Functional domains of the TGF-beta-inducible transcription factor Tieg3 and detection of two putative nuclear localization signals within the zinc finger DNA-binding domain.
The recently identified TGF-beta-inducible early gene 3 (Tieg3) belongs to the gene family of Sp1/Klf-like transcription factors and is upregulated immediately after TGF-beta treatment. To explore the molecular mechanisms of Tieg3-mediated transcriptional control, GAL4-based luciferase assays were performed in order to determine regulatory domains within the Tieg3 protein. Using EGFP-fusion proteins, we monitored the intracellular localization and mapped putative nuclear localization signals (NLS). We provide evidence that the amino-terminus of Tieg3 is essential to repress the transcription and that the loss of the mSin3A interacting domain (SID) disrupts the repressive effects of Tieg3 in the oligodendroglial cell line OLI-neu. Herein we also demonstrate that the zinc finger containing DNA-binding domain (DBD) alone is able to activate the transcription of a reporter gene. Sequence analysis of the zinc finger region revealed no similarities to known activation domains. Analysis of the subcellular localization disclosed Tieg3 as a nuclear protein. Further, we identified the DBD as being essential for the nuclear localization of Tieg3. We detected two closely located putative bipartite NLS within the second and third zinc finger, which are conserved among the members of the Tieg family of proteins. Together these results may help to increase the understanding of Tieg3-mediated transcriptional control and to characterize this TGF-beta-induced Sp1/Klf-like transcription factor.
Spittau
Björn
B
Center of Anatomy, Department of Neuroanatomy, University of Göttingen, Kreuzbergring, Germany. bspitta@gwdg.de
Wang
Ziyuan
Z
Boinska
Dagmara
D
Krieglstein
Kerstin
K
D000595:Amino Acid Sequence; D051017:Apoptosis Regulatory Proteins; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D016335:Zinc Fingers / Q000235:genetics*
D051017:Apoptosis Regulatory Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C115117:KLF11 protein, human; D019913:Nuclear Localization Signals; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins