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Congenital CMV (cCMV) infection causes neurological sequelae in newborns. Recent studies of cCMV pathogenesis in animal models reveal ventriculomegaly and cortical atrophy associated with impaired neural progenitor cell (NPC) proliferation and migration.

During the long coevolution of human cytomegalovirus (HCMV) and humans, the host has formed a defense system of multiple layers to eradicate the invader, and the virus has developed various strategies to evade host surveillance programs. The intrinsic immunity primarily orchestrated by promyelocytic leukemia (PML) nuclear bodies (PML-NBs) represents the first line of defense against HCMV infection. Here, we demonstrate that microrchidia family CW-type zinc finger 3 (MORC3), a PML-NBs component, is a restriction factor targeting HCMV infection. We show that depletion of MORC3 through knockdown by RNA interference or knockout by CRISPR-Cas9 augmented immediate-early protein 1 (IE1) gene expression and subsequent viral replication, and overexpressing MORC3 inhibited HCMV replication by suppressing IE1 gene expression. To relief the restriction, HCMV induces transient reduction of MORC3 protein level via the ubiquitin-proteasome pathway during the immediate-early to early stage. However, MORC3 transcription is upregulated, and the protein level recovers in the late stages. Further analyses with temporal-controlled MORC3 expression and the major immediate-early promoter (MIEP)-based reporters show that MORC3 suppresses MIEP activity and consequent IE1 expression with the assistance of PML. Taken together, our data reveal that HCMV enforces temporary loss of MORC3 to evade its repression against the initiation of immediate-early gene expression.

Human cytomegalovirus (HCMV) has a large (∼235 kb) genome with more than 200 predicted open reading frames that exploits numerous cellular factors to facilitate its replication. A key feature of HCMV-infected cells is the emergence of a distinctive membranous cytoplasmic compartment termed the virion assembly compartment (vAC). Here, we report that host protein WD repeat domain 11 (WDR11) plays a key role in vAC formation and virion morphogenesis. We found that WDR11 was upregulated at both mRNA and protein levels during HCMV infection. At the late stage of HCMV replication, WDR11 relocated to the vAC and colocalized with markers of the trans-Golgi network (TGN) and vAC. Depletion of WDR11 hindered HCMV-induced membrane reorganization of the Golgi and TGN, altered vAC formation, and impaired HCMV secondary envelopment and virion morphogenesis. Further, motifs critical for the localization of WDR11 in TGN were identified by alanine-scanning mutagenesis. Mutation of these motifs led to WDR11 mislocation outside the TGN and loss of vAC formation. Taken together, these data indicate that host protein WDR11 is required for efficient viral replication at the stage of virion assembly, possibly by facilitating the remodeling of the endomembrane system for vAC formation and virion morphogenesis. IMPORTANCE During the late phase of human cytomegalovirus (HCMV) infection, the endomembrane system is dramatically reorganized, resulting in the formation of a unique structure termed the virion assembly compartment (vAC), which is critical for the assembly of infectious virions. The mechanism of HCMV-induced vAC formation is still not fully understood. In this report, we identified a host factor, WDR11, that plays an important role in vAC formation. Our findings argue that WDR11 contributes to the relocation of the Golgi and trans-Golgi network to the vAC, a membrane reorganization process that appears to be required for efficient virion maturation. The present work provides new insights into the vAC formation and HCMV virion morphogenesis and a potential novel target for antiviral treatment.

We previously reported that human cytomegalovirus (HCMV) utilizes the cellular protein WD repeat-containing protein 5 (WDR5) to facilitate capsid nuclear egress. Here, we further show that HCMV infection results in WDR5 localization in a juxtanuclear region and that its localization to this cellular site is associated with viral replication and late viral gene expression. Furthermore, WDR5 accumulated in the virion assembly compartment (vAC) and colocalized with vAC markers of γ-tubulin, early endosomes, and viral vAC marker proteins pp65, pp28, and glycoprotein B (gB). WDR5 coimmunoprecipitated with multiple virion proteins, including major capsid protein (MCP), pp150, pp65, pIRS1, and pTRS1, which may explain WDR5 accumulation in the vAC during infection. WDR5 fractionated with virions in either the presence or absence of Triton X-100 and was present in purified viral particles, suggesting that WDR5 was incorporated into HCMV virions. Thus, WDR5 localized to the vAC and was incorporated into virions, raising the possibility that in addition to capsid nuclear egress, WDR5 could also participate in cytoplasmic HCMV virion morphogenesis. IMPORTANCE Human cytomegalovirus (HCMV) has a large (∼235-kb) genome that contains over 170 open reading frames (ORFs) and exploits numerous cellular factors to facilitate its replication. In the late phase of HCMV infection, cytoplasmic membranes are reorganized to establish the virion assembly compartment (vAC), which has been shown to be necessary for efficient assembly of progeny virions. We previously reported that WDR5 facilitates HCMV nuclear egress. Here, we show that WDR5 is localized to the vAC and incorporated into virions, perhaps contributing to efficient virion maturation. Thus, findings in this study identified a potential role for WDR5 in HCMV assembly in the cytoplasmic phase of virion morphogenesis.

WD repeat-containing protein 5 (WDR5) is essential for assembling the VISA-associated complex to induce a type I interferon antiviral response to Sendai virus infection. However, the roles of WDR5 in DNA virus infections are not well described. Here, we report that human cytomegalovirus exploits WDR5 to facilitate capsid nuclear egress. Overexpression of WDR5 in fibroblasts slightly enhanced the infectious virus yield. However, WDR5 knockdown dramatically reduced infectious virus titers with only a small decrease in viral genome replication or gene expression. Further investigation of late steps of viral replication found that WDR5 knockdown significantly impaired formation of the viral nuclear egress complex and induced substantially fewer infoldings of the inner nuclear membrane. In addition, fewer capsids were associated with these infoldings, and there were fewer capsids in the cytoplasm. Restoration of WDR5 partially reversed these effects. These results suggest that WDR5 knockdown impairs the nuclear egress of capsids, which in turn decreases virus titers. These findings reveal an important role for a host factor whose function(s) is usurped by a viral pathogen to promote efficient replication. Thus, WDR5 represents an interesting regulatory mechanism and a potential antiviral target. IMPORTANCE Human cytomegalovirus (HCMV) has a large (∼235-kb) genome with over 170 open reading frames and exploits numerous cellular factors to facilitate its replication. HCMV infection increases protein levels of WD repeat-containing protein 5 (WDR5) during infection, overexpression of WDR5 enhances viral replication, and knockdown of WDR5 dramatically attenuates viral replication. Our results indicate that WDR5 promotes the nuclear egress of viral capsids, the depletion of WDR5 resulting in a significant decrease in production of infectious virions. This is the first report that WDR5 favors HCMV, a DNA virus, replication and highlights a novel target for antiviral therapy.

The mechanisms underlying neurodevelopmental damage caused by virus infections remain poorly defined. Congenital human cytomegalovirus (HCMV) infection is the leading cause of fetal brain development disorders. Previous work has linked HCMV to perturbations of neural cell fate, including premature differentiation of neural progenitor cells (NPCs). Here we show that HCMV infection of NPCs results in the loss of the SOX2 protein, a key pluripotency-associated transcription factor. SOX2 depletion maps to the HCMV major immediate-early (IE) transcription unit and is individually mediated by the IE1 and IE2 proteins. IE1 causes SOX2 down-regulation by promoting the nuclear accumulation and inhibiting the phosphorylation of STAT3, a transcriptional activator of SOX2 expression. Deranged signaling resulting in depletion of a critical stem cell protein is an unanticipated mechanism by which the viral major IE proteins may contribute to brain development disorders caused by congenital HCMV infection.IMPORTANCEHuman cytomegalovirus (HCMV) infections are a leading cause of brain damage, hearing loss and other neurological disabilities in children. We report that the HCMV proteins known as IE1 and IE2 target expression of human SOX2, a central pluripotency-associated transcription factor that governs neural progenitor cell (NPC) fate and is required for normal brain development. Both during HCMV infection and when expressed alone, IE1 causes the loss of SOX2 from NPCs. IE1 mediates SOX2 depletion by targeting STAT3, a critical upstream regulator of SOX2 expression. Our findings reveal an unanticipated mechanism by which a common virus may cause damage to the developing nervous system and suggest novel targets for medical intervention.

The mechanisms underlying neurodevelopmental damage caused by virus infections remain poorly defined. Congenital human cytomegalovirus (HCMV) infection is the leading cause of fetal brain development disorders. Previous work has linked HCMV infection to perturbations of neural cell fate, including premature differentiation of neural progenitor cells (NPCs). Here, we show that HCMV infection of NPCs results in loss of the SOX2 protein, a key pluripotency-associated transcription factor. SOX2 depletion maps to the HCMV major immediate early (IE) transcription unit and is individually mediated by the IE1 and IE2 proteins. IE1 causes SOX2 downregulation by promoting the nuclear accumulation and inhibiting the phosphorylation of STAT3, a transcriptional activator of SOX2 expression. Deranged signaling resulting in depletion of a critical stem cell protein is an unanticipated mechanism by which the viral major IE proteins may contribute to brain development disorders caused by congenital HCMV infection.IMPORTANCE Human cytomegalovirus (HCMV) infections are a leading cause of brain damage, hearing loss, and other neurological disabilities in children. We report that the HCMV proteins known as IE1 and IE2 target expression of human SOX2, a central pluripotency-associated transcription factor that governs neural progenitor cell (NPC) fate and is required for normal brain development. Both during HCMV infection and when expressed alone, IE1 causes the loss of SOX2 from NPCs. IE1 mediates SOX2 depletion by targeting STAT3, a critical upstream regulator of SOX2 expression. Our findings reveal an unanticipated mechanism by which a common virus may cause damage to the developing nervous system and suggest novel targets for medical intervention.

Electronic transport through a double quantum dot interferometer with Rashba spin–orbit interaction is investigated. By means of the slave-boson mean-field approximation and Green function technique, we calculated the local density of states (DOS) and the linear conductance in the Kondo regime at zero temperature. It is shown that the local DOS and the linear conductance of each spin component can be tuned by the Rashba spin–orbit interaction with help of the magnetic flux. In particular, by modulating the strength of the Rashba spin–orbit interaction properly, only one spin component electron can be allowed to transport through this structure.

Combine olfactory ensheathing glia (OEG) implantation with ex vivo non-viral vector-based neurotrophin-3 (NT-3) gene therapy in attempting to enhance regeneration after thoracic spinal cord injury (SCI). Primary OEG were transfected with cationic liposome-mediated recombinant plasmid pcDNA3.1(+)-NT3 and subsequently implanted into adult Wistar rats directly after the thoracic spinal cord (T9) contusion by the New York University impactor. The animals in 3 different groups received 4×105 OEG transfected with pcDNA3.1(+)-NT3 or pcDNA3.1(+) plasmids, or the OEGs without any plasmid transfection, respectively; the fourth group was untreated group, in which no OEG was implanted. NT-3 production was seen increased both ex vivo and in vivo in pcDNA3.1(+)-NT3 transfected OEGs. Three months after implantation of NT-3-transfected OEGs, behavioral analysis revealed that the hindlimb function of SCI rats was improved. All spinal cords were filled with regenerated neurofilament-positive axons. Retrograde tracing revealed enhanced regenerative axonal sprouting. Non-viral vector-mediated genetic engineering of OEG was safe and more effective in producing NT-3 and promoting axonal outgrowth followed by enhancing SCI recovery in rats.