Virus-against-virus dominant-negative interference strategy targeting a viral CC chemokine prevents cytomegalovirus-related neurodevelopmental pathogenesis
Authors: Bauer Sylvian , Tarhini Sarah , Buhler Emmanuelle , Saha Saswati , Stamminger Thomas , Streblow Daniel N. , Burnashev Nail , Luche Hervé , Szepetowski Pierre 
Published: February 2025

Background

Congenital cytomegalovirus (CMV) infections are one leading cause of human neurodevelopmental disorders. Increasing evidence for the pathogenic involvement of brain immune alterations was obtained in the recent years. Host and virus-encoded chemokines might play important roles in CMV-related neuropathogenesis by regulating leukocyte trafficking and microglia recruitment in the CMV-infected brains, and by interfering with key neurodevelopmental steps. In a rat model of CMV infection of the fetal brainin uterothat leads to detrimental neurologic and other severe phenotypes postnatally, we reported on the early alteration of microglia and on the infiltration of the infected brains by lymphoid and myeloid cells. Particularly, expression of ther129gene encoding the viral chemokine RCK3 was detected as early as 24h post-infection; together with the previously reported chemotaxis properties exerted by RCK3 on lymphocytes and on macrophagesin vitro, this suggested that RCK3 might be involved in the brain immune cell alterations seen in the CMV-infected developing brains, and in the related neuropathogenesis.

Methods

Infection of the rat fetal brain was done by intracerebroventricular injectionsin uteroof either rat CMV encoding wild-type (wt) RCK3 (RCMV-wt), or a mutant CMV counterpart encoding RCK3 with a deletion in its chemokine domain (RCMV-r129ΔNT). As RCMV-r129ΔNT had shown dominant-negative effects on the chemotaxis properties of RCK3in vitro, simultaneous and successive co-infection rescue assays were also performed. The detrimental postnatal phenotypesin vivoand the epileptiform activityex vivousually detected after infection of the rat developing brain with RCMV-wt were monitored in the RCMV-r129ΔNT condition and in co-infection assays.

Results

In sharp contrast with RCMV-wt whose infection of the fetal brain led to decreased postnatal survival, impaired sensorimotor development, hindlimb hyperextension and epileptic seizures in neonatal pups, RCMV-r129ΔNT infection was not associated with any severe postnatal phenotypein vivo. Consistently, the epileptiform activity recorded in most neocortical slices from RCMV-wt-infected pups was not detected in any slice from RCMV-r129ΔNT-infected pups. Simultaneous co-infection assays led to dramatic prevention against the postnatal phenotypesin vivoand the altered network activityex vivo, revealing a dose-dependent rescuing effect exerted by RCMV-r129ΔNT on RCMV-wt. Importantly, successful rescue was also obtained when the mutant RCMV-r129ΔNT was inoculated in the fetal brains either before or after infection with RCMV-wt.

Significance

Our data demonstrate the crucial neuropathogenic role of RCK3 CMV chemokinein vivo. The dramatic success and apparent safety of the dominant-negative RCK3 rescue assaysin vivoprovide a proof-of-principle for the beneficial use of a virus-against-virus approach against CMV-related pathogenesis.