Characterization of essential domains for the functionality of the MHBst transcriptional activator and identification of a minimal MHBst activator.
Integrated hepatitis B virus DNA derived from hepatocellular carcinomas can express, in one third of the cases investigated so far, a transcriptional activator encoded from 3' terminal truncated surface (preS/S) genes resulting in a C-terminally truncated middle surface protein (MHBst). Since MHBst, in contrast to the secreted MHBs, is retained in the secretory pathway at the ER, the question as to whether the retention generates the transcriptional activator function was investigated. Through fusion of MHBs to the ER-retention signal KDEL, it was shown that the intracellular retention does not generate the transcriptional activator function. Tryptic digestions of microsomal vesicles revealed that the amino terminal domain of MHBst directs into the cytoplasmic compartment, whereas in MHBs this domain directs into the lumen of the ER. This structural difference appears to be why transcriptional activator function arises. Through deletion analysis it was shown that non-membrane-associated MHBst proteins are also functional activators. Nonmembrane associated MHBst proteins represent a second class of MHBst proteins. These MHBst-proteins are homogenously distributed all over the cell and show no difference in functionality as compared to the membrane-associated MHBst proteins. MHBst53 (truncated at aa53) was shown to be a minimal activator of this class. Both classes of MHBst proteins were found to form dimers; an which is involved in mediating the dimerization. The integrity of this domain was also revealed to be a prerequisite for the functionality of the activator, suggesting a linkage between dimerization and functionality.
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000821:virology; D002462:Cell Membrane / Q000378:metabolism; D006367:HeLa Cells; D006515:Hepatitis B virus / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism; D008113:Liver Neoplasms / Q000821:virology; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis; D017433:Protein Structure, Secondary; D018411:Spodoptera / Q000821:virology; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014162:Transfection
D008562:Membrane Glycoproteins; D010446:Peptide Fragments; D015534:Trans-Activators