FUNCTIONS OF THE INTERNAL PRE-S DOMAIN OF THE LARGE SURFACE PROTEIN IN HEPATITIS-B VIRUS PARTICLE MORPHOGENESIS

The large hepatitis B virus (HBV) surface protein (L) forms two isomer s which display their N-terminal pre-S domain at the internal and exte rnal side of the viral envelope, respectively, The external pre-S doma in has been implicated in binding to a virus receptor, To investigate functions of the internal pre-S domain, a secretion signal sequence wa s fused to the N terminus of L (sigL), causing exclusive expression of external pre-S domains. A fusion construct with a nonfunctional signa l (s25L), which corresponds in its primary sequence to sigL cleaved by signal peptidase, was used as a control. SigL was N glycosylated in t ransfected COS cells at both potential sites in pre-S in contrast to s 25L or wild-type L, confirming the expected transmembrane topologies o f sigL and s25L. Phenotypic characterization revealed the following po ints. (i) SigL lost the inhibitory effect of L or s25L on secretion of subviral hepatitis B surface antigen particles, suggesting that the r etention signal mapped to the N terminus of L is recognized in the cyt osol and not in the lumen of the endoplasmic reticulum. (ii) SigL was secreted into the culture medium even in the absence of the major HBV surface protein (S), while release of an L mutant lacking the retentio n signal was still dependent on S coexpression. (iii) s25L but not sig L could complement an L negative HBV genome defective for virion secre tion in cotransfections. This suggests that the cytosolic pre-S domain , like a matrix protein, is involved in the interaction of the viral e nvelope with preformed cytosolic nucleocapsids during virion assembly.