HEPATIC NUCLEAR FACTOR-3 AND HIGH-MOBILITY GROUP I Y PROTEINS BIND THE INSULIN-RESPONSE ELEMENT OF THE INSULIN-LIKE GROWTH FACTOR-BINDING PROTEIN-1 PROMOTER/

The insulin response element (IRE) of the human insulin-like growth fa ctor-binding protein-1 (IGFBP-1) promoter contains a palindrome of the T(A/G)TIT sequence crucial to hormonal regulation of many genes. In i nitial studies of how this IRE participates in hormonal regulation, th e electromobility shift assay was used under a variety of conditions t o identify IRE-binding proteins. An exhaustive search identified five proteins that specifically bind this IRE; purified proteins were used to show that all five are related to either the high mobility group I/ Y (HMGI/Y) or hepatic nuclear factor 3 (HNF3) protein families. Furthe r studies used purified HNF3 and HMGI proteins to show: 1) each protec ts the IGFBP-1 IRE from deoxyribonuclease I (DNaseI) digestion; and 2) HNF3 but not HMGI/Y binds to the related phosphoenolpyruvate carboxyk inase and Apo CIII IREs. A series of IRE mutants with variable respons iveness to insulin were used to show that the presence of a TGTTT sequ ence in the mutants did parallel, but HMGI/Y and HNF3 binding to the m utants did not parallel, the ability of the mutants to confer the inhi bitory effect of insulin. In contrast, HNF3 binding to these IRE mutan ts roughly correlates with response of the mutants to glucocorticoids. The way by which HNF3 and/or other as yet unidentified IRE-binding pr oteins confer insulin inhibition to IGFBP-1 transcription and the role of HMGI/Y in IRE function have yet to be established.