scholarly journals Expression of the phosphoenolpyruvate carboxykinase gene in 3T3-F442A adipose cells: opposite effects of dexamethasone and isoprenaline on transcription

1995 ◽  
Vol 305 (1) ◽  
pp. 65-71 ◽  
Author(s):  
S Franckhauser ◽  
J Antras-Ferry ◽  
P Robin ◽  
D Robin ◽  
D K Granner ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Gene Promoter ◽  
Adrenergic Agonist ◽  
Inductive Effects ◽  
Fold Induction ◽  
Beta Adrenergic Agonist ◽  
Cat Gene ◽  
Adipose Cells ◽  
Cat Expression

The enzyme phosphoenolpyruvate carboxykinase (PEPCK) plays a key role in gluconeogenesis in liver and in glyceroneogenesis in adipose tissue. These processes, and PEPCK, are regulated by a number of hormones, some of which have different effects on the enzyme in liver and adipose tissue. To explore this phenomenon, PEPCK gene expression was studied in 3T3-F442A adipocytes maintained in a serum-free medium. The beta-adrenergic agonist isoprenaline (isoproterenol) and a cyclic AMP analogue (8-CPT-cAMP) increased PEPCK mRNA. A maximal 3-fold induction occurred in 2 h. Dexamethasone decreased PEPCK mRNA by 80% in 4 h. Dexamethasone also counteracted the inductive effects of isoprenaline and 8-CPT-cAMP. Run-on transcription experiments showed that the isoprenaline and dexamethasone actions were, at least in part, exerted at the level of PEPCK gene transcription. These effects were further analysed by using transient and stable transfection of adipocytes with a plasmid containing bp -2100 to 69 of the PEPCK gene promoter fused to the chloramphenicol acetyltransferase (CAT) gene. In such cells isoprenaline stimulated CAT expression, an effect that was prevented if the cells were also exposed to dexamethasone.

Transient activity assays of the Trypanosoma brucei variant surface glycoprotein gene promoter: control of gene expression at the posttranscriptional level

1991 ◽  
Vol 11 (1) ◽  
pp. 338-343
Author(s):  
D Jefferies ◽  
P Tebabi ◽  
E Pays
Keyword(s):  
Trypanosoma Brucei ◽  
Gene Promoter ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Gene Promoters ◽  
Control Of Gene Expression ◽  
Transient Activity ◽  
Expression Site ◽  
Cat Gene ◽  
Cat Expression

The putative promoter of the variant surface glycoprotein (VSG) gene of Trypanosoma brucei was cloned into a plasmid containing the chloramphenicol acetyltransferase (CAT) gene. After electroporation into trypanosomes, this construct directed the expression of the CAT reporter gene. The essential region for promoter activity was found to reside within 88 bp upstream of the putative transcription start site. Transcription of the CAT construct occurred at approximately the same level in both bloodstream and procyclic forms and was resistant to alpha-amanitin. However, CAT expression appeared to be modulated in the two forms of the parasite. Sequences 3' to the gene seemed to be important in this respect, as CAT activity in bloodstream forms was readily detectable only when the 3' region of a VSG cDNA was placed downstream of the CAT gene. Two separate VSG gene promoter sequences, both cloned from T. brucei AnTat 1.3A, were equally able to direct CAT expression, which suggests that there are a number of potential VSG gene promoters in the genome, although usually only one expression site is fully active at any one time.

scholarly journals Interaction between two different regulatory elements activates the murine alpha A-crystallin gene promoter in explanted lens epithelia.

1987 ◽  
Vol 7 (5) ◽  
pp. 1807-1814 ◽  
Author(s):  
A B Chepelinsky ◽  
B Sommer ◽  
J Piatigorsky
Keyword(s):  
Dna Sequences ◽  
Gene Promoter ◽  
Regulatory Elements ◽  
Regulatory Sequences ◽  
Base Pairs ◽  
Hybrid Gene ◽  
Promoter Sequences ◽  
Hybrid Genes ◽  
Cat Gene ◽  
Cat Expression

Previous experiments have indicated that 5' flanking DNA sequences (nucleotides-366 to +46) are capable of regulating the lens-specific transcription of the murine alpha A-crystallin gene. Here we have analyzed these 5' regulatory sequences by transfecting explanted embryonic chicken lens epithelia with different alpha A-crystallin-CAT (chloramphenicol acetyltransferase) hybrid genes (alpha A-crystallin promoter sequences fused to the bacterial CAT gene in the pSVO-CAT expression vector). The results indicated the presence of a proximal (-88 to +46) and a distal (-111 to -88) domain which must interact for promoter function. Deletion experiments showed that the sequence between -88 and -60 was essential for function of the proximal domain in the explanted epithelia. A synthetic oligonucleotide containing the sequence between -111 and -84 activated the proximal domain when placed in either orientation 57 base pairs upstream from position -88 of the alpha A-crystallin-CAT hybrid gene.

scholarly journals Activities of enzymes of glycerolipid synthesis in brown adipose tissue after treatment of rats with the adrenergic agonists BRL 26830A and phenylephrine, after exposure to cold and in streptozotocin-diabetes

1991 ◽  
Vol 277 (3) ◽  
pp. 665-669 ◽  
Author(s):  
J R D Mitchell ◽  
E D Saggerson
Keyword(s):  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Streptozotocin Diabetes ◽  
Fatty Acyl ◽  
Adrenergic Agonist ◽  
Interscapular Brown Adipose Tissue ◽  
Exposure To Cold ◽  
Brown Adipose ◽  
Glycerolipid Synthesis ◽  
Beta Adrenergic Agonist

1. Measurements were made, relative to tissue DNA, of the activities of enzymes of glycerolipid synthesis in homogenates of interscapular brown adipose tissue. These were: mitochondrial and microsomal forms of glycerolphosphate acyltransferase (GPAT), Mg(2+)-dependent phosphatidate phosphohydrolase (PPH) and fatty acyl-CoA synthetase (FAS). 2. In normal animals, 3 days of cold-exposure (4 degrees C) increased all activities. The increase in mitochondrial GPAT activity was particularly pronounced (5-fold). Administration of the beta-adrenergic agonist BRL 26830A mimicked the effect of cold on microsomal GPAT activity. Mitochondrial GPAT, PPH and FAS activities were unresponsive to BRL 26830A. The alpha-adrenergic agonist phenylephrine significantly decreased activities of GPAT and PPH. 3. Streptozotocin-diabetes decreased mitochondrial GPAT activity, but did not abolish the effect of cold to increase this activity or the activity of microsomal GPAT. Diabetes abolished the effect of cold on PPH and FAS activities. 4. The findings are relevant to signals that drive early events in mitochondriogenesis and cell proliferation in brown adipose tissue on exposure to cold.

Interaction between two different regulatory elements activates the murine alpha A-crystallin gene promoter in explanted lens epithelia

1987 ◽  
Vol 7 (5) ◽  
pp. 1807-1814
Author(s):  
A B Chepelinsky ◽  
B Sommer ◽  
J Piatigorsky
Keyword(s):  
Dna Sequences ◽  
Gene Promoter ◽  
Regulatory Elements ◽  
Regulatory Sequences ◽  
Base Pairs ◽  
Hybrid Gene ◽  
Promoter Sequences ◽  
Hybrid Genes ◽  
Cat Gene ◽  
Cat Expression

Previous experiments have indicated that 5' flanking DNA sequences (nucleotides-366 to +46) are capable of regulating the lens-specific transcription of the murine alpha A-crystallin gene. Here we have analyzed these 5' regulatory sequences by transfecting explanted embryonic chicken lens epithelia with different alpha A-crystallin-CAT (chloramphenicol acetyltransferase) hybrid genes (alpha A-crystallin promoter sequences fused to the bacterial CAT gene in the pSVO-CAT expression vector). The results indicated the presence of a proximal (-88 to +46) and a distal (-111 to -88) domain which must interact for promoter function. Deletion experiments showed that the sequence between -88 and -60 was essential for function of the proximal domain in the explanted epithelia. A synthetic oligonucleotide containing the sequence between -111 and -84 activated the proximal domain when placed in either orientation 57 base pairs upstream from position -88 of the alpha A-crystallin-CAT hybrid gene.

scholarly journals Expression of the phosphoenolpyruvate carboxykinase gene in 3T3-F442A adipose cells: effects of retinoic acid and differentiation

1994 ◽  
Vol 302 (3) ◽  
pp. 943-948 ◽  
Author(s):  
J Antras-Ferry ◽  
S Franckhauser ◽  
D Robin ◽  
P Robin ◽  
D K Granner ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Dna Sequences ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Gene Encoding ◽  
Specific Expression ◽  
Stable Transfectants ◽  
Half Maximal Effective Concentration ◽  
Fold Induction ◽  
Cat Gene

3T3-F442A adipocytes express the gene encoding cytosolic phosphoenolpyruvate carboxykinase (GTP) (PEPCK). Retinoic acid (RA) caused a 5-fold induction of PEPCK mRNA within 6 h in these cells with a half-maximal effective concentration of approximately 75 microM. This effect was independent of cycloheximide and inhibited by actinomycin D. In vitro run-on experiments using isolated nuclei confirmed that the RA-induced increase was mainly due to an increased rate of transcription of the gene. Stable transfectants bearing either the region of the PEPCK promoter from -2100 to +69 fused to the chloramphenicol acetyltransferase (CAT) gene (pPL1-CAT) or -600 to +69 fused to CAT (pPL9-CAT) were used to study PEPCK gene regulation during differentiation. The same transfected cells were used to analyse the RA effect. Preadipocytes containing pPL1-CAT expressed a much lower level of CAT activity than did adipocytes. pPL9-CAT was not expressed in either preadipocytes or adipocytes. RA induced the expression of CAT activity in preadipocytes and adipocytes transfected with pPL1-CAT, but had no effect in cells transfected with pPL9-CAT. These results suggest that one or more DNA sequences located between -2100 and -600 bp of the PEPCK promoter is required for adipocyte-specific expression of this gene. RA action is independent of the state of differentiation and appears to require different elements in fat cells from those required in liver.

scholarly journals Effect of suckling and adrenergic stimulation on peripheral deiodination in lactating rats: differential expression of type 1 deiodinase mRNA forms

2001 ◽  
Vol 171 (3) ◽  
pp. 533-540 ◽  
Author(s):  
C Aceves ◽  
R Rojas-Huidobro
Keyword(s):  
Adipose Tissue ◽  
Mammary Gland ◽  
Brown Adipose Tissue ◽  
Adrenergic Stimulation ◽  
Adrenergic Agonist ◽  
Large Form ◽  
Brown Adipose ◽  
Sympathetic Nervous ◽  
Beta Adrenergic Agonist

Previous works led us to propose that peripheral iodothyronine deiodination is mainly regulated by the reciprocal interaction between the thyroid and the sympathetic nervous system (SNS). In this study, we analyzed the role suckling exerts, through SNS activation, upon deiodination of thyronines in liver, heart, brown adipose tissue and mammary gland during lactation. Our results showed that resuckling causes a concurrent stimulatory response on deiodinase type 1 (D1) in heart and mammary gland, but not in liver and brown adipose tissue. The stimulatory response was mimicked by norepinephrine and by the beta-adrenergic agonist isoproterenol, through the overexpression of the large form of D1 mRNA. These results suggested that, during lactation, peripheral thyronine deiodination is co-ordinated by the SNS, and suckling is a major modulatory influence.

Specificity of a retinoic acid response element in the phosphoenolpyruvate carboxykinase gene promoter: consequences of both retinoic acid and thyroid hormone receptor binding

1991 ◽  
Vol 11 (10) ◽  
pp. 5164-5170
Author(s):  
P C Lucas ◽  
B M Forman ◽  
H H Samuels ◽  
D K Granner
Keyword(s):  
Gene Expression ◽  
Retinoic Acid ◽  
Thyroid Hormone ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Thyroid Hormone Receptor ◽  
Gene Promoter ◽  
Response Element ◽  
Promoter Sequences ◽  
Retinoic Acid Response Element ◽  
Cat Expression

The ability of a retinoic acid (RA) response element (RARE) in the phosphoenolpyruvate carboxykinase (PEPCK) gene promoter to mediate effects of either RA or thyroid hormone (T3) on gene expression was studied. Fusion gene constructs consisting of PEPCK promoter sequences ligated to the chloramphenicol acetyltransferase (CAT) reporter gene were used for this analysis. While T3 induced CAT expression to a small degree (about twofold) when such constructs were transiently transfected into H4IIE rat hepatoma cells, along with an expression vector encoding the alpha subtype of the T3 receptor (TR), this effect was mediated by promoter sequences distinct from the PEPCK RARE. Although TRs were capable of binding the PEPCK RARE in the form of putative monomers, dimers, and heterodimers with RA receptors (RARs), this element failed to mediate any positive effect of T3 on gene expression. In contrast, the PEPCK RARE mediated six- to eightfold induction of CAT expression by RA. When TRs were coexpressed along with RARs in transfected H4IIE cells, this RA induction was substantially blunted in a T3-independent manner. This inhibitory effect may be due to the binding of nonfunctional TRs or TR-RAR heterodimers to the PEPCK RARE. A model is proposed to explain the previously observed in vivo effects of T3 on PEPCK gene expression.

scholarly journals Transient activity assays of the Trypanosoma brucei variant surface glycoprotein gene promoter: control of gene expression at the posttranscriptional level.

1991 ◽  
Vol 11 (1) ◽  
pp. 338-343 ◽  
Author(s):  
D Jefferies ◽  
P Tebabi ◽  
E Pays
Keyword(s):  
Trypanosoma Brucei ◽  
Gene Promoter ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Gene Promoters ◽  
Control Of Gene Expression ◽  
Transient Activity ◽  
Expression Site ◽  
Cat Gene ◽  
Cat Expression

The putative promoter of the variant surface glycoprotein (VSG) gene of Trypanosoma brucei was cloned into a plasmid containing the chloramphenicol acetyltransferase (CAT) gene. After electroporation into trypanosomes, this construct directed the expression of the CAT reporter gene. The essential region for promoter activity was found to reside within 88 bp upstream of the putative transcription start site. Transcription of the CAT construct occurred at approximately the same level in both bloodstream and procyclic forms and was resistant to alpha-amanitin. However, CAT expression appeared to be modulated in the two forms of the parasite. Sequences 3' to the gene seemed to be important in this respect, as CAT activity in bloodstream forms was readily detectable only when the 3' region of a VSG cDNA was placed downstream of the CAT gene. Two separate VSG gene promoter sequences, both cloned from T. brucei AnTat 1.3A, were equally able to direct CAT expression, which suggests that there are a number of potential VSG gene promoters in the genome, although usually only one expression site is fully active at any one time.

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