GEOapplication/xmlftp://ftp.ncbi.nlm.nih.gov/geo/series/GSE325nnn/GSE325506/primaryOK200OtherHomo sapiensOtherhttps://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE325506GEOGSEfalseHSV-1 temporally orchestrates RAP80 ubiquitination to balance viral replication and evasion [Hi-C]The mechanisms governing the complete replication cycle of herpes simplex virus type 1 (HSV-1) remain incompletely elucidated. Here, we showed that HSV-1 employs a biphasic strategy to dynamically regulate the host protein RAP80 through stage-specific ubiquitination, thereby coordinating its replication cycle. Upon entry, RAP80 undergoes phase separation to form antiviral condensates that encapsulate the viral genome and suppress viral transcription. The HSV-1-encoded E3 ubiquitin ligase ICP0 counteracts this antiviral mechanism by mediating RAP80 ubiquitination to dissolve these condensates, facilitating viral replication. In middle stage, RAP80 depletion leads to accumulation of R-loops and activation of the cGAS–STING–apoptosis axis, threatening viral persistence. In response, in middle and late stage, the viral deubiquitinase UL36USP deubiquitinates and stabilizes RAP80 to suppress R-loop accumulation and this immune-triggered apoptosis. This biphasic regulation of a single host factor represents a sophisticated viral strategy to ensure productive replication and dissemination. Notably, pharmacological disruption of RAP80 ubiquitination using engineered inhibitory peptides potently inhibits HSV-1 replication in vitro and in vivo. Furthermore, RAP80 inhibition enhances both chemosensitivity and oncolytic virotherapy in tumors. These findings reveal a ubiquitination-mediated host–pathogen equilibrium with therapeutic potential in antiviral and anticancer contexts.2026/03/20GSE325506GSM9605332GSM9605310GSM9605331GSM9605330GSM9605319GSM9605318GSM9605339GSM9605317GSM9605338GSM9605316GSM9605315GSM9605337GSM9605336GSM9605314GSM9605335GSM9605313GSM9605312GSM9605334GSM9605333GSM9605311GSM9605321GSM9605320GSM9605309GSM9605308GSM9605329GSM9605307GSM9605306GSM9605328GSM9605327GSM9605305GSM9605304GSM9605326GSM9605325GSM9605324GSM9605323GSM960532234284325506Homo sapiens