Uukuniemi virus maturation: accumulation of virus particles and viral antigens in the Golgi complex.

Abstract

We studied the maturation of Uukuniemi virus and the localization of the viral surface glycoproteins and nucleocapsid protein in infected cells by electron microscopy, indirect immunofluorescence, and immunoelectron microscopy with specific antisera prepared in rabbits against the two glycoproteins Gl and G2 and the nucleocapsid protein N. Electron microscopy of thin sections from infected cells showed virus particles maturing at smooth-surfaced membranes close to the nucleus. Localization of the G1/G2 and N proteins by indirect immunofluorescence at different stages after infection showed the antigens to be present throughout the cell interior but concentrated in the juxtanuclear region. The Gi/ G2 antiserum also appeared to stain the nuclear and plasma membranes. Double staining with tetramethylrhodamine isothiocyanate-conjugated wheat germ agglutinin, which preferentially stains the Golgi complex, and fluorescein isothiocyanate-conjugated anti-rabbit immunoglobulin G, which stained the G1/G2 or N proteins, showed that the staining of the juxtanuclear region coincided. Similarly, double staining for thiamine pyrophosphatase, an enzyme activity specific for the Golgi complex, showed the fluorescence and the cytochemical stain to coincide in the juxtanuclear region. Immunoperoxidase electron microscopy of cells permeabilized with saponin revealed that the viral glycoproteins were present in the rough endoplasmic reticulum and the nuclear and Golgi membranes; the latter was heavily stained. With this method, the N protein was localized to the cytoplasm, especially around smooth-surfaced vesicles in the Golgi region. Taken together, the results indicate that Uukuniemi virus and its structural proteins accumulate in the Golgi complex, supporting the idea that this compartment rather than the plasma membrane is the site of virus maturation. This raises the interesting possibility that deficient transport of the glycoproteins to the plasma membrane and hence their accumulation in the Golgi complex determines the site of virus maturation.A common feature of the viruses belonging to the large Bunyaviridae family (3) is that they appear to mature intracellularly by a budding process at the smooth-surfaced vesicles in the Golgi region (2,18,21,37). If this finding is indeed true, the maturation of the bunyaviruses differs from that of other enveloped viruses, which receive their envelopes either by budding at the plasma membrane (e.g., alpha-, myxo-, paramyxo-, and rhabdoviruses [4,12,13]), nuclear membrane (e.g., herpesviruses [5]), or the rough endoplasmic reticulum (e.g., coronaviruses [20]). The factors determining the site of maturation of enveloped viruses have remained largely unknown.

Hantaan Virus: Identification of Virion Proteins

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Semliki Forest Virus-specific Non-structural Protein nsP3 Is a Phosphoprotein

“…BHK-21 cells infected with SFV were fixed at 4 h p.i. with 3 % paraformaldehyde and permeabilized with 0.05% Triton X-100 (Kuismanen et al, 1982). Preimmune sera and immune sera were diluted 1:50.…”

Heterogeneous Distribution of the Unusual Phospholipid Semilysobisphosphatidic Acid through the Golgi Complex

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