REPRESSION OF ENHANCER-II ACTIVITY BY A NEGATIVE REGULATORY ELEMENT IN THE HEPATITIS-B VIRUS GENOME

Enhancer II of human hepatitis B virus has dual functions in vivo. Loc ated at nucleotides (nt) 1646 to 1741, it can stimulate the surface an d X promoters from a downstream position. Moreover, the same sequence can also function as upstream regulatory element that activates the co re promoter in a position- and orientation-dependent manner. In this s tudy, we report the identification and characterization of a negative regulatory element (NRE) upstream of enhancer II (nt 1613 to 1636) whi ch can repress both the enhancer and upstream stimulatory function of the enhancer LT sequence in differentiated liver cells. This NRE has m arginal inhibitory effect by itself but a strong repressive function i n the presence of a functional enhancer II. Mutational analysis reveal s that sequence from nt 1616 to 1621 is required for repression of enh ancer activity by the NRE. Gel shift analysis reveals that this negati ve regulatory region can be recognized by a specific protein factor(s) present at the 0.4 M NaCl fraction of HepG2 nuclear extracts. The dis covery of the NRE indicates that HBV gene transcription is controlled by combined efforts of both positive and negative regulation. It also provides a unique system with which to study the mechanism of negative regulation of gene expression.