Modulation of PLAGL2 transactivation by positive cofactor 2 (PC2), a component of the ARC/Mediator complex

Abstract

The pleomorphic adenoma gene (PLAG) family of transcription factors regulate a wide-range of physiological processes, including cell proliferation, tissue-specific gene regulation, and embryonic development, although little is known regarding the mechanisms that regulate PLAG protein activity. In this study, a yeast two-hybrid screen identified PC2, a component of the Mediator complex, as a PLAGL2-binding protein. We show that PC2 cooperates with PLAGL2 and PU.1 to enhance the activity of a known PLAGL2 target promoter (NCF2). The PLAGL2 binding element in the NCF2 promoter consisted of the core sequence of the bipartite PLAG1 consensus site, but lacked the Gcluster motif, and was recognized by PLAGL2 zinc fingers 5 and 6. Promoter and PLAGL2 mutants showed that PLAGL2 and PU.1 were required to bind to their respective sites in the promoter, and PC2 knockdown demonstrated that PC2 was essential for enhanced promoter activity. Coimmunoprecipitation and promoter-reporter studies reveal that the effect of PC2 on PLAGL2 target promoter activity was conferred via the C-terminus of PLAGL2, the region that is required for PC2 binding and contains the PLAGL2 activation domain. Importantly, chromatin immunoprecipitation analysis and PC2 knockdown studies confirmed that endogenous PC2 protein associated with the NCF2 promoter in MM1 cells in the region occupied by PLAGL2, and was required for PLAGL2 target promoter activity in TNF-α-treated MM1 cells, respectively. Lastly, the expression of another known PLAGL2 target gene, insulin-like growth factor II (IGF-II), was greatly diminished in the presence of PC2 siRNA. Together, the data identify PC2 as a novel PLAGL2-binding protein and important mediator of PLAGL2 transactivation.

DNA-methylation-mediated silencing of miR-486-5p promotes colorectal cancer proliferation and migration through activation of PLAGL2/IGF2/β-catenin signal pathways

“…Pleomorphic adenoma gene-like 2 (PLAGL2), a member of the pleomorphic adenoma gene (PLAG)-family, is a zinc-finger transcription factor that is located in the nucleus 15 , 16 . PLAGL2 was initially found in mouse cell lines 17 .…”

Studying the mechanism of PLAGL2 overexpression and its carcinogenic characteristics based on 3'-untranslated region in colorectal cancer

“…The pleomorphic adenoma gene like (PLAGL) gene family includes PLAGL1, PLAG1 and PLAGL2. As a zinc finger protein transcription factor, PLAGL2 possesses seven C2H2 zinc finger domains on the N-terminal ( 1 – 3 ). PLAGL2 shows similar DNA-binding affinity to the homologous PLAG1, which serves a carcinogenic function in tumor development ( 4 ).…”

The PLAGL2/MYCN/miR-506-3p interplay regulates neuroblastoma cell fate and associates with neuroblastoma progression

“…Here we identified for the first time PLAGL2 as a direct target of miR-506-3p in neuroblastoma cells, expanding the miRNA regulatome of PLAGL2 expression. Additional mechanisms that regulate PLAGL2 expression and functions, including gene amplification and DNA insertion [44,49], posttranslational modifications of PLAGL2 protein [53,54] and regulation of PLAGL2 activity by PLAGL2-binding proteins [55], have been reported in other cell types but have not been investigated in neuroblastoma. Overall, the molecular mechanisms that regulate PLAGL2 expression and function have been demonstrated to be complex, and future effort is certainly needed to fully dissect the molecular network involved in modulating PLAGL2 expression and functions in neuroblastoma.…”