CONCERTED ACTIVITY OF HOST-CELL FACTOR SUBREGIONS IN PROMOTING STABLEVP16 COMPLEX ASSEMBLY AND PREVENTING INTERFERENCE BY THE ACIDIC ACTIVATION DOMAIN

In contrast to our understanding af the roles of Oct-1 and VP16 in VP1 6-mediated transcriptional activation, virtually nothing is known of t he role of tile second cellular component, termed host cell factor (HC F), or of its structure-function relationships. We show that the major ity of the internal region of HCF, including the repeats involved in H CF cleavage, is dispensable for complex assembly with VP16 and Oct-1, The N-terminal domain of HCF (HCF.N) had only weak VP16 binding and co mplex promoting activity, while the C-terminal region (HCF.C) had no i ntrinsic activity. However, the C-terminal region strongly enhanced co mplex formation and reduced dissociation kinetics when linked to the N -terminal domain (HCF.NC). The potent activity of the HCF.NC fusion in complex assembly was recapitulated in vivo in yeast and mammalian cel ls, Moreover, HCF.N could promote increased complex formation when the acidic activation domain of VP16 was deleted. Restoration of the acti vation domain strongly inhibited complex formation with HCF.N, hut the addition of the C-terminal domain of HCF restored strong stable compl ex formation with intact VP16, The results indicate that this C-termin al domain is critically required to alter the presentation of the acid ic domain of VP16, Additional results are consistent with the interpre tation that this alteration in acidic domain presentation for complex assembly also facilitates the activation function in VP16, The sequenc e of an HCF homolog from Caenorhabditis elegans shows it to be a natur al HCF.NC construct, reinforcing the conclusions from our functional a nalysis.