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Identification of hPYGO2 Interacting Protein by Yeast Two-Hybrid

Author: LinChen
Tutor: LiBoAn
School: Xiamen University
Course: Biochemistry and Molecular Biology
Keywords: Yeast two-hybrid hPYGO2 △ NHD Wnt signaling pathway
CLC: Q78
Type: Master's thesis
Year: 2008
Downloads: 87
Quote: 1
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Abstract


The earliest Pygopus protein in Drosophila as a special component of Wg (mammalian homologous protein Wnt) signaling pathway was found, which led to its functional mechanism. Surprisingly, that has a specific function in Drosophila Wg signal transduction pathway Pygopus has a dual function in mice, and play a role in the Wnt and Wnt independent pathway. This article describes the classical Wnt signaling pathway and of this stage Pygopus protein function studies, and further analysis of the role of the the two conservative Pygopus protein domains in the transcription process. HPYGO2 △ NHD bait protein to screen a human breast library, 70 positive clones. And the sequencing results of the positive clone was sequenced using BLAST analysis of the amino acid sequence of positive clones 13,5,22,35 and CDC73 (Cell Division Cycle 73), respectively, of BCL6 (B-cell CLL / lymphoma 6), cdc34 (Cell Division Cycle 34homolog ), EEF1G (the Gamma the eukaryotic TRANSLATION elongmion factor 1) four proteins having a high degree of homology. In further experiments, we successfully constructed the mammalian expression vector p3 × Flag-CMV-hpygopus ΔNHD , pcDNA3.1-Myc-cdc73 and pCMV5-Myc-bc16 three fusion proteins in 293T cells are effective expression. Interactions between other proteins also need to continue to verify the report by Co-IP and fluorescence Su the enzyme detection experiments prove pcDNA3.1-Myc-cdc73 and p3 × Flag-CMV-hpygopus ΔNHD . Through the above experiments, we believe that the interaction between the screened four proteins in yeast and hPYGO2 △ NHD . Wherein, CDC73 in humans Parafibromin, it is a member of the PAF protein complexes involved in nuclear Wg / Wnt signal transduction process: BCL6 is a transcriptional repressor. Speculated the BCL6 protein can inhibit the expression of the p53 tumor suppressor factor gene; CDC34 is a ubiquitin ligase family, this protein is a large part of the protein complex, the degradation in the ubiquitination mediated cell cycle G1 regulators need for its participation in the process and the initiation of DNA replication. EEF1G eukaryotic transcription elongation factor, because PYGO2 transcriptional activation, guess its possible related transcription elongation. The significance of these findings to further understand the function of hPYGO2 △ NHD , eventually clarify the mechanism of the Wnt signaling pathway.

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CLC: > Biological Sciences > Molecular Biology > Genetic engineering (genetic engineering)
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