Part:BBa_K2591014

TCF-TATA-GFP

TCF-TATA-GFP

Usage and Biology

The WNT signaling pathway is one of the cellular signal transduction pathways, which begins with reception of WNT ligands in plasma membrane, and initiate a series of downstream events. If there is no reception of WNT ligands, the subunits of disruption complex is free, and it can bind the downstream protein β-catenin in cytoplasm, mediating its degradation, which means there is no more downstream events.(Fig.1)

Figure1. Condition of no WNT signal However, when WNT ligands bind to the receptor Frizzled, the disruption complex is fixed by WNT-Frizzled in plasma membrane, which leads to release and accumulation of β-catenin. Then, the β-catenin can finally translocated into the nucleus and acts as transcriptional co-activator of transcriptional factor TCF. The β-catenin-TCF complex then interacts with TCF transcriptional response element(TCF TRE) and start the transcription of WNT target genes.(Fig.2)

Figure2. Condition of WNT on In the absence of β-catenin, TCF acts as a repressor of WNT target genes by occupying the TCF TRE, but β-catenin can convert the TCF to a transcriptional activator of the same genes. This novel process can be extract from nature and make up a reporter system when we replace the target genes into a fluorescence gene like GFP. When this whole construct stably insert to a receptor cell line, it can sense WNT signal and give green fluorescence.(Fig.3)

Figure3. GFP followed by TCF TRE

Reference

      Akiri, G., Cherian, M. M., Vijayakumar, S., Liu, G., Bafico, A., & Aaronson, S. A. (2009). Wnt pathway aberrations including autocrine Wnt activation occur at high frequency in human non-small-cell lung carcinoma. Oncogene, 28(21), 2163.
      Blauwkamp, T. A., Nigam, S., Ardehali, R., Weissman, I. L., & Nusse, R. (2012). Endogenous Wnt signalling in human embryonic stem cells generates an equilibrium of distinct lineage-specified progenitors. Nature communications, 3, 1070.
      LEF, G. W., & GFP, T. D. (2002). LEF/TCF-Dependent, Fluorescence-Based Re-porter Gene Assay For Wnt Signaling. BioTechniques, 32, 1000-1002.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Unknown
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 56
  Illegal BamHI site found at 42
  Illegal XhoI site found at 52
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 120
  
• 1000
  COMPATIBLE WITH RFC[1000]