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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)

Multiple Sp1 sites efficiently drive transcription of the TATA-less promoter of the human glypican 3 (GPC3) gene.

Simpson-Golabi-Behmel Syndrome (SGBS) is an X-linked disease characterized by pre- and postnatal overgrowth. Recently, we have shown that mutations in the glypican family gene, GPC3, cause SGBS. This gene is predominantly expressed in the same mesoderm-derived tissues that overgrow in its absence. To investigate the basis for promoter function, 3.3kb of GC-rich DNA 5' of the transcribed region were fused to a luciferase cDNA, transfected into Caco-2 and NT2 cells, and assayed for activity. Deletion analysis identified a 218-bp fragment upstream of the transcription start site that conferred more than 80% of maximal reporter gene activation. This fragment contains five putative Sp1 binding sites, three of which (centered at nt -14, -34, and -92) were active when assessed by DNaseI footprinting and gel shift/supershift assays. Additionally, Sp1 specifically transactivated transcription in Sp1-deficient Drosophila SL2 cells, demonstrating the functionality of Sp1 on the GPC3 promoter. A full-length promoter construct was also highly active in HeLa cells, which do not express endogenous GPC3. These results indicate that the GPC3 promoter is dependent on Sp1 for proper activation, but tissue-specific repression in non-expressing cells must involve either DNA that lies outside the region tested or auxiliary structural features of chromatin.[1]


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