Palmitoylation of the human cytomegalovirus tegument protein pp28 facilitates virus release

Abstract

Tegument proteins of human cytomegalovirus (HCMV) play essential roles in viral assembly, coordinating interactions among capsids, membranes, and host-derived components. pp28 (UL99), a dominant tegument protein expressed during late infection, is essential for cytoplasmic envelopment and proper trafficking to the viral assembly compartment (vAC). Here, we identify a critical role for palmitoylation in pp28 function. Using site-directed mutagenesis and acyl-resin assisted capture (acyl-RAC) assays, we show that palmitoylation occurs at conserved cysteine residues (Cys6, Cys10, Cys11) near the N-terminus. Disruption of these residues impairs pp28 stability, alters its subcellular localization, and reduces the release of infectious virions without affecting intracellular viral replication. Confocal imaging and proteasome inhibition experiments reveal that palmitoylation-deficient pp28 is more susceptible to degradation and fails to accumulate at ERGIC-derived membranes. Consistent with these findings, recombinant HCMVs encoding pp28 mutants impaired in palmitoylation exhibit reduced extracellular viral titers. These results define palmitoylation as a key modification of pp28 that ensures proper compartmental targeting and virion maturation, underscoring a broader role for tegument lipidation in herpesvirus assembly and egress.