Upstream stimulatory factor (USF) and neurogenic differentiation/beta-cell E box transactivator 2 (NeuroD/BETA2) contribute to islet-specific glucose-6-phosphatase catalytic-subunit-related protein (IGRP) gene expression.

Martin CC, Svitek CA, Oeser JK, Henderson E, Stein R, O'Brien RM
Biochem J. 2003 371 (Pt 3): 675-86

PMID: 12540293 · PMCID: PMC1223330 · DOI:10.1042/BJ20021585

Islet-specific glucose-6-phosphatase (G6Pase) catalytic-subunit-related protein (IGRP) is a homologue of the catalytic subunit of G6Pase, the enzyme that catalyses the final step of the gluconeogenic pathway. The analysis of IGRP-chloramphenicol acetyltransferase (CAT) fusion-gene expression through transient transfection of islet-derived beta TC-3 cells revealed that multiple promoter regions, located between -306 and -97, are required for maximal IGRP-CAT fusion-gene expression. These regions correlated with trans -acting factor-binding sites in the IGRP promoter that were identified in beta TC-3 cells in situ using the ligation-mediated PCR (LMPCR) footprinting technique. However, the LMPCR data also revealed additional trans -acting factor-binding sites located between -97 and +1 that overlap two E-box motifs, even though this region by itself conferred minimal fusion-gene expression. The data presented here show that these E-box motifs are important for IGRP promoter activity, but that their action is only manifest in the presence of distal promoter elements. Thus mutation of either E-box motif in the context of the -306 to +3 IGRP promoter region reduces fusion-gene expression. These two E-box motifs have distinct sequences and preferentially bind NeuroD/BETA2 (neurogenic differentiation/beta-cell E box transactivator 2) and upstream stimulatory factor (USF) in vitro, consistent with the binding of both factors to the IGRP promoter in situ, as determined using the chromatin-immunoprecipitation (ChIP) assay. Based on experiments using mutated IGRP promoter constructs, we propose a model to explain how the ubiquitously expressed USF could contribute to islet-specific IGRP gene expression.

MeSH Terms (10)

Animals Basic Helix-Loop-Helix Transcription Factors Catalytic Domain Cricetinae Gene Expression Regulation Glucose-6-Phosphatase Humans Islets of Langerhans Mice Nerve Tissue Proteins

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