Sp1 and Smad proteins cooperate to mediate transforming growth factor-β1-induced α2(I) collagen expression in human glomerular mesangial cells

被引:203
作者
Poncelet, AC
Schnaper, HW
机构
[1] Northwestern Univ, Sch Med, Dept Pediat, Chicago, IL 60611 USA
[2] Childrens Mem Inst Educ & Res, Chicago, IL 60611 USA
关键词
D O I
10.1074/jbc.M006442200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The mechanism(s) by which Smads mediate and modulate the transforming growth factor (TGF)-beta signal transduction pathway in fibrogenesis are not well characterized. We previously showed that Smads promotes alpha2(I) collagen gene (COL1A2) activation in human glomerular mesangial cells, potentially contributing to glomerulosclerosis. Here, we report that Sp1 binding is necessary for TGF-beta1-induced type I collagen mRNA expression. Deletion of three Spl sites (GC box) between -376 and -268 or mutation of a CAGA box at -268/-260 inhibited TGF-beta1-induced alpha2(I) collagen promoter activity. TGF-beta1 inducibility was also blocked by a SmadS dominant negative mutant, Chemical inhibition of Spl binding with mithramycin A, or deletion of the GC boxes, inhibited COLIA2 activation by SmadS, suggesting cooperation between SmadS and Spl in the TGF-beta1 response. Electrophoretic mobility shift assay showed that Spl and Smads form complexes with -283/-250 promoter sequences. Coimmunoprecipitation experiments demonstrate that endogenous Spl, Smad3, and Smad4 form complexes in mesangial cells. In a Gal4-LUC reporter assay system, Spl stimulated the TGF-beta1-induced transcriptional activity of Gal4-Smads, Gal4-Smad4 (266-552), or both. Using the transactivation domain B of Spl fused to the Gal4 DNA binding domain, we show that, in our system, the transcriptional activity of this Spl domain is not regulated by TGF-beta1, but it becomes responsive to this factor when Smad8 is coexpressed. Finally, combined Spl and Smads overexpression induces marked ligand-independent and ligand-dependent promoter activity of COLIA2, Thus, Spl and Smad proteins form complexes and their synergy plays an important role in mediating TGF-beta1-induced alpha2(I) collagen expression in human mesangial cells.
引用
收藏
页码:6983 / 6992
页数:10
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共 66 条
  • [61] Transforming growth factor-β1 stimulates protein kinase A in mesangial cells
    Wang, LW
    Zhu, YQ
    Sharma, K
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 1998, 273 (14) : 8522 - 8527
  • [62] Wong C, 1999, MOL CELL BIOL, V19, P1821
  • [63] Human Smad3 and Smad4 are sequence-specific transcription activators
    Zawel, L
    Dai, JL
    Buckhaults, P
    Zhou, SB
    Kinzler, KW
    Vogelstein, B
    Kern, SE
    [J]. MOLECULAR CELL, 1998, 1 (04) : 611 - 617
  • [64] Synergistic cooperation between Sp1 and Smad3/Smad4 mediates transforming growth factor β1 stimulation of α2(I)-collagen (COL1A2) transcription
    Zhang, W
    Ou, JW
    Inagaki, Y
    Greenwel, P
    Ramirez, F
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 2000, 275 (50) : 39237 - 39245
  • [65] Smad3 and Smad4 cooperate with c-Jun/c-Fos to mediate TGF-β-induced transcription
    Zhang, Y
    Feng, XH
    Derynck, R
    [J]. NATURE, 1998, 394 (6696) : 909 - 913
  • [66] Characterization of human FAST-1, a TGFβ and activin signal transducer
    Zhou, S
    Zawel, L
    Lengauer, C
    Kinzler, KW
    Vogelstein, B
    [J]. MOLECULAR CELL, 1998, 2 (01) : 121 - 127