Interactions between areas I and II direct pdx-1 expression specifically to islet cell types of the mature and developing pancreas.

Van Velkinburgh JC, Samaras SE, Gerrish K, Artner I, Stein R
J Biol Chem. 2005 280 (46): 38438-44

PMID: 16147997 · DOI:10.1074/jbc.M508594200

PDX-1 regulates transcription of genes involved in islet beta cell function and pancreas development. Islet-specific expression is controlled by 5'-flanking sequences from base pair (bp) -2917 to -1918 in transgenic experiments, which encompasses both conserved (i.e. Area I (bp -2761/-2457), Area II (bp -2153/-1923)) and non-conserved pdx-1 sequences. However, only an Area II-driven transgene is independently active in vivo, albeit in only a fraction of islet PDX-1-producing cells. Our objective was to identify the sequences within the -2917/-1918-bp region that act in conjunction with Area II to allow comprehensive expression in islet PDX-1(+) cells. In cell line-based transfection assays, only Area I effectively potentiated Area II activity. Both Area I and Area II functioned in an orientation-independent manner, whereas synergistic, enhancer-like activation was uniquely found with duplicated Area II. Chimeras of Area II and the generally active SV40 enhancer or the beta cell-specific insulin enhancer suggested that islet cell-enriched activators were necessary for Area I activation, because Area II-mediated stimulation was reduced by the SV40 enhancer and activated by the insulin enhancer. Several conserved sites within Area I were important in Area I/Area II activation, with binding at bp -2614/-2609 specifically controlled by Nkx2.2, an insulin gene regulator that is required for terminal beta cell differentiation. The ability of Area I to modulate Area II activation was also observed in vivo, as an Area I/Area II-driven transgene recapitulated the endogenous pdx-1 expression pattern in developing and adult islet cells. These results suggest that Area II is a central pdx-1 control region, whose islet cell activity is uniquely modified by Area I regulatory factors.

MeSH Terms (32)

Animals Animals, Newborn Base Sequence Binding Sites Cell Differentiation Cell Line Cell Nucleus Chromatin Chromatin Immunoprecipitation DNA Mutational Analysis Enhancer Elements, Genetic Galactosides Homeodomain Proteins Humans Immunohistochemistry Indoles Insulin Insulinoma Islets of Langerhans Lac Operon Mice Molecular Sequence Data NIH 3T3 Cells Pancreas Protein Binding Protein Structure, Tertiary Recombinant Fusion Proteins Trans-Activators Transcription, Genetic Transcription Factors Transfection Transgenes

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