title: Xylosyltransferase-1 expression is refractory to inhibition by the inflammatory cytokines tumor necrosis factor α and IL-1β in nucleus pulposus cells: novel regulation by AP-1, Sp1, and Sp3.
abstract: We investigated 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 Jun, 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.
authors.affiliation: Department of Orthopaedic Surgery, Thomas Jefferson University, Philadelphia, Pennsylvania; Department of Orthopaedic Surgery, Sun Yat-sen Memorial Hospital of Sun Yat-Sen University, Guangzhou, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Ye
authors.affiliation: Department of Cancer Biology, Thomas Jefferson University, Philadelphia, Pennsylvania; Department of Surgery, Affiliated Cancer Hospital of Guangzhou Medical University, Guangzhou, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhou
authors.affiliation: Department of Orthopaedic Surgery, Thomas Jefferson University, Philadelphia, Pennsylvania.
authors.forename: Dessislava Z
authors.initials: DZ
authors.lastname: Markova
authors.affiliation: Department of Orthopaedic Surgery, Thomas Jefferson University, Philadelphia, Pennsylvania; Department of Orthopaedic Surgery, Changzheng Hospital of the Second Military Medical University, Shanghai, China.
authors.forename: Ye
authors.initials: Y
authors.lastname: Tian
authors.affiliation: Department of Orthopaedic Surgery, Thomas Jefferson University, Philadelphia, Pennsylvania; Department of Orthopaedic Surgery, Changzheng Hospital of the Second Military Medical University, Shanghai, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Li
authors.affiliation: Department of Orthopaedic Surgery, Thomas Jefferson University, Philadelphia, Pennsylvania.
authors.forename: D Greg
authors.initials: DG
authors.lastname: Anderson
authors.affiliation: Department of Orthopaedic Surgery, Thomas Jefferson University, Philadelphia, Pennsylvania.
authors.forename: Irving M
authors.initials: IM
authors.lastname: Shapiro
authors.affiliation: Department of Orthopaedic Surgery, Thomas Jefferson University, Philadelphia, Pennsylvania. Electronic address: makarand.risbud@jefferson.edu.
authors.forename: Makarand V
authors.initials: MV
authors.lastname: Risbud
mesh_terms: D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D053583:Interleukin-1beta / Q000235:genetics / Q000378:metabolism*; D055959:Intervertebral Disc Degeneration / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D010430:Pentosyltransferases / Q000096:biosynthesis* / Q000235:genetics; D051381:Rats; D020218:Response Elements; D017384:Sequence Deletion; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D051705:Sp3 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D013131:Spine; D018808:Transcription Factor AP-1 / Q000235:genetics / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism*; D000097608:UDP Xylose-Protein Xylosyltransferase
chemical_list: D053583:Interleukin-1beta; D010430:Pentosyltransferases; D016329:Sp1 Transcription Factor; D051705:Sp3 Transcription Factor; D018808:Transcription Factor AP-1; D014409:Tumor Necrosis Factor-alpha; D000097608:UDP Xylose-Protein Xylosyltransferase; C508585:IL1B protein, human; C000594089:IL1B protein, rat; C497121:SP3 protein, human; C108209:SP1 protein, human; C497123:Sp3 protein, rat