Glenn F. Rall

Viruses, including members of the herpes-, entero-, and morbillivirus families, are the most common cause of infectious encephalitis in mammals worldwide. During most instances of acute viral encephalitis, neurons are typically the initial cell type that is infected. However, as replication and spread ensue, other parenchymal cells can become viral targets, especially in chronic infections. Consequently, to ascertain how neurotropic viruses trigger neuropathology, it is crucial to identify which central nervous system (CNS) cell populations are susceptible and permissive throughout the course of infection, and to define how viruses spread between distinct cell types. Using a measles virus (MV) transgenic mouse model that expresses human CD46 (hCD46), the MV vaccine strain receptor, under the control of a neuron-specific enolase promoter (NSE-hCD46⁺ mice), a novel mode of viral spread between neurons and astrocytes was identified. Although hCD46 is required for initial neuronal infection, it is dispensable for heterotypic spread to astrocytes, which instead depends on glutamate transporters and direct neuron-astrocyte contact. Moreover, in the presence of RNase A, astrocyte infection is reduced, suggesting that nonenveloped ribonucleoproteins (RNP) may cross the neuron-astrocyte synaptic cleft. The characterization of this novel mode of intercellular transport offers insights into the unique interaction of neurons and glia and may reveal therapeutic targets to mitigate the life-threatening consequences of measles encephalitis. IMPORTANCE Viruses are the most important cause of infectious encephalitis in mammals worldwide; several thousand people, primarily the very young and the elderly, are impacted annually, and few therapies are reliably successful once neuroinvasion has occurred. To understand how viruses contribute to neuropathology, and to develop tools to prevent or ameliorate such infections, it is crucial to define if and how viruses disseminate among the different cell populations within the highly complex central nervous system. This study defines a noncanonical mode of viral transmission between neurons and astrocytes within the brain.

Wild elephant populations are declining rapidly due to rampant killing for ivory and body parts, range fragmentation, and human-elephant conflict. Wild and captive elephants are further impacted by viruses, including highly pathogenic elephant endotheliotropic herpesviruses. Moreover, while the rich genetic diversity of the ancient elephant lineage is disappearing, elephants, with their low incidence of cancer, have emerged as a surprising resource in human cancer research for understanding the intrinsic cellular response to DNA damage. However, studies on cellular resistance to transformation and herpesvirus reproduction have been severely limited, in part due to the lack of established elephant cell lines to enablein vitroexperiments. This report describes creation of a recombinant plasmid, pAelPyV-1-Tag, derived from a wild isolate of African Elephant Polyomavirus (AelPyV-1), that can be used to create immortalized lines of elephant cells. This isolate was extracted from a trunk nodule biopsy isolated from a wild African elephant,Loxodonta africana, in Botswana. The AelPyV-1 genome contains open-reading frames encoding the canonical large (LTag) and small (STag) tumor antigens. We cloned the entire early region spanning the LTag and overlapping STag genes from this isolate into a high-copy vector to construct a recombinant plasmid, pAelPyV-1-Tag, which effectively transformed primary elephant endothelial cells. We expect that the potential of this reagent to transform elephant primary cells will, at a minimum, facilitate study of elephant-specific herpesviruses.

Influenza A viruses (IAV) are lytic viruses that have recently been found to activate necroptosis in many of the cell types they infect. Necroptotic cell death is potently immunogenic and limits IAV spread by directly eliminating infected cells and by mobilizing both innate and adaptive immune responses. The benefits of necroptosis to the host, however, may sometimes be outweighed by the potentially deleterious hyperinflammatory consequences of activating this death modality in pulmonary and other tissues.

Genomic material from many neurotropic RNA viruses (e.g., measles virus [MV], West Nile virus [WNV], Sindbis virus [SV], rabies virus [RV], and influenza A virus [IAV]) remains detectable in the mouse brain parenchyma long after resolution of the acute infection. The presence of these RNAs in the absence of overt central nervous system (CNS) disease has led to the suggestion that they are viral remnants, with little or no potential to reactivate. Here we show that MV RNA remains detectable in permissive mouse neurons long after challenge with MV and, moreover, that immunosuppression can cause RNA and protein synthesis to rebound, triggering neuropathogenesis months after acute viral control. Robust recrudescence of viral transcription and protein synthesis occurs after experimental depletion of cells of the adaptive immune response and is associated with a loss of T resident memory (T_rm) lymphocytes within the brain. The disease associated with loss of immune control is distinct from that seen during the acute infection: immune cell-depleted, long-term-infected mice display severe gait and motor problems, in contrast to the wasting and lethal disease that occur during acute infection of immunodeficient hosts. These results illuminate the potential consequences of noncytolytic, immune-mediated viral control in the CNS and demonstrate that what were once considered “resolved” RNA viral infections may, in fact, induce diseases later in life that are distinct from those caused by acute infection. IMPORTANCE Viral infections of neurons are often not cytopathic; thus, once-infected neurons survive, and viral RNAs can be detected long after apparent viral control. These RNAs are generally considered viral fossils, unlikely to contribute to central nervous system (CNS) disease. Using a mouse model of measles virus (MV) neuronal infection, we show that MV RNA is maintained in the CNS of infected mice long after acute control and in the absence of overt disease. Viral replication is suppressed by the adaptive immune response; when these immune cells are depleted, viral protein synthesis recurs, inducing a CNS disease that is distinct from that observed during acute infection. The studies presented here provide the basis for understanding how persistent RNA infections in the CNS are controlled by the host immune response, as well as the pathogenic consequences of noncytolytic viral control.

T cell-mediated adaptive immunity requires naïve, unstimulated T cells to transition from a quiescent metabolic state into a highly proliferative state upon T cell receptor engagement. This complex process depends on transcriptional changes mediated by Ca²⁺-dependent NFAT signaling, mTOR-mediated signaling and increased activity of the guanine nucleotide biosynthetic inosine-5′-monophosphate (IMP) dehydrogenase 1 and 2 enzymes (IMPDH1 and IMPDH2, hereafter IMPDH). Inhibitors of these pathways serve as potent immunosuppressants. Unexpectedly, we discovered that all three pathways converge to promote the assembly of IMPDH protein into micron-scale macromolecular filamentous structures in response to T cell activation. Assembly is post-transcriptionally controlled by mTOR and the Ca²⁺ influx regulator STIM1. Furthermore, IMPDH assembly and catalytic activity were negatively regulated by guanine nucleotide levels, suggesting a negative feedback loop that limits biosynthesis of guanine nucleotides. Filamentous IMPDH may be more resistant to this inhibition, facilitating accumulation of the higher GTP levels required for T cell proliferation.

T cell-mediated adaptive immunity requires naïve, unstimulated T cells to transition from a quiescent metabolic state into a highly proliferative state upon T cell receptor engagement. This complex process depends on transcriptional changes mediated by Ca2+-dependent NFAT signaling, mTOR-mediated signaling and increased activity of the guanine nucleotide biosynthetic enzyme inosine-5'-monophosphate (IMP) dehydrogenase (IMPDH). Inhibitors of these pathways serve as potent immunosuppressants. Unexpectedly, we discovered that all three pathways converge to promote the assembly of IMPDH protein into micron-scale macromolecular filamentous structures in response to T cell activation. Assembly is post-transcriptionally controlled by mTOR and the Ca2+ influx regulator STIM1. Furthermore, IMPDH assembly and catalytic activity were negatively regulated by guanine nucleotide levels, suggesting a negative feedback loop that limits biosynthesis of guanine nucleotides. Filamentous IMPDH may be more resistant to this inhibition, facilitating accumulation of the higher GTP levels required for T cell proliferation.

Three routes of viral entry into the brain have been identified: direct infection of the cells that comprise the blood–brain barrier and blood–cerebrospinal fluid barrier, infection of cells that are licensed to cross these barriers, and transneuronal migration across synapses from the periphery to the CNS. Some viruses that are cytopathic in renewable cell types can switch to a non-productive, non-syncytia-forming mode of spread when infecting neurons, promoting neuron survival. Long-lasting viral infections within the brain may be classified based on the state of the viral genome, its ability to produce infectious progeny and whether such progeny can infect other hosts. Neurotropic viral infections pose unique challenges for the host, including the need to detect antigens within the CNS, the requirement for T lymphocytes to engage with neurons that express negligible levels of the proteins that are typically present on target cells, and the need to mitigate the risk of neuroinflammation and widespread loss of generally non-renewable neurons. The identification of lymphatic drainage portals from the CNS into deep cervical lymph nodes and the presence of a fluid gradient that flushes the brain of extracellular proteins have helped to define how antigens leave the brain to educate the host response in local lymph nodes. The host response to a viral infection may be tailored to promote survival of infected neurons but to destroy similarly infected epithelial or endothelial cells. Non-lytic clearance of neuronal infections may allow for persistence of RNA viruses that induce pathogenesis long after primary exposure.