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Human cytomegalovirus (HCMV) open reading frame UL32 codes for the basic phosphoprotein pp150 (ppUL32), an abundant constituent of the virion tegument. In order to study its potential role in the assembly and/or transport of progeny particles, astrocytoma cell lines (U373MG) were generated, stably expressing a 2.1 kb 5' fragment of UL32 in antisense orientation under the control of the HCMV major immediate early promoter. The steady-state level of the UL32 sense mRNA and pp150 synthesis were strongly reduced in infected antisense cell lines. Neither immediate early and early gene expression, nor viral DNA replication, was inhibited; the expression of the late gene product gB (gpUL55) was also reduced, but mainly at the level of translation. Control experiments indicated that this differential effect of UL32 antisense expression on the synthesis of viral products was specific. As a consequence of the inhibitory effect, virus yield was significantly reduced in antisense mRNA cell lines. Ultrastructural comparison of control and antisense cells revealed no difference in nucleocapsid forms in the nucleus. However, in the cytoplasm of antisense cells, DNA-containing C capsids and virions were absent and abnormal forms of non-infectious enveloped particles were observed. The data suggest the involvement of pp150 either in the transport of DNA-containing particles through the nuclear envelope or in the stabilization of capsids in the cytoplasm. Thus, UL32 antisense mRNA appears to interfere strongly with virus maturation during the late phase of the infectious cycle.

Human cytomegalovirus late-phase maturation is blocked by stably expressed UL32 antisense mRNA in astrocytoma cells.