Transcription factor Sp3 is regulated by acetylation - PowerPoint PPT Presentation

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Transcription factor Sp3 is regulated by acetylation

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Harald Braun, Ronald Koop, Alexa Ertmer, Silvina Nacht and Guntram Suske ... Sp3 is a ubiquitously expressed transcription factor closely related to Sp1. ... – PowerPoint PPT presentation

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Title: Transcription factor Sp3 is regulated by acetylation


1
Transcription factor Sp3 is regulated by
acetylation Harald Braun, Ronald Koop, Alexa
Ertmer, Silvina Nacht and Guntram Suske
Institute of Molecular Biology and Tumor
Research (IMT), Philipps-University Marburg,
Emil-Mannkopff-Strasse 2, D-35037 Marburg,
Germany Received September 4, 2001 Revised and
Accepted October 12, 2001.
  • Sp3 is a ubiquitously expressed transcription
    factor closely related to Sp1.
  • Sp3 is, unlike Sp1, inactive or acts only as a
    weak activator
  • contains ID located between the 2nd Glu-rich AD
    and the Zn finger region
  • The integrity of a charged amino acid triplet
    (KEE) within this domain is absolutely essential
    for inhibitor function.
  • Mutation of these amino acids converted Sp3 to a
    strong activator that is almost indistinguishable
    from Sp1

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Expression and purification of epitope-tagged Sp3
and Sp3SD in stable transfected SL2 cells
0.5 mM CuSO4
Flag peptide
Nuclear Ex aSp3-protein A column aFLAG-protein
A column Elute with FLAG peptide
BT aSp3
4
Sp3 acts as a strong transcriptional activator in
vitro
In vitro transcription assay
Similar activity
C, GC box affinity - -
5
A single lysine residue is essential for
inhibitory function
6
Sp3 is acetylated in vivo
IP with aFLAG
1mM 3H-acetate for 1h Nuclear Ex
7
CBP and p300 but not PCAF can acetylate Sp3 in
vitro
Thrombin clevage
PCAF (amino acids 1832) CBP (amino acids
10981758) p300 (amino acids 10711715)
8
  • Discussion
  • Purified recombinant Sp3 is a strong activator in
    vitro.
  • Sp3 is acetylated in vivo
  • What might be the consequences of acetylation?
  • stimulation of DNA binding p53, E2F1, MyoD and
    Tal1 .
  • transcriptional potency is enhanced EKLF
  • Alterations of proteinprotein interactions and
    protein stability, dTCF and E2F1
  • Our finding that the critical lysine residue,
    which is essential for silencing of Sp3 activity,
    is acetylated in vivo suggests that acetylation
    is involved in the process of silencing.
  • The amino acids surrounding the lysine residue
    within the inhibitory domain of Sp3 resemble
    sites that are known to be modified by the small
    ubiquitin-like modifier (SUMO).
  • Thus, it seems not unlikely that the same lysine
    residue could be modified not only by acetylation
    but also by other modifications.
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