Accumulation of virion tegument and envelope proteins in a stable cytoplasmic compartment during human cytomegalovirus replication: Characterization of a potential site of virus assembly

The assembly of human cytomegalovirus (HCMV) is thought to be similar to th at which has been proposed for alphaherpesviruses and involve envelopment o f tegumented subviral particles at the nuclear membrane followed by export from the cell by a poorly defined pathway. However, several studies have sh own that at least two tegument virion proteins remain in the cytoplasm duri ng the HCMV replicative cycle, thereby suggesting that HCMV cannot acquire its final envelope at the nuclear envelope, We investigated the assembly of HCMV by determining the intracellular trafficking of the abundant tegument protein pp150 (UL32) in productively infected human fibroblasts. Our resul ts indicated that pp150 remained within the cytoplasm throughout the replic ative cycle of HCMV and accumulated in a stable, juxtanuclear structure lat e in infection. Image analysis using a variety of cell protein-specific ant ibodies indicated that the pp150-containing structure was not a component o f the endoplasmic reticulum, (ER), ER-Golgi intermediate compartment, cis o r medial Golgi, or lysosomes, Partial colocalization of the structure was n oted with the trans-Golgi network, and it appeared to lie in close proximit y to the microtubule organizing center, Two additional tegument proteins (p p28 and pp65) and three envelope glycoproteins (gB, gH, and gp65) localized in this same structure late infection. This compartment appeared to be rel atively stable since pp150, pp65, and the processed form of gB could be coi solated following cell fractionation, Our findings indicated that pp150 was expressed exclusively within the cytoplasm throughout the infectious cycle of HCMV and that the accumulation of the pp150 in this cytoplasmic structu re was accompanied by at least five other virion proteins. These results su ggested the possibility that this virus-induced structure represented a cyt oplasmic site of virus assembly.