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

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.

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Interferon-responsive regulatory elements in the promoter of the human 2',5'-oligo(A) synthetase gene.

Molecular and Cellular Biology ◽

10.1128/mcb.7.12.4498 ◽

1987 ◽

Vol 7 (12) ◽

pp. 4498-4504 ◽

Cited By ~ 95

Author(s):

P Benech ◽

M Vigneron ◽

D Peretz ◽

M Revel ◽

J Chebath

Keyword(s):

Dna Sequences ◽

Regulatory Region ◽

Gene Promoter ◽

Regulatory Elements ◽

Base Pairs ◽

Independent Manner ◽

E Gene ◽

Constitutive Enhancer ◽

Chloramphenicol Acetyltransferase Gene ◽

Simplex Virus

The interferon (IFN)-activated human 2',5'-oligo(A) synthetase E gene contains 11 RNA starts and lacks TATA and CAAT signals. DNA sequences around the promoter make the expression of the chloramphenicol acetyltransferase gene (CAT) inducible over 20-fold by IFN. A 72-base-pair segment (E-IRS) immediately upstream of the RNA starts was defined as being required for IFN-activated expression of the E-gene promoter-CAT constructs and acts in a position-independent manner. It also confers IFN-activated enhancement to the herpes simplex virus thymidine kinase promoter. On this promoter, the 5' part of the E-IRS functions as a constitutive enhancer, while the last 16 base pairs of the E-IRS is sufficient to give IFN-induced expression. On the E-gene promoter, the constitutive enhancer and the IFN-activated sequence are both needed but can be separated. In addition, promoter competition experiments indicate a third regulatory region which helps to repress expression of the E gene in uninduced cells.

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Interferon-responsive regulatory elements in the promoter of the human 2',5'-oligo(A) synthetase gene

Molecular and Cellular Biology ◽

10.1128/mcb.7.12.4498-4504.1987 ◽

1987 ◽

Vol 7 (12) ◽

pp. 4498-4504

Author(s):

P Benech ◽

M Vigneron ◽

D Peretz ◽

M Revel ◽

J Chebath

Keyword(s):

Dna Sequences ◽

Regulatory Region ◽

Gene Promoter ◽

Regulatory Elements ◽

Base Pairs ◽

Independent Manner ◽

E Gene ◽

Constitutive Enhancer ◽

Chloramphenicol Acetyltransferase Gene ◽

Simplex Virus

The interferon (IFN)-activated human 2',5'-oligo(A) synthetase E gene contains 11 RNA starts and lacks TATA and CAAT signals. DNA sequences around the promoter make the expression of the chloramphenicol acetyltransferase gene (CAT) inducible over 20-fold by IFN. A 72-base-pair segment (E-IRS) immediately upstream of the RNA starts was defined as being required for IFN-activated expression of the E-gene promoter-CAT constructs and acts in a position-independent manner. It also confers IFN-activated enhancement to the herpes simplex virus thymidine kinase promoter. On this promoter, the 5' part of the E-IRS functions as a constitutive enhancer, while the last 16 base pairs of the E-IRS is sufficient to give IFN-induced expression. On the E-gene promoter, the constitutive enhancer and the IFN-activated sequence are both needed but can be separated. In addition, promoter competition experiments indicate a third regulatory region which helps to repress expression of the E gene in uninduced cells.

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Arabidopsis thaliana Myb59 Gene Is Involved in the Response to Heterodera schachtii Infestation, and Its Overexpression Disturbs Regular Development of Nematode-Induced Syncytia

International Journal of Molecular Sciences ◽

10.3390/ijms22126450 ◽

2021 ◽

Vol 22 (12) ◽

pp. 6450

Author(s):

Anita Wiśniewska ◽

Kamila Wojszko ◽

Elżbieta Różańska ◽

Klaudia Lenarczyk ◽

Karol Kuczerski ◽

...

Keyword(s):

Cell Metabolism ◽

Gene Promoter ◽

Promoter Sequence ◽

Regulatory Elements ◽

Heterodera Schachtii ◽

Myb Transcription Factor ◽

Parasitic Nematodes ◽

Regulatory Sequences ◽

Gene Encoding ◽

Constitutive Overexpression

Transcription factors are proteins that directly bind to regulatory sequences of genes to modulate and adjust plants’ responses to different stimuli including biotic and abiotic stresses. Sedentary plant parasitic nematodes, such as beet cyst nematode, Heterodera schachtii, have developed molecular tools to reprogram plant cell metabolism via the sophisticated manipulation of genes expression, to allow root invasion and the induction of a sequence of structural and physiological changes in plant tissues, leading to the formation of permanent feeding sites composed of modified plant cells (commonly called a syncytium). Here, we report on the AtMYB59 gene encoding putative MYB transcription factor that is downregulated in syncytia, as confirmed by RT-PCR and a promoter pMyb59::GUS activity assays. The constitutive overexpression of AtMYB59 led to the reduction in A. thaliana susceptibility, as indicated by decreased numbers of developed females, and to the disturbed development of nematode-induced syncytia. In contrast, mutant lines with a silenced expression of AtMYB59 were more susceptible to this parasite. The involvement of ABA in the modulation of AtMYB59 gene transcription appears feasible by several ABA-responsive cis regulatory elements, which were identified in silico in the gene promoter sequence, and experimental assays showed the induction of AtMYB59 transcription after ABA treatment. Based on these results, we suggest that AtMYB59 plays an important role in the successful parasitism of H. schachtii on A. thaliana roots.

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Ecdysterone regulatory elements function as both transcriptional activators and repressors

Molecular and Cellular Biology ◽

10.1128/mcb.11.4.1846-1853.1991 ◽

1991 ◽

Vol 11 (4) ◽

pp. 1846-1853

Author(s):

L Dobens ◽

K Rudolph ◽

E M Berger

Keyword(s):

Regulatory Element ◽

Regulatory Elements ◽

Upstream Region ◽

Transcriptional Activators ◽

Reporter Plasmid ◽

Bacterial Gene ◽

Simplex Virus ◽

Very High ◽

Cat Gene ◽

Cat Expression

A synthetic, 23-bp ecdysterone regulatory element (EcRE), derived from the upstream region of the Drosophila melanogaster hsp27 gene, was inserted adjacent to the herpes simplex virus thymidine kinase promoter fused to a bacterial gene for chloramphenicol acetyltransferase (CAT). Hybrid constructs were transfected into Drosophila S3 cells and assayed for ecdysterone-inducible CAT expression. In the absence of ecdysterone a tandem pair of EcREs repressed the high constitutive level of CAT activity found after transfection with the parent reporter plasmid alone. After hormone addition very high levels of CAT activity were observed. Insertion of the EcRE pair 3' of the CAT gene also led to high levels of ecdysterone-induced CAT expression, but the repression of high constitutive levels of CAT activity failed to occur. The EcRE-CAT construct was cotransfected with plasmids containing tandem 10-mers or 40-mers of the EcRE but lacking a reporter gene. These additional EcREs led to a reduced level of ecdysterone-induced CAT activity and to an elevation of basal CAT activity in the absence of hormone. The data suggest that the receptor binds to the EcRE in the absence of hormone, blocking basal transcription from a constitutive promoter. In the presence of ecdysterone, receptor-hormone binding to the EcRE leads to greatly enhanced transcription.

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

Molecular and Cellular Biology ◽

10.1128/mcb.11.1.338-343.1991 ◽

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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Expression of the K-fgf proto-oncogene is controlled by 3' regulatory elements which are specific for embryonal carcinoma cells

Molecular and Cellular Biology ◽

10.1128/mcb.10.6.2475-2484.1990 ◽

1990 ◽

Vol 10 (6) ◽

pp. 2475-2484

Author(s):

A M Curatola ◽

C Basilico

Keyword(s):

Dna Sequences ◽

Embryonal Carcinoma ◽

Protein S ◽

Cell Types ◽

Regulatory Elements ◽

Developmentally Regulated ◽

Cis Acting ◽

Dna Elements ◽

Ec Cells ◽

Cat Expression

Expression of the K-fgf/hst proto-oncogene appears to be restricted to cells in the early stages of development, such as embryonal carcinoma (EC) cells. When EC cells are induced to differentiate, K-fgf expression is drastically repressed. To identify cis-acting DNA elements responsible for this type of regulation, we constructed a plasmid in which cat gene expression was driven by about 1 kilobase of upstream K-fgf human DNA sequences, including the putative promoter, and transfected it into undifferentiated F9 EC cells or HeLa cells as prototypes of cells which express or do not express, respectively, the K-fgf proto-oncogene. This plasmid was essentially inactive in both cell types, and the addition of more than 8 kilobases of DNA sequences upstream of the K-fgf promoter did not lead to any increase in chloramphenicol acetyltransferase (CAT) expression. On the other hand, when we inserted in this plasmid DNA sequences which are 3' of the human K-fgf coding sequences, we could detect a significant stimulation of CAT activity. Analysis of these sequences led to the identification of enhancerlike DNA elements which are part of the 3' noncoding region of K-fgf exon 3 and promote CAT expression only in undifferentiated mouse F9 or human NT2/D1 EC cells, but not in HeLa, 3T3, or differentiated F9 cells, therefore mimicking the physiological expression of the K-fgf proto-oncogene. Similar elements are also present in the 3' region of the murine K-fgf proto-oncogene, in a region showing high homology to the human K-fgf sequences. These regulatory elements can promote CAT expression from heterologous promoters in an EC-specific manner, suggesting that they interact with a specific cellular transacting protein(s) whose expression is developmentally regulated.

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Interaction of a tissue-specific factor with an essential rat growth hormone gene promoter element.

Molecular and Cellular Biology ◽

10.1128/mcb.7.3.1193 ◽

1987 ◽

Vol 7 (3) ◽

pp. 1193-1197 ◽

Cited By ~ 51

Author(s):

B L West ◽

D F Catanzaro ◽

S H Mellon ◽

P A Cattini ◽

J D Baxter ◽

...

Keyword(s):

Growth Hormone ◽

Dna Sequences ◽

Site Directed Mutagenesis ◽

Specific Factor ◽

Growth Hormone Gene ◽

Base Pairs ◽

Hybrid Gene ◽

Specific Expression ◽

Tissue Specific ◽

Rat Growth

Rat growth hormone (rGH) gene expression is normally restricted to the anterior pituitary. As a model of this tissue specificity, we compared the transient expression of an rGH-chloramphenicol acetyltransferase (CAT) hybrid gene in rGH-producing rat pituitary tumor (GC) cells and in non-rGH-producing rat fibroblast (rat-2) cells. Deletion analysis of the rGH portion of this hybrid gene demonstrated that DNA sequences within 140 base pairs 5' to the rGH gene were sufficient for correct cell type-specific expression. Deletion of an additional 35 base pairs of the rGH 5'-flanking DNA resulted in a loss of expression of the transfected hybrid gene and correlated with the interaction of a putative trans-acting factor with this region of the rGH promoter. This factor was detectable by DNase I footprinting in a crude nuclear extract from GC cells but not from rat-2 cells. Site-directed mutagenesis of the footprint region caused complete loss of expression of a hybrid gene containing 530 base pairs 5' to the rGH gene. Thus, the interaction of this factor, which we term GC2, is likely to be essential for the tissue-specific expression of the rGH gene.

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

Biochemical Journal ◽

10.1042/bj3050065 ◽

1995 ◽

Vol 305 (1) ◽

pp. 65-71 ◽

Cited By ~ 28

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.

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Ecdysterone regulatory elements function as both transcriptional activators and repressors.

Molecular and Cellular Biology ◽

10.1128/mcb.11.4.1846 ◽

1991 ◽

Vol 11 (4) ◽

pp. 1846-1853 ◽

Cited By ~ 59

Author(s):

L Dobens ◽

K Rudolph ◽

E M Berger

Keyword(s):

Regulatory Element ◽

Regulatory Elements ◽

Upstream Region ◽

Transcriptional Activators ◽

Reporter Plasmid ◽

Bacterial Gene ◽

Simplex Virus ◽

Very High ◽

Cat Gene ◽

Cat Expression

A synthetic, 23-bp ecdysterone regulatory element (EcRE), derived from the upstream region of the Drosophila melanogaster hsp27 gene, was inserted adjacent to the herpes simplex virus thymidine kinase promoter fused to a bacterial gene for chloramphenicol acetyltransferase (CAT). Hybrid constructs were transfected into Drosophila S3 cells and assayed for ecdysterone-inducible CAT expression. In the absence of ecdysterone a tandem pair of EcREs repressed the high constitutive level of CAT activity found after transfection with the parent reporter plasmid alone. After hormone addition very high levels of CAT activity were observed. Insertion of the EcRE pair 3' of the CAT gene also led to high levels of ecdysterone-induced CAT expression, but the repression of high constitutive levels of CAT activity failed to occur. The EcRE-CAT construct was cotransfected with plasmids containing tandem 10-mers or 40-mers of the EcRE but lacking a reporter gene. These additional EcREs led to a reduced level of ecdysterone-induced CAT activity and to an elevation of basal CAT activity in the absence of hormone. The data suggest that the receptor binds to the EcRE in the absence of hormone, blocking basal transcription from a constitutive promoter. In the presence of ecdysterone, receptor-hormone binding to the EcRE leads to greatly enhanced transcription.

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