Production of HVJ (Sendai virus) virion is suppressed in the infected LLC-MK2 cells in the low Ca2+ condition. To clarify this phenomenon, the effect of Ca2+ on HVJ morphogenesis was examined under normal and low Ca2+ conditions. The HVJ production was observed in standard medium (containing 1.8 mM Ca2+) within 24 hr after infection and increased with the culture time. Indirect immunofluorescent staining with an anti-HVJ envelope antibody showed that the viral glycoproteins, HANA (HN) and F proteins, were synthesized within 10 hr after infection, diffused to the cell surface, and then many patches strongly stained with this antibody were formed on the cell surface. In contrast, when the infected cells were cultured in low Ca2+ medium, viral glycoproteins were synthesized as in the case of standard medium. However, the glycoproteins were almost all accumulated in the Golgi apparatus, suggesting that expression of viral glycoproteins at the cell surface was suppressed, and the patches were hardly seen on the cell surface. When the low Ca2+ medium was replaced by standard medium, the accumulated viral glycoproteins were rapidly diffused and expressed on the cell surface, HVJ production was restored and patches formed on the cell surface. By confocal laser scanning microscopy, the results were seen more clearly; Ca2+ is necessary for the expression of viral glycoproteins at the cell surface. By electron microscopy under normal conditions, groups of viral budding sites were seen collected in restricted areas, corresponding to the patches observed on immunofluorescent staining, but the budding sites were not seen on the cells cultured in low Ca2+ medium. These results suggest that Ca2+ specifically affects the expression of viral glycoproteins on the cell surface, and that Ca2+ is possibly associated with the assembly of HVJ-specific components necessary for formation of viral budding sites. © 1994, Japan Society for Cell Biology. All rights reserved.