Angiotensin II–Inducible Platelet-Derived Growth Factor-D Transcription Requires Specific Ser/Thr Residues in the Second Zinc Finger Region of Sp1

• Published in: Circulation research 2008-02, Vol.102 (4), p.e38-e51
• Main Authors: Tan, Nicole Y, Midgley, Valerie C, Kavurma, Mary M, Santiago, Fernando S, Luo, Xiao, Peden, Ryan, Fahmy, Roger G, Berndt, Michael C, Molloy, Mark P, Khachigian, Levon M
• Format: Article
• Language: English
• Subjects: Angiotensin II - pharmacology; Animals; Antibodies - pharmacology; Carotid Artery Diseases - metabolism; Carotid Artery Diseases - physiopathology; Catheterization - adverse effects; Cells, Cultured; Disease Models, Animal; Humans; Lymphokines - genetics; Muscle, Smooth, Vascular - cytology; Muscle, Smooth, Vascular - drug effects; Muscle, Smooth, Vascular - metabolism; Mutagenesis; Phosphorylation - drug effects; Platelet-Derived Growth Factor - genetics; Promoter Regions, Genetic; Protein Kinase C - metabolism; Protein Structure, Tertiary; Rats; Rats, Sprague-Dawley; Serine - metabolism; Sp1 Transcription Factor - chemistry; Sp1 Transcription Factor - immunology; Sp1 Transcription Factor - metabolism; Spectrometry, Mass, Electrospray Ionization; Threonine - metabolism; Transcription, Genetic - drug effects; Transcription, Genetic - physiology; Vasoconstrictor Agents - pharmacology; Zinc Fingers - physiology
• ISSN: 0009-7330; 1524-4571
• DOI: 10.1161/CIRCRESAHA.107.167395

Sp1, the first identified and cloned transcription factor, regulates gene expression via multiple mechanisms including direct protein–DNA interactions, protein–protein interactions, chromatin remodeling, and maintenance of methylation-free CpG islands. Sp1 is itself regulated at different levels, for example, by glycosylation, acetylation, and phosphorylation by kinases such as the atypical protein kinase C-ζ. Although Sp1 controls the basal and inducible regulation of many genes, the posttranslational processes regulating its function and their relevance to pathology are not well understood. Here we have used a variety of approaches to identify 3 amino acids (Thr668, Ser670, and Thr681) in the zinc finger domain of Sp1 that are modified by PKC-ζ and have generated novel anti-peptide antibodies recognizing the PKC-ζ–phosphorylated form of Sp1. Angiotensin II, which activates PKC-ζ phosphorylation (at Thr410) via the angiotensin II type 1 receptor, stimulates Sp1 phosphorylation and increases Sp1 binding to the platelet-derived growth factor-D promoter. All 3 residues in Sp1 (Thr668, Ser670, and Thr681) are required for Sp1-dependent platelet-derived growth factor-D activation in response to angiotensin II. Immunohistochemical analysis revealed that phosphorylated Sp1 is expressed in smooth muscle cells of human atherosclerotic plaques and is dynamically expressed together with platelet-derived growth factor-D in smooth muscle cells of the injured rat carotid artery wall. This study provides new insights into the regulatory mechanisms controlling the PKC-ζ–phospho-Sp1 axis and angiotensin II–inducible gene expression.