Tags

Type your tag names separated by a space and hit enter

Role of CAGA boxes in the plasminogen activator inhibitor-1 promoter in mediating oxidized low-density lipoprotein-induced transcriptional activation in mesangial cells.
Transl Res. 2007 Sep; 150(3):180-8.TR

Abstract

Oxidized low-density lipoprotein (Ox-LDL) activates transforming growth factor-beta (TGF-beta)/Smad signaling to stimulate plasminogen activator inhibitor-1 (PAI-1) expression in mesangial cells. Smad-binding sequences, termed CAGA boxes, are present in the promoter of human PAI-1 gene, and they mediate TGF-beta transcriptional induction. However, the functional role of each CAGA box in the Ox-LDL-induced PAI-1 promoter activation is unknown. In this study, mutation of 1 of the 3 CAGA boxes located at -730, -580, and -280 of the PAI-1 promoter decreased the Ox-LDL-induced luciferase activity by 40 to 58%, whereas mutations in 2 sites reduced it over 75% or completely abolished it. Overexpression of Smad3 in N-terminal tagged Smad3-transfected cells increased the Ox-LDL-induced transcriptional activation of the PAI-1 promoter, whereas mutation of Smad3 abolished it. Electrophoretic mobility shift assay showed that the labeled -280, -580, and -730 CAGA box probes detected DNA/protein complexes induced by Ox-LDL, whereas mutant probes did not. When nuclear extracts were preincubated with a 100-fold of an unlabeled -280, -580, and -730 CAGA oligonucleotide, the formation of complexes was prevented but not with mutant CAGA box competitors. The addition of anti-Smad3 to the reaction with the labeled -280 or -580 CAGA box probe resulted in a supershift, but not with the -730 CAGA box probe. These results suggest that the 3 CAGA elements in the PAI-1 promoter mediate the Ox-LDL-induced PAI-1 transcription to a different degree, of which the -280 and -580 CAGA regions directly bind to Smad3.

Authors+Show Affiliations

Department of Pathology, Seoul National University College of Medicine, Seoul, Korea.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

17761371

Citation

Kim, Bong Cho, et al. "Role of CAGA Boxes in the Plasminogen Activator Inhibitor-1 Promoter in Mediating Oxidized Low-density Lipoprotein-induced Transcriptional Activation in Mesangial Cells." Translational Research : the Journal of Laboratory and Clinical Medicine, vol. 150, no. 3, 2007, pp. 180-8.
Kim BC, Song CY, Hong HK, et al. Role of CAGA boxes in the plasminogen activator inhibitor-1 promoter in mediating oxidized low-density lipoprotein-induced transcriptional activation in mesangial cells. Transl Res. 2007;150(3):180-8.
Kim, B. C., Song, C. Y., Hong, H. K., & Lee, H. S. (2007). Role of CAGA boxes in the plasminogen activator inhibitor-1 promoter in mediating oxidized low-density lipoprotein-induced transcriptional activation in mesangial cells. Translational Research : the Journal of Laboratory and Clinical Medicine, 150(3), 180-8.
Kim BC, et al. Role of CAGA Boxes in the Plasminogen Activator Inhibitor-1 Promoter in Mediating Oxidized Low-density Lipoprotein-induced Transcriptional Activation in Mesangial Cells. Transl Res. 2007;150(3):180-8. PubMed PMID: 17761371.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Role of CAGA boxes in the plasminogen activator inhibitor-1 promoter in mediating oxidized low-density lipoprotein-induced transcriptional activation in mesangial cells. AU - Kim,Bong Cho, AU - Song,Chi Young, AU - Hong,Hye Kyoung, AU - Lee,Hyun Soon, Y1 - 2007/05/25/ PY - 2007/03/02/received PY - 2007/04/03/revised PY - 2007/04/03/accepted PY - 2007/9/1/pubmed PY - 2007/10/13/medline PY - 2007/9/1/entrez SP - 180 EP - 8 JF - Translational research : the journal of laboratory and clinical medicine JO - Transl Res VL - 150 IS - 3 N2 - Oxidized low-density lipoprotein (Ox-LDL) activates transforming growth factor-beta (TGF-beta)/Smad signaling to stimulate plasminogen activator inhibitor-1 (PAI-1) expression in mesangial cells. Smad-binding sequences, termed CAGA boxes, are present in the promoter of human PAI-1 gene, and they mediate TGF-beta transcriptional induction. However, the functional role of each CAGA box in the Ox-LDL-induced PAI-1 promoter activation is unknown. In this study, mutation of 1 of the 3 CAGA boxes located at -730, -580, and -280 of the PAI-1 promoter decreased the Ox-LDL-induced luciferase activity by 40 to 58%, whereas mutations in 2 sites reduced it over 75% or completely abolished it. Overexpression of Smad3 in N-terminal tagged Smad3-transfected cells increased the Ox-LDL-induced transcriptional activation of the PAI-1 promoter, whereas mutation of Smad3 abolished it. Electrophoretic mobility shift assay showed that the labeled -280, -580, and -730 CAGA box probes detected DNA/protein complexes induced by Ox-LDL, whereas mutant probes did not. When nuclear extracts were preincubated with a 100-fold of an unlabeled -280, -580, and -730 CAGA oligonucleotide, the formation of complexes was prevented but not with mutant CAGA box competitors. The addition of anti-Smad3 to the reaction with the labeled -280 or -580 CAGA box probe resulted in a supershift, but not with the -730 CAGA box probe. These results suggest that the 3 CAGA elements in the PAI-1 promoter mediate the Ox-LDL-induced PAI-1 transcription to a different degree, of which the -280 and -580 CAGA regions directly bind to Smad3. SN - 1931-5244 UR - https://www.unboundmedicine.com/medline/citation/17761371/Role_of_CAGA_boxes_in_the_plasminogen_activator_inhibitor_1_promoter_in_mediating_oxidized_low_density_lipoprotein_induced_transcriptional_activation_in_mesangial_cells_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1931-5244(07)00112-0 DB - PRIME DP - Unbound Medicine ER -