HIV-1 Tat regulates the SOD2 basal promoter by altering Sp1/Sp3 binding activity.
Regulation of the basal manganese superoxide dismutase (SOD2) promoter depends on the transcriptional activity of the Sp family of transcription factors. Here we report that reduced expression in the presence of Tat is independent of induction with Tumor necrosis factor alpha and that Tat affects the interaction of Sp1 and Sp3 with the basal promoter. Footprinting and electrophoretic mobility shift assay (EMSA) analyses with extracts from HeLa cells showed that Sp1/Sp3 complexes populate the proximal SOD2 promoter, and that Tat leads to an increase in the binding activity of Sp3. In Drosophila S2 cells, both Sp1 and Sp3 activated the basal SOD2 promoter (88.1 +/- 39.4 fold vs. 10.3 +/- 3.5 fold, respectively), demonstrating a positive, yet lower transcriptional regulatory function for Sp3. Additionally, the inability of Sp3 to synergistically affect promoter activity indicates an efficient competition of Sp3 with Sp1 for the multiple Sp binding sites in the SOD2 basal promoter. Tat potentiated both Sp1 and Sp3 activation of the promoter in S2 cells, though the activity of Sp3 was still lower than that of Sp1. Thus, the consequence of a shift by Tat to increased Sp3-containing complexes on the basal SOD2 promoter is decreased SOD2 expression. Together, our studies demonstrate the functional importance of the interaction of Sp1, Sp3, and Tat, revealing a possible mechanism for the attenuation of basal manganese superoxide dismutase expression.
Marecki
John C
JC
Cardiovascular Pulmonary Research Laboratory, University of Colorado Health Sciences Center, Denver, CO 80262, USA. John.Marecki@UCHSC.edu
Cota-Gomez
Adela
A
Vaitaitis
Gisela M
GM
Honda
Jennifer R
JR
Porntadavity
Sureerut
S
St Clair
Daret K
DK
Flores
Sonia C
SC
D000111:Acetylcysteine / Q000737:chemistry; D000818:Animals; D001665:Binding Sites; D015152:Blotting, Northern; D015153:Blotting, Western; D002454:Cell Differentiation; D002460:Cell Line; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004330:Drosophila; D005609:Free Radicals; D015967:Gene Expression Regulation, Viral; D015696:Gene Products, tat / Q000235:genetics* / Q000502:physiology*; D015497:HIV-1 / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D008956:Models, Chemical; D018384:Oxidative Stress; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D051705:Sp3 Transcription Factor; D013482:Superoxide Dismutase / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D054322:tat Gene Products, Human Immunodeficiency Virus; D000099156:Superoxide Dismutase 2
D000111:Acetylcysteine; D004247:DNA; D004268:DNA-Binding Proteins; D005609:Free Radicals; D015696:Gene Products, tat; D008156:Luciferases; D016329:Sp1 Transcription Factor; D051705:Sp3 Transcription Factor; D013482:Superoxide Dismutase; D014157:Transcription Factors; D054322:tat Gene Products, Human Immunodeficiency Virus; D000099156:Superoxide Dismutase 2; C497121:SP3 protein, human