EXPRESSION IN ANIMAL-CELLS AND CHARACTERIZATION OF THE HEPATITIS-E VIRUS STRUCTURAL PROTEINS

Hepatitis E virus (HEV) is a major human pathogen in much of the devel oping world. It is a positive-strand RNA virus with a 7.5 . kb polyade nylated genome consisting of three open reading frames (ORFs), In the absence of an in vitro culture system, the replication and expression strategy of HEV and the nature of its encoded polypeptides are not wel l understood, We have expressed the two ORFs constituting the structur al portion of the HEV genome in COS-1 cells by using simian virus 40-b ased expression vectors and in vitro by using a coupled transcription- translation system. We show here that the major capsid protein, encode d by ORF2, is an 88-kDa glycoprotein which is expressed intracellularl y as well as on the cell surface and has the potential to form noncova lent homodimers, It is synthesized as a precursor (ppORF2) which is pr ocessed through signal sequence cleavage into the mature protein (pORF 2), which is then glycosylated (gpORF2), The minor protein, pORF3, enc oded by ORF3 is a 13.5 . kDa nonglycosylated protein expressed intrace llularly and does not show any major processing. pORF3 interacts with a cellular protein of about 18 kDa which we call 3IP, the pORF3-intera cting protein, The significance of these findings are discussed in lig ht of an existing model of HEV genome replication and expression.