Binding of the Ets factor GA-binding protein to an upstream site in the factor IX promoter is a critical event in transactivation.

Factor IX is an essential vitamin K-dependent serine protease that participates in the intrinsic pathway of coagulation. The protein is expressed exclusively in the liver. The rare Leyden form of hemophilia B (inherited factor IX deficiency) results from point mutations in three proximal promoter elements that decrease factor IX expression. Recovery of expression occurs following puberty, with factor IX protein levels rising into the normal range. We have previously implicated the PAR domain D-site-binding protein (DBP) as well as an upstream element, site 5, as playing important roles in the phenotypic recovery of hemophilia B Leyden. Here we demonstrate that site 5 binds both the CCAAT/enhancer-binding protein (C/EBPalpha) and the ubiquitous Ets factor GA-binding protein (GABPalpha/beta). Transactivation of the factor IX promoter by the PAR proteins DBP and hepatic leukemia factor (HLF) is dependent on the binding of GABPalpha/beta to site 5, and coexpression of these two factors is required for optimal activation of this promoter. The binding of C/EBPalpha to site 5 also augments the activity of GABPalpha/beta. Analysis of the developmental regulation of site 5-binding proteins in rat liver has shown that C/EBPalpha and the GABPbeta subunit increase markedly in the 2 weeks after birth. These observations establish a functional association between the Ets factor GABPalpha/beta and C/EBPalpha and indicate that the two PAR proteins, DBP and HLF, may play complementary roles in factor IX activation. Given the developmental changes exhibited by these proteins, it is likely that they play a role in regulation of the normal factor IX promoter as well as promoters carrying hemophilia B Leyden mutations.     

Two alpha subunits of the Gq class of G proteins stimulate phosphoinositide phospholipase C-beta 1 activity.

Two G protein alpha subunits were detected in preparations of GTP gamma S-dependent, phosphoinositide-specific phospholipase C-activating proteins from bovine liver membranes. Partial resolution of the two alpha subunits, of molecular mass 42 and 43 kDa, was achieved by Mono Q chromatography. Quantitation of the levels of each alpha subunit and reconstitution assays demonstrated that each possessed stimulatory activity towards the beta 1 isozyme of phospholipase C. Immunoblot analysis showed that the 42 kDa protein was immunologically related to alpha q, whereas the 43 kDa protein was related to alpha 11, another member of the Gq class. The data thus show that two different alpha subunits of the Gq class of G proteins stimulate phospholipase C-beta 1 Activity.