NUCLEAR-LOCALIZATION OF THE HUMAN CYTOMEGALOVIRUS TEGUMENT PROTEIN PP150 (PPUL32)

The human cytomegalovirus (HCMV) basic phosphoprotein pp150, encoded b y the UL32 gene, together with the two other major phosphoproteins, pp 65 (ppUL83) and pp71 (ppUL82) and several minor structural proteins, f orm the tegument around the viral nucleocapsid. Experiments were under taken to locate the area of assembly of tegument proteins pp150 and pp 65 and nucleocapsids in fibroblasts, in order to assess the functional role of these two structural proteins in HCMV morphogenesis. Whereas pp150 expression starts during the cytoplasmic maturation of HCMV, pp6 5 is expressed in the early and late phases of HCMV gene transcription . Western blot analysis of isolated cell fractions showed that pp150 i s initially (48 h postinfection) localized in the nucleus, associated either with the nuclear membrane or with viral assembly regions, and l ater (72 h post-infection) in the cytoplasm. By indirect immunofluores cence, pp150 and pp65 could be detected in nuclear subcompartments and were strongly associated with the nuclear membrane. Using immunogold analysis by electron microscopy, pp65 was exclusively detected within the matrix of cytoplasmic and extracellular dense bodies and of dense body-like structures in the nucleoplasm. These were localized in close contact with hypertrophic nucleoli, in the proximity of developing nu cleocapsids and in special patches at the inner nuclear membrane. Posi tive immunostaining of pp150 was observed at the surface of developing nucleocapsids concentrated within viral assembly regions in the nucle oplasm. Additionally, the tegument of cytoplasmic and extracellular vi rions was stained, whereas dense bodies or nuclear dense body-like str uctures did not react. Thus, the acquisition of the tegument protein p p150 seems to start in special nuclear subcompartments of the HCMV-inf ected fibroblasts.