Article

Stability of the Sp3-DNA complex is promoter-specific: Sp3 efficiently competes with Sp1 for binding to promoters containing multiple Sp-sites.

Center for Molecular Biology of Oral Diseases, College of Dentistry (M/C 860), University of Illinois at Chicago, 801 South Paulina Street, Chicago, IL 60612, USA.
Nucleic Acids Research (Impact Factor: 8.81). 10/2003; 31(18):5368-76.
Source: PubMed

ABSTRACT The transcription regulatory protein Sp3 shares more than 90% sequence homology with Sp1 in the DNA-binding domain and they bind to the same cognate DNA-element. However, the transcriptional activities of these two Sp-family factors are not equivalent. While Sp1 functions strictly as a transcriptional activator, Sp3 has been shown to be transcriptionally inactive for promoters containing multiple Sp-binding sites. In the present study, we show that the DNA-binding property of Sp3 is promoter dependent and is different from Sp1. The 116 kDa Sp3 polypeptide binds as a monomer to a single Sp-binding site but readily forms slower migrating complexes with adjacent Sp-binding sites. The slower migrating Sp3-DNA complexes are significantly more stable than monomeric Sp3-DNA complexes or multimeric Sp1-DNA complexes. As a consequence, Sp3 can efficiently compete with Sp1 for binding to regions containing multiple Sp sites. The transcription regulatory function of Sp3 is also significantly different from Sp1. Unlike Sp1, Sp3 does not synergistically activate transcription of promoters containing multiple Sp-binding sites. Therefore, although Sp3 is a transcription activator, Sp3 reduces Sp1-dependent transcription of promoters containing adjacent Sp-binding sites by competing with Sp1 for promoter occupancy and thereby blocking the synergistic transactivation function of Sp1. Taken together, this study provides a possible mechanism of the promoter-specific transcription repression function of Sp3.

0 Followers
 · 
71 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: In cultured human colonic epithelial cells and mouse colonic tissue, exposure to the common food additive carrageenan leads to inflammation, activation of Wnt signaling, increased Wnt9A expression, and decline in the activity of the enzyme arylsulfatase B (ARSB; N-acetylgalactosamine-4-sulfatase). In this study, the novel transcriptional mechanism by which carrageenan and decline in ARSB increase Wnt9A expression in NCM460 and HT-29 human colonic epithelial cells and in mouse colon is presented. Increased expression of Wnt9A has been associated with multiple malignancies, including colon carcinoma, and with ectodermal and mesoendodermal morphogenesis. When ARSB activity was reduced by siRNA or by exposure to carrageenan (1 μg/ml x 24 h), degradation of chondroitin-4-sulfate (C4S) was inhibited, leading to accumulation of more highly sulfated C4S which binds less galectin-3, a β-galactoside binding protein. Nuclear galectin-3 increased and mediated increased binding of Sp1 to the Sp1 consensus sequence in the Wnt9A promoter, shown by oligonucleotide binding assay and by chromatin immunoprecipitation assay. When galectin-3 was silenced, the increases in Sp1 binding to the Wnt9A promoter following carrageenan or ARSB silencing and in Wnt9A expression were inhibited. Mithramycin A, a specific inhibitor of Sp1 oligonucleotide binding, blocked the carrageenan- and ARSB siRNA-induced increases in Wnt9A expression. These studies reveal how carrageenan exposure can lead to transcriptional events in colonic epithelial cells through decline in arylsulfatase B activity, with subsequent impact on C4S, galectin-3, Sp1, and Wnt9A and can exert significant effects on Wnt-initiated signaling and related vital cell processes.
    Journal of Biological Chemistry 04/2014; 289(25). DOI:10.1074/jbc.M114.561589 · 4.60 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Although structurally and biochemically similar to the cellular prion (PrP(C)), doppel (Dpl) is unique in its biological functions. There are no reports about any neurodegenerative diseases induced by Dpl. However the artificial expression of Dpl in the PrP-deficient mouse brain causes ataxia with Purkinje cell death. Abundant Dpl proteins have been found in testis and depletion of the Dpl gene (Prnd) causes male infertility. Therefore, we hypothesize different regulations of Prnd in the nerve and male productive systems. In this study, by electrophoretic mobility shift assays we have determined that two different sets of transcription factors are involved in regulation of the Prnd promoter in mouse neuronal N2a and GC-1 spermatogenic (spg) cells, i.e., upstream stimulatory factors (USF) in both cells, Brn-3 and Sp1 in GC-1 spg cells, and Sp3 in N2a cells, leading to the expression of Dpl in GC-1 spg but not in N2a cells. We have further defined that, in N2a cells, Dpl induces oxidative stress and apoptosis, which stimulate ataxia-telangiectasia mutated (ATM)-modulating bindings of transcription factors, p53 and p21, to Prnp promoter, resulting the PrP(C) elevation for counteraction of the Dpl cytotoxicity; in contrast, in GC-1 spg cells, phosphorylation of p21 and N-terminal truncated PrP may play roles in the control of Dpl-induced apoptosis, which may benefit the physiological function of Dpl in the male reproduction system.
    PLoS ONE 12/2013; 8(12):e82130. DOI:10.1371/journal.pone.0082130 · 3.53 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The objective of this study was to investigate whether expression of xylosyltransferase-1 (XT-1), a key enzyme in glycosaminoglycan biosynthesis, is responsive to disk degeneration and to inhibition by the inflammatory cytokines tumor necrosis factor α and IL-1β in nucleus pulposus (NP) cells. Analysis of human NP tissues showed that XT-1 expression is unaffected by degeneration severity; XT-1 and c-Jun, c-Fos, and Sp1 mRNA were positively correlated. Cytokines failed to inhibit XT-1 promoter activity and expression. However, cytokines decreased activity of XT-1 promoters containing deletion and mutation of the -730/-723 bp AP-1 motif, prompting us to investigate the role of AP-1 and Sp1/Sp3 in the regulation of XT-1 in healthy NP cells. Overexpression and suppression of AP-1 modulated XT-1 promoter activity. Likewise, treatment with the Sp1 inhibitors WP631 and mithramycin A or cotransfection with the plasmid DN-Sp1 decreased XT-1 promoter activity. Inhibitors of AP-1 and Sp1 and stable knockdown of Sp1 and Sp3 resulted in decreased XT-1 expression in NP cells. Genomic chromatin immunoprecipitation analysis showed AP-1 binding to motifs located at -730/-723 bp and -684/-677 bp and Sp1 binding to -227/-217 bp and -124/-114 bp in XT-1 promoter. These results suggest that XT-1 expression is refractory to the disease process and to inhibition by inflammatory cytokines and that signaling through AP-1, Sp1, and Sp3 is important in the maintenance of XT-1 levels in NP cells. Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.
    American Journal Of Pathology 12/2014; 185(2). DOI:10.1016/j.ajpath.2014.09.021 · 4.60 Impact Factor

Preview

Download
0 Downloads
Available from