Developmental and hormonal regulation of carbamoyl-phosphate synthase gene expression in rat liver: Evidence for control mechanisms at different levels in the perinatal period

Groot, C. J. De; Zonneveld, D.; Laaf, R.T.M. De; Dingemanse, Maria A.; Mooren, P.G.; Moorman, Antoon F.M.; Lamers, Wouter H.; Charles, Roger

doi:10.1016/0167-4781(86)90101-6

Cited by 55 publications

(33 citation statements)

References 28 publications

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“…The data also show that a response to fasting is seen in both periportal and pericentral hepatocytes, and are in agreement with available data for PEPCK, CPS, and OAT. 15,36,37 Only the observed increase in GS expression upon fasting appears to distinguish the mouse from the rat, in which GS activity decreases after 24 hours of starvation. 38,39 Even though the parameters describing the porto-central gradient in PEPCK are similar in fed and fasted animals, the response of PEPCK to fasting differs qualitatively from the other enzymes in that the absolute increase in mRNA concentration is similar in all hepatocytes.…”

Section: Discussionmentioning

confidence: 99%

“…After histological processing and embedding in Paraplast (Tyco Healthcare Group, Mansfield, MA), each liver was cut into serial sections (7 m). Adjacent series of 4 sections were stained by in situ hybridization with [35] S-labeled complementary RNA probes of glutamine synthetase (GS), 12 phosphoenolpyruvate carboxykinase (PEPCK), 13 ornithine aminotransferase (OAT), 14 or carbamoylphosphate synthetase (CPS), 15 exactly as described. 16 The sections were simultaneously processed under well-defined conditions to ensure that the optical density (OD) of the autoradiography signals linearly reflected the cellular messenger RNA levels (mRNA).…”

Section: Methodsmentioning

confidence: 99%

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Stereological measurement of porto-central gradients in gene expression in mouse liver

et al. 2004

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The liver is thought to consist of lobules, numerous repeating, randomly oriented units. Within these lobules, genes are expressed in gradients along the porto-central axis, which spans the distance between portal and central veins. We have developed a robust stereological method to map all points in an image to their position on this porto-central axis. This approach is based on the distribution of well-characterized periportal and pericentral enzymes, which are visualized on sections preceding and following the section of interest. Because expression of the model genes phosphoenolpyruvate carboxykinase and ornithine aminotransferase declines gradually with increasing distance from the portal vein and central vein, respectively, these genes can be used to prepare images with topographical information without any assumption about the shape of the hepatic unit, or about the direction or shape of the gradient to be determined. The "relative distance" image is a 2-dimensional image that accurately maps the relative position of hepatocytes on the porto-central axis in 3-dimensional space. It is superimposed on the serial section under investigation to relate local staining density to position on the porto-central axis and obtain the gene expression gradient. The method was used to determine the expression gradient of 2 periportal and 2 pericentral enzymes and their response to fasting. The "total distance" image was used to measure the length of the porto-central axis, which was approximately 210 m in mice and found to decrease 13% after 1 day of starvation. The method can be applied to any tissue component that can be stained quantitatively. (HEPATOLOGY 2004;39:343-352.)

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Stereological measurement of porto-central gradients in gene expression in mouse liver

et al. 2004

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“…Filters were hybridized with antisense RNA probes in vitro transcribed from rat CPS cDNA (58) and mouse cytochrome oxidase (8) or with CPS and GAPDH (glyceraldehyde-3-phosphate dehydrogenase) DNA probes (11,58).…”

Section: Methodsmentioning

confidence: 99%

Glucocorticoid Receptor, C/EBP, HNF3, and Protein Kinase A Coordinately Activate the Glucocorticoid Response Unit of the Carbamoylphosphate Synthetase I Gene

Christoffels

Grange

Kaestner

et al. 1998

Molecular and Cellular Biology

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A single far-upstream enhancer is sufficient to confer hepatocyte-specific, glucocorticoid-and cyclic AMPinducible periportal expression to the carbamoylphosphate synthetase I (CPS) gene. To identify the mechanism of hormone-dependent activation, the composition and function of the enhancer have been analyzed. DNase I protection and gel mobility shift assays revealed the presence of a cyclic AMP response element, a glucocorticoid response element (GRE), and several sites for the liver-enriched transcription factor families HNF3 and C/EBP. The in vivo relevance of the transcription factors interacting with the enhancer in the regulation of CPS expression in the liver was assessed by the analysis of knockout mice. A strong reduction of CPS mRNA levels was observed in glucocorticoid receptor-and C/EBP␣-deficient mice, whereas the CPS mRNA was normally expressed in C/EBP␤ knockout mice and in HNF3␣ and -␥ double-knockout mice. (The role of HNF␤ could not be assessed, because the corresponding knockout mice die at embryonic day 10). In hepatoma cells, most of the activity of the enhancer is contained within a 103-bp fragment, which depends for its activity on the simultaneous occupation of the GRE, HNF3, and C/EBP sites, thus meeting the requirement of a glucocorticoid response unit. In fibroblast-like CHO cells, on the other hand, the GRE in the CPS enhancer does not cooperate with the C/EBP and HNF3 elements in transactivation of the CPS promoter. In both hepatoma and CHO cells, stimulation of expression by cyclic AMP depends mainly on the integrity of the glucocorticoid pathway, demonstrating cross talk between this pathway and the cyclic AMP (protein kinase A) pathway.

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“…Pretreatment of the sections, hybridization, and washes have been described in detail (Moorman et al 1993). 35 S-labeled transcripts were made by in vitro transcription of linearized pBluescript in which the 564-bp BamHI-Smal DNA fragment localized at position 719-1283 of the CPS cDNA clone CPS-KdG (de Groot et al 1986) was inserted. CPS probe was prepared with a specific activity of 1500 cpm/pg.…”

Section: In Situ Hybridizationmentioning

confidence: 99%

“…blots were hybridized with a 32 P-labeled, random-primed CPS cDNA probe (1.9 kb ; position 2667-4545) (de Groot et al 1986). …”

Section: Northern Blot Analysismentioning

confidence: 99%

Towards Quantitative In Situ Hybridization

Jonker

Boer

Hoff

et al. 1997

J Histochem Cytochem.

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SUMMARYIn situ hybridization analysis of tissue mRNA concentrations remains to be accepted as a quantitative technique, even though exposure of tissue sections to photographic emulsion is equivalent to Northern blot analysis. Because of the biological importance of in situ quantification of RNA sequences within a morphological context, we evaluated the quantitative aspects of this technique. In calibrated microscopic samples, autoradiographic signal (density of silver grains) was proportionate to the radioactivity present, to the exposure time, and to time of development of the photographic emulsion. Similar results were obtained with tissue sections, showing that all steps of the in situ hybridization protocol, before and including the detection of the signal, can be reproducibly performed. Furthermore, the integrated density of silver grains produced in liver and intestinal sections by the in situ hybridization procedure using 35 S-labeled riboprobes is directly proportionate to the signal obtained by quantitative Northern blot analysis. The significance of this finding is that in situ quantification of RNA can be realized with high sensitivity and with the additional advantage of the possibility of localizing mRNA within the cells of interest. Application of this procedure on fetal and adult intestinal tissue showed that the carbamoylphosphate synthetase (CPS)-expressing epithelial cells of both tissues accumulated CPS mRNA to the same level but that whole-organ CPS mRNA levels decreased four-to fivefold in the same period, owing to a comparable decrease in the number of CPSexpressing cells in total intestinal tissue.

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Developmental and hormonal regulation of carbamoyl-phosphate synthase gene expression in rat liver: Evidence for control mechanisms at different levels in the perinatal period

Cited by 55 publications

References 28 publications

Stereological measurement of porto-central gradients in gene expression in mouse liver

Stereological measurement of porto-central gradients in gene expression in mouse liver

Glucocorticoid Receptor, C/EBP, HNF3, and Protein Kinase A Coordinately Activate the Glucocorticoid Response Unit of the Carbamoylphosphate Synthetase I Gene

Towards Quantitative In Situ Hybridization

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