IDENTIFICATION AND CHARACTERIZATION OF BASAL AND CYCLIC-AMP RESPONSE ELEMENTS IN THE PROMOTER OF THE RAT HEXOKINASE-II GENES

Hexokinases catalyze the phosphorylation of glucose and initiate cellu lar glucose metabolism. Hexokinase II (HKII) is the principal hexokina se isoform in skeletal muscle, heart, and adipose tissue. Isoprotereno l and exogenous cyclic AMP (cAMP) increase HKII gene transcription in L6 myotubes. Various segments of the HKII promoter that direct the exp ression of the chloramphenicol acetyltransferase reporter gene were tr ansfected into L6 myotubes to identify basal and cAMP response element s. The 5'-flanking region that extends 90 base pairs upstream of the t ranscription start site includes a CCAAT box and a cAMP response eleme nt (CRE); both contribute to basal promoter activity and each provides an independent, maximal response 60 cAMP. An inverted CCAAT motif, or Y box, located just upstream of the CCAAT box, contributes to basal p romoter activity but is not involved in the cAMP response. Home- and h eterodimers composed of the CRE-binding protein and activating transcr iption factor-1 bind specifically to the CRE. The Y box and the CCAAT box specifically bind the factor NF-Y (also known as CBF).