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Regulation of MMP-9 by p53 in first trimester cytotrophoblastic cells.

Cohen M, Wuillemin C, Irion O, Bischof P - Hum. Reprod. (2008)

Bottom Line: However, endogenous p53 is not able to regulate MMP-9 expression in CTB.The presence of high molecular weight complexes of p53 in CTB suggests a potential mechanism of inactivation of p53 transcriptional activity towards MMPs in these cells.Although p53 is mutated in trophoblast, it is functionally incompetent towards MMPs in these cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynaecology, Laboratory of Hormonology, University of Geneva, Geneva, Switzerland. marie.cohen@hcuge.ch

ABSTRACT

Background: The matrix metalloproteinase (MMP) family is known to play a key role in tissue remodelling during embryonic development and in pathological conditions, such as cardiovascular disease, arthritis and cancer metastasis. It has been shown previously that p53 regulates positively or negatively the expression of different MMPs. Because of p53 overexpression in trophoblastic cells, and its potential role in regulating MMP-2 and MMP-9 expression in different cell lines, we hypothesized that the expression of MMP-9 could also be regulated by p53 in first trimester cytotrophoblasts (CTB).

Methods and results: Transfection experiments in CTB demonstrated that wild-type p53 down-regulates the -670 (P < 0.001) but not the -531 and -90 human MMP-9 promoter/CAT reporter plasmid activity, whereas p53 mutants partially lost this repressive activity. However, endogenous p53 is not able to regulate MMP-9 expression in CTB. The presence of high molecular weight complexes of p53 in CTB suggests a potential mechanism of inactivation of p53 transcriptional activity towards MMPs in these cells.

Conclusions: Although p53 is mutated in trophoblast, it is functionally incompetent towards MMPs in these cells.

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Related in: MedlinePlus

Determination of the minimal promoter sequence required for p53 repression.CTB were transfected with 3 µg of −670, −531 or −90 hMMP-9 promoter/CAT reporter plasmid and 0.5 µg wt-p53 or control vector. After 48 h, cells were harvested and CAT activity was assayed and normalized to the amount of proteins in the cellular lysates. Three independent experiments, each run in triplicate, were performed and the results expressed as mean ± SEM. Paired Student's t-test was used to compare transfected cells with wt-p53 versus control plasmids.
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DEN264F3: Determination of the minimal promoter sequence required for p53 repression.CTB were transfected with 3 µg of −670, −531 or −90 hMMP-9 promoter/CAT reporter plasmid and 0.5 µg wt-p53 or control vector. After 48 h, cells were harvested and CAT activity was assayed and normalized to the amount of proteins in the cellular lysates. Three independent experiments, each run in triplicate, were performed and the results expressed as mean ± SEM. Paired Student's t-test was used to compare transfected cells with wt-p53 versus control plasmids.

Mentions: Transcription assays were performed with CAT reporter driven by −670 promoter of the hMMP-9 gene cotransfected with the expression vector of p53 in CTB. As shown in Fig. 2, wt-p53 and -p53 mutants commonly found in human cancer, p53-175H and p53-143A, are able to down-regulate −670 promoter activity. CAT activity is significantly reduced by wt-p53 and by p53-175H (∼50%), whereas p53-143A has partially lost the ability to repress the hMMP-9 promoter activity (∼25%). In order to map the site(s) responsible for p53-mediated repression, CTB have been cotransfected with wt-p53 expression plasmid or control vector and various 5′ deletions of hMMP-9/CAT promoter. As shown in Fig. 3, only −670 hMMP-9 CAT activity is significantly inhibited by wt-p53. So, cis-acting elements involved in the p53-induced down-regulation of hMMP-9 gene expression are localized between −670 and −531 bp of the 5′ flanking region of hMMP-9 gene.


Regulation of MMP-9 by p53 in first trimester cytotrophoblastic cells.

Cohen M, Wuillemin C, Irion O, Bischof P - Hum. Reprod. (2008)

Determination of the minimal promoter sequence required for p53 repression.CTB were transfected with 3 µg of −670, −531 or −90 hMMP-9 promoter/CAT reporter plasmid and 0.5 µg wt-p53 or control vector. After 48 h, cells were harvested and CAT activity was assayed and normalized to the amount of proteins in the cellular lysates. Three independent experiments, each run in triplicate, were performed and the results expressed as mean ± SEM. Paired Student's t-test was used to compare transfected cells with wt-p53 versus control plasmids.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2538584&req=5

DEN264F3: Determination of the minimal promoter sequence required for p53 repression.CTB were transfected with 3 µg of −670, −531 or −90 hMMP-9 promoter/CAT reporter plasmid and 0.5 µg wt-p53 or control vector. After 48 h, cells were harvested and CAT activity was assayed and normalized to the amount of proteins in the cellular lysates. Three independent experiments, each run in triplicate, were performed and the results expressed as mean ± SEM. Paired Student's t-test was used to compare transfected cells with wt-p53 versus control plasmids.
Mentions: Transcription assays were performed with CAT reporter driven by −670 promoter of the hMMP-9 gene cotransfected with the expression vector of p53 in CTB. As shown in Fig. 2, wt-p53 and -p53 mutants commonly found in human cancer, p53-175H and p53-143A, are able to down-regulate −670 promoter activity. CAT activity is significantly reduced by wt-p53 and by p53-175H (∼50%), whereas p53-143A has partially lost the ability to repress the hMMP-9 promoter activity (∼25%). In order to map the site(s) responsible for p53-mediated repression, CTB have been cotransfected with wt-p53 expression plasmid or control vector and various 5′ deletions of hMMP-9/CAT promoter. As shown in Fig. 3, only −670 hMMP-9 CAT activity is significantly inhibited by wt-p53. So, cis-acting elements involved in the p53-induced down-regulation of hMMP-9 gene expression are localized between −670 and −531 bp of the 5′ flanking region of hMMP-9 gene.

Bottom Line: However, endogenous p53 is not able to regulate MMP-9 expression in CTB.The presence of high molecular weight complexes of p53 in CTB suggests a potential mechanism of inactivation of p53 transcriptional activity towards MMPs in these cells.Although p53 is mutated in trophoblast, it is functionally incompetent towards MMPs in these cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynaecology, Laboratory of Hormonology, University of Geneva, Geneva, Switzerland. marie.cohen@hcuge.ch

ABSTRACT

Background: The matrix metalloproteinase (MMP) family is known to play a key role in tissue remodelling during embryonic development and in pathological conditions, such as cardiovascular disease, arthritis and cancer metastasis. It has been shown previously that p53 regulates positively or negatively the expression of different MMPs. Because of p53 overexpression in trophoblastic cells, and its potential role in regulating MMP-2 and MMP-9 expression in different cell lines, we hypothesized that the expression of MMP-9 could also be regulated by p53 in first trimester cytotrophoblasts (CTB).

Methods and results: Transfection experiments in CTB demonstrated that wild-type p53 down-regulates the -670 (P < 0.001) but not the -531 and -90 human MMP-9 promoter/CAT reporter plasmid activity, whereas p53 mutants partially lost this repressive activity. However, endogenous p53 is not able to regulate MMP-9 expression in CTB. The presence of high molecular weight complexes of p53 in CTB suggests a potential mechanism of inactivation of p53 transcriptional activity towards MMPs in these cells.

Conclusions: Although p53 is mutated in trophoblast, it is functionally incompetent towards MMPs in these cells.

Show MeSH
Related in: MedlinePlus