A leucine zipper motif of a tegument protein triggers final envelopment of human cytomegalovirus.
The product of the human cytomegalovirus (HCMV) UL71 gene is conserved throughout the herpesvirus family. During HCMV infection, protein pUL71 is required for efficient virion egress and is involved in the final steps of secondary envelopment leading to infectious viral particles. We found strong indications for oligomerization of pUL71 under native conditions when recombinant pUL71 was negatively stained and analyzed by electron microscopy. Oligomerization of pUL71 during infection was further verified by native and reducing polyacrylamide gel electrophoresis (PAGE). By in silico analyses of the pUL71 sequence, we noticed a basic leucine zipper (bZIP)-like domain, which might serve as an oligomerization domain. We demonstrated the requirement of the bZIP-like domain for pUL71 oligomerization by coimmunoprecipitation and bimolecular fluorescence complementation using a panel of pUL71 mutants. These studies revealed that the mutation of two leucine residues is sufficient to abrogate oligomerization but that intracellular localization of pUL71 was unaffected. To investigate the relevance of the bZIP domain in the viral context, recombinant viruses carrying mutations identical to those in the panel of pUL71 mutants were generated. bZIP-defective viral mutants showed impaired viral growth, a small-plaque phenotype, and an ultrastructural phenotype similar to that of the previously described UL71 stop mutant virus. The majority of virus particles within the viral assembly compartment exhibited various stages of incomplete envelopment, which is consistent with the growth defect for the bZIP mutants. From these data we conclude that the bZIP-like domain is required for oligomerization of pUL71, which seems to be essential for correct envelopment of HCMV.
Institute of Medical Virology, Charité Universitätsmedizin Berlin, Berlin, Germany.
Christina Sylvia
CS
Meissner
Sascha
S
Suffner
Martin
M
Schauflinger
Jens
J
von Einem
Elke
E
Bogner
D000595:Amino Acid Sequence; D002460:Cell Line; D003587:Cytomegalovirus / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D003586:Cytomegalovirus Infections / Q000821:virology*; D006801:Humans; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D015678:Viral Structural Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019065:Virus Assembly
D015678:Viral Structural Proteins