formed. The first is an internal site that utilizes valine from the intracellular pool, and the second is an external (possibly membranous) system that converts extracellular valine directly to valyl-tRNA. Valine originating from protein degradation flows into the intracellular pool, from which it can be reutilized by the internal system. According to these assumptions, in the incorporation experiment and at low valine concentrations, the specific activity of valyl-tRNA is higher than that of the intracellular pool of free valine, due to the contribution of the external system. On the other hand, in the reincorporation experiment the specific activity of extracellular valine is negligible in comparison with that of the intracellular pool. Therefore, in this case the specific activity of valyl-tRNA is proportional to that of the intracellular pool, with a constant dilution by unlabeled valine of extracellular origin...

Cell Line↗