Biology subjects

Koyuncu, O. O.

Publications and source records attributed to Koyuncu, O. O..

6 recordsLinked to original sources

bioRxiv2026

Axonal entry and retrograde transport define HSV-1 latency establishment and reactivation potential in neurons

Herpes simplex virus 1 (HSV-1) establishes life-long latency in peripheral neurons following neuroinvasion, yet the early determinants governing latency versus productive infection remain poorly understood. Available in vitro models bypass the physiological route of infection by directly infecting neuronal cell bodies under antiviral suppression. Here, we established a compartmented axonal infection model that recapitulates HSV-1 neuroinvasion and latency establishment without pharmacological inhibitors. Using primary superior cervical ganglion (SCG) neurons, we demonstrated that low-dose axonal infection consistently establishes latency, whereas higher axonal doses or direct somatic infection led to productive replication. Latently infected neurons exhibit accumulation of the latency-associated transcript (LAT) and can be efficiently reactivated by UV-inactivated virus or VP16 expression. Remarkably, productive infection can be induced during low-dose axonal infection by simultaneous exposure of neuronal cell bodies, but not axons, to replication-deficient virions, implicating tegument delivery in soma overrides latency establishment. Conversely, excess replication-incompetent particles in axons suppress productive outcomes and subsequent reactivations, suggesting competition for retrograde transport. Together, these findings identify axonal entry and transport as critical regulatory checkpoints in HSV-1 latency establishment and suggest that interference with retrograde trafficking may represent a strategy to limit neuroinvasion and life-long reactivations.

microbiology↗

bioRxiv2026

Differential Sensitivity of HSV-1 and PRV to IFN-{lambda} Reveals a Neuron-Specific Antiviral Role for RSAD2

Alpha herpesviruses (-HV) initially infect mucosal epithelial cells and subsequently establish lifelong latency in the peripheral nervous system (PNS). Herpes simplex virus-1 (HSV-1), a human pathogen persisting in the majority of the adult population, shares neuroinvasive properties with Pseudorabies virus (PRV), a swine -HV, commonly used as a model -HV. Utilizing primary peripheral neuronal cultures, we previously showed that IFN-{lambda} pre-treatment significantly reduced PRV yield. In this paper, we further characterized the early and late neuronal responses to IFN-{lambda} by RNA-seq, and the antiviral potential of this response against HSV-1. Notably, HSV-1 exhibited neuron-specific resistance to IFN-{lambda} mediated antiviral responses both in murine primary neurons and human neuronal cells. An ICP34.5-deficient HSV-1 ({Delta}34.5) mutant showed IFN-{lambda} sensitivity in neurons, while replicating normally in untreated neurons showing that ICP34.5 is responsible for the neuron specific IFN-{lambda} resistance of HSV-1. Our results further demonstrate that RSAD2 is strongly induced by IFN-{lambda} in neurons, localizing to ER-associated membranes, and effectively restricting -HV protein synthesis in the absence of ICP34.5. siRNA-mediated RSAD2 knockdown in IFN-{lambda}-primed primary neurons largely restored replication of {Delta}34.5 HSV-1, highlighting the role of this IFN-{lambda} induced host factor in neuronal infections. Together, neuronal IFN-{lambda}-induced RSAD2 and HSV-1 ICP34.5 define a neuron-specific antagonistic mechanism that collectively determines the replication efficiency of HSV-1 in the PNS.

microbiology↗

bioRxiv2023

Pseudorabies virus hijacks DDX3X, initiating an addictive 'mad itch' and immune suppression, to facilitate viral spread.

Infections with defined Herpesviruses, such as Pseudorabies virus (PRV) and Varicella zoster virus (VZV) can cause neuropathic itch, referred to as "mad itch" in multiple species. The underlying mechanisms involved in neuropathic "mad itch" are poorly understood. Here, we show that PRV infections hijack the RNA helicase DDX3X in sensory neurons to facilitate anterograde transport of the virus along axons. PRV induces re-localization of DDX3X from the cell body to the axons which ultimately leads to death of the infected sensory neurons. Inducible genetic ablation of Ddx3x in sensory neurons results in neuronal death and "mad itch" in mice. This neuropathic "mad itch" is propagated through activation of the opioid system making the animals "addicted to itch". Moreover, we show that PRV co-opts and diverts T cell development in the thymus via a sensory neuron-IL-6-hypothalamus-corticosterone stress pathway. Our data reveal how PRV, through regulation of DDX3X in sensory neurons, travels along axons and triggers neuropathic itch and immune deviations to initiate pathophysiological programs which facilitate its spread to enhance infectivity.

neuroscience↗

bioRxiv2023

Neuronal expression of herpes simplex virus type-1 VP16 protein induces pseudorabies virus escape from silencing and reactivation by activating Jun

Alpha herpesvirus (-HV) particles enter their hosts from mucosal surfaces and efficiently maintain fast transport in peripheral nervous system (PNS) axons to establish infections in the peripheral ganglia. The path from axons to distant neuronal nuclei is challenging to dissect due to the difficulty of monitoring early events in a dispersed neuron culture model. We have established well-controlled, reproducible, and reactivateable latent infections in compartmented rodent neurons by infecting physically isolated axons with a small number of viral particles. This system not only recapitulates the physiological infection route, but also facilitates independent treatment of isolated cell bodies or axons. Consequently, this system enables study not only of the stimuli that promote reactivation, but also the factors that regulate the initial switch from productive to latent infection. Adeno associated virus (AAV) mediated expression of herpes simplex type 1 (HSV-1) VP16 alone in neuronal cell bodies enabled the escape from silencing of incoming pseudorabies virus (PRV) genomes. Furthermore, expression of HSV VP16 alone reactivated a latent PRV infection in this system. Surprisingly, expression of PRV VP16 protein supported neither PRV escape from silencing nor reactivation. We compared transcription transactivation activity of both VP16 proteins in primary neurons by RNA sequencing and found that these homolog viral proteins produce different gene expression profiles. AAV transduced HSV VP16 specifically induced expression of proto-oncogenes including Jun and Pim2. In addition, HSV VP16 induces phosphorylation of Jun in neurons, and when this activity is inhibited, escape of PRV silencing is dramatically reduced.

microbiology↗

bioRxiv2021

A pseudorabies virus serine/threonine kinase, US3, promotes retrograde transport in axons via Akt/mToRC1

Infection of peripheral axons by alpha herpesviruses (AHVs) is a critical stage in establishing a life-long infection in the host. Upon entering the cytoplasm of axons, AHV nucleocapsids and associated inner-tegument proteins must engage the cellular retrograde transport machinery to promote the long-distance movement of virion components to the nucleus. The current model outlining this process is incomplete and further investigation is required to discover all viral and cellular determinants involved as well as the temporality of the events. Using a modified tri-chamber system, we have discovered a novel role of the pseudorabies virus (PRV) serine/threonine kinase, US3, in promoting efficient retrograde transport of nucleocapsids. We discovered that transporting nucleocapsids move at similar velocities both in the presence and absence of a functional US3 kinase; however fewer nucleocapsids are moving when US3 is absent and move for shorter periods of time before stopping, suggesting US3 is required for efficient nucleocapsid engagement with the retrograde transport machinery. This led to fewer nucleocapsids reaching the cell bodies to produce a productive infection 12hr later. Furthermore, US3 was responsible for the induction of local translation in axons as early as 1hpi through the stimulation of a PI3K/Akt-mToRC1. These data describe a novel role for US3 in the induction of local translation in axons during AHV infection, a critical step in transport of nucleocapsids to the cell body. ImportanceNeurons are highly polarized cells with axons that can reach centimeters in length. Communication between axons at the periphery and the distant cell body is a relatively slow process involving the active transport of chemical messengers. Theres a need for axons to respond rapidly to extracellular stimuli. Translation of repressed mRNAs present within the axon occurs to enable rapid, localized responses independently of the cell body. AHVs have evolved a way to hijack local translation in the axons to promote their transport to the nucleus. We have determined the cellular mechanism and viral components involved in the induction of axonal translation. The US3 serine/threonine kinase of PRV activates Akt-mToRC1 signaling pathways early during infection to promote axonal translation. When US3 is not present, the number of moving nucleocapsids and their processivity are reduced, suggesting that US3 activity is required for efficient engagement of nucleocapsids with the retrograde transport machinery.

molecular biology↗

bioRxiv2019

Pseudorabies Virus Infection Accelerates Degradation of the Kinesin-3 Motor KIF1A

Alphaherpesviruses, including pseudorabies virus (PRV), are neuroinvasive pathogens that establish life-long latency in peripheral ganglia following the initial infection at mucosal surfaces. The establishment of latent infection and the subsequent reactivations during which newly-assembled virions are sorted into and transported anterogradely inside axons to the initial mucosal site of infection, rely on axonal bidirectional transport mediated by microtubule-based motors. Previous studies using cultured peripheral nervous system (PNS) neurons have demonstrated that KIF1A, a kinesin-3 motor, mediates the efficient axonal sorting and transport of newly-assembled PRV virions. In this study, we report that KIF1A, unlike other axonal kinesins, is an intrinsically unstable protein prone to proteasomal degradation. Interestingly, PRV infection of neuronal cells leads not only to a non-specific depletion of KIF1A mRNA, but also to an accelerated proteasomal degradation of KIF1A proteins, leading to a near depletion of KIF1A protein late in infection. Using a series of PRV mutants deficient in axonal sorting and anterograde spread, we identified the PRV US9/gE/gI protein complex as a viral factor facilitating the proteasomal degradation of KIF1A proteins. Moreover, by using compartmented neuronal cultures that fluidically and physically separate axons from cell bodies, we found that the proteasomal degradation of KIF1A occurs in axons during infection. We propose that PRV anterograde sorting complex, gE/gI/US9, recruits KIF1A to viral transport vesicles for axonal sorting and transport, and eventually accelerates the proteasomal degradation of KIF1A in axons. ImportancePseudorabies virus (PRV) is an alphaherpesvirus related to human pathogens herpes simplex virus -1, -2 and varicella zoster virus. Alphaherpesviruses are neuroinvasive pathogens that establish life-long latent infections in the host peripheral nervous system (PNS). Following reactivation from latency, infection spreads from the PNS back via axons to the peripheral mucosal tissues, a process mediated by kinesin motors. Here, we unveil and characterize the underlying mechanisms for a PRV-induced, accelerated degradation of KIF1A, a kinesin-3 motor promoting the sorting and transport of PRV virions in axons. We show that PRV infection disrupts the synthesis of KIF1A, and simultaneously promotes the degradation of intrinsically unstable KIF1A proteins by proteasomes in axons. Our work implies that the timing of motor reduction after reactivation would be critical because progeny particles would have a limited time window for sorting into and transport in axons for further host-to-host spread.

microbiology↗