Human reproduction is regulated by retrotransposons derived from ancient Hominidae-specific viral infections.

Germ cells are essential to pass DNA from one generation to the next. In human reproduction, germ cell development begins with the specification of primordial germ cells (PGCs) and a failure to specify PGCs leads to human infertility. Recent studies have revealed that the transcription factor network required for PGC specification has diverged in mammals, and this has a significant impact on our understanding of human reproduction. Here, we reveal that the Hominidae-specific Transposable Elements (TEs) LTR5Hs, may serve as TEENhancers (TE Embedded eNhancers) to facilitate PGC specification. LTR5Hs TEENhancers become transcriptionally active during PGC specification both in vivo and in vitro with epigenetic reprogramming leading to increased chromatin accessibility, localized DNA demethylation, enrichment of H3K27ac, and occupation of key hPGC transcription factors. Inactivation of LTR5Hs TEENhancers with KRAB mediated CRISPRi has a significant impact on germ cell specification. In summary, our data reveals the essential role of Hominidae-specific LTR5Hs TEENhancers in human germ cell development.

Avian leukosis virus subgroup J and reticuloendotheliosis virus coinfection induced TRIM62 regulation of the actin cytoskeleton.

BACKGROUND: Coinfection with avian leukosis virus subgroup J (ALV-J) and reticuloendotheliosis virus (REV) is common in chickens, and the molecular mechanism of the synergistic pathogenic effects of the coinfection is not clear. Exosomes have been identified as new players in the pathogenesis of retroviruses. The different functions of exosomes depend on their cargo components. OBJECTIVES: The aim of this study was to investigate the function of co-regulation differentially expressed proteins in exosomes on coinfection of ALV-J and REV. METHODS: Here, viral replication in CEF cells infected with ALV-J, REV or both was detected by immunofluorescence microscopy. Then, we analyzed the exosomes isolated from supernatants of chicken embryo fibroblast (CEF) cells single infected and coinfected with ALV-J and REV by mass spectrometry. KEGG pathway enrichment analyzed the co-regulation differentially expressed proteins in exosomes. Next, we silenced and overexpressed tripartite motif containing 62 (TRIM62) to evaluate the effects of TRIM62 on viral replication and the expression levels of NCK-association proteins 1 (NCKAP1) and actin-related 2/3 complex subunit 5 (ARPC5) determined by quantitative reverse transcription polymerase chain reaction. RESULTS: The results showed that coinfection of ALV-J and REV promoted the replication of each other. Thirty proteins, including TRIM62, NCK-association proteins 1 (NCKAP1, also known as Nap125), and Arp2/3-5, ARPC5, were identified. NCKAP1 and ARPC5 were involved in the actin cytoskeleton pathway. TRIM62 negatively regulated viral replication and that the inhibition of REV was more significant than that on ALV-J in CEF cells coinfected with TRIM62. In addition, TRIM62 decreased the expression of NCKAP1 and increased the expression of ARPC5 in coinfected CEF cells. CONCLUSIONS: Collectively, our results indicated that coinfection with ALV-J and REV competitively promoted each other's replication, the actin cytoskeleton played an important role in the coinfection mechanism, and TRIM62 regulated the actin cytoskeleton.

Up-to-date knowledge about the association between multiple sclerosis and the reactivation of human endogenous retrovirus infections.

BACKGROUND: Although existing studies show that reactivation of the human endogenous retrovirus (HERVs) plays a leading role in multiple sclerosis (MS) progression, the practitioners are yet to establish effective approaches for managing MS without jeopardizing the patients' immune systems. AIM: To provide up-to-date knowledge on the specific roles played by the reactivation of the HERVs in the pathogenesis of MS. MATERIALS AND METHODS: A systematic review of 70 peer-reviewed journals accessed via PubMed was conducted. The searches generated more than 600 sources that were evaluated based on three step in and exclusion criteria. The selected sources were critically analyzed vis-à-vis the paper's hypothesis which posits that the HERVs reactivation does not directly cause the MS, but triggers a demyelination process by promoting the pathogenic effects of the retroviruses. The paper further documents the advancements in the therapeutic applications resulting from the immunohistological analysis and pathological studies aimed at minimizing the adverse consequences of the HERVs reactivation. RESULTS AND DISCUSSIONS: Only three out of the 70 reviewed sources did not find provide evidence linking the reactivation of HERV and MS progression. On the other hand, overwhelming pieces of evidence confirm that the reactivations often drive the demyelinating plaques by initiating microglial inflammation. Pathological examinations reveal that the inflammatory monocytes (Ly6ChiCCR2 + CX3CR1lo) trigger the reactivation of the malignant T cells that are responsible for the progression of MS. These findings are promoting new discoveries as far as managing MS is concerned.

Mouse Siglec-1 Mediates trans-Infection of Surface-bound Murine Leukemia Virus in a Sialic Acid N-Acyl Side Chain-dependent Manner.

Siglec-1 (sialoadhesin, CD169) is a surface receptor on human cells that mediates trans-enhancement of HIV-1 infection through recognition of sialic acid moieties in virus membrane gangliosides. Here, we demonstrate that mouse Siglec-1, expressed on the surface of primary macrophages in an interferon-α-responsive manner, captures murine leukemia virus (MLV) particles and mediates their transfer to proliferating lymphocytes. The MLV infection of primary B-cells was markedly more efficient than that of primary T-cells. The major structural protein of MLV particles, Gag, frequently co-localized with Siglec-1, and trans-infection, primarily of surface-bound MLV particles, efficiently occurred. To explore the role of sialic acid for MLV trans-infection at a submolecular level, we analyzed the potential of six sialic acid precursor analogs to modulate the sialylated ganglioside-dependent interaction of MLV particles with Siglec-1. Biosynthetically engineered sialic acids were detected in both the glycolipid and glycoprotein fractions of MLV producer cells. MLV released from cells carrying N-acyl-modified sialic acids displayed strikingly different capacities for Siglec-1-mediated capture and trans-infection; N-butanoyl, N-isobutanoyl, N-glycolyl, or N-pentanoyl side chain modifications resulted in up to 92 and 80% reduction of virus particle capture and trans-infection, respectively, whereas N-propanoyl or N-cyclopropylcarbamyl side chains had no effect. In agreement with these functional analyses, molecular modeling indicated reduced binding affinities for non-functional N-acyl modifications. Thus, Siglec-1 is a key receptor for macrophage/lymphocyte trans-infection of surface-bound virions, and the N-acyl side chain of sialic acid is a critical determinant for the Siglec-1/MLV interaction.

Postinhibitory rebound neurons and networks are disrupted in retrovirus-induced spongiform neurodegeneration.

Certain retroviruses induce progressive spongiform motor neuron disease with features resembling prion diseases and amyotrophic lateral sclerosis. With the neurovirulent murine leukemia virus (MLV) FrCasE, Env protein expression within glia leads to postsynaptic vacuolation, cellular effacement, and neuronal loss in the absence of neuroinflammation. To understand the physiological changes associated with MLV-induced spongiosis, and its neuronal specificity, we employed patch-clamp recordings and voltage-sensitive dye imaging in brain slices of the mouse inferior colliculus (IC), a midbrain nucleus that undergoes extensive spongiosis. IC neurons characterized by postinhibitory rebound firing (PIR) were selectively affected in FrCasE-infected mice. Coincident with Env expression in microglia and in glia characterized by NG2 proteoglycan expression (NG2 cells), rebound neurons (RNs) lost PIR, became hyperexcitable, and were reduced in number. PIR loss and hyperexcitability were reversed by raising internal calcium buffer concentrations in RNs. PIR-initiated rhythmic circuits were disrupted, and spontaneous synchronized bursting and prolonged depolarizations were widespread. Other IC neuron cell types and circuits within the same degenerative environment were unaffected. Antagonists of NMDA and/or AMPA receptors reduced burst firing in the IC but did not affect prolonged depolarizations. Antagonists of L-type calcium channels abolished both bursts and slow depolarizations. IC infection by the nonneurovirulent isogenic virus Friend 57E (Fr57E), whose Env protein is structurally similar to FrCasE, showed no RN hyperactivity or cell loss; however, PIR latency increased. These findings suggest that spongiform neurodegeneration arises from the unique excitability of RNs, their local regulation by glia, and the disruption of this relationship by glial expression of abnormal protein.

Murine leukemia virus Gag localizes to the uropod of migrating primary lymphocytes.

UNLABELLED: B and CD4(+) T lymphocytes are natural targets of murine leukemia virus (MLV). Migrating lymphocytes adopt a polarized morphology with a trailing edge designated the uropod. Here, we demonstrate that MLV Gag localizes to the uropod in polarized B cells and CD4(+) T cells. The uropod localization of MLV Gag was dependent on plasma membrane (PM) association and multimerization of Gag but independent of the viral glycoprotein Env. Basic residues in MA that are required for MLV Gag recruitment to virological synapses between HEK293 and XC cells were dispensable for uropod localization in migrating B cells. Ultrastructural studies indicated that both wild-type and basic-residue mutant Gag localized to the outer surface of the PM at the uropod. Late-domain mutant virus particles were seen at the uropod in form of budding-arrested intermediates. Finally, uropods mediated contact between MLV-infected B cells and uninfected T cells to form virological synapses. Our results suggest that MLV, not unlike HIV, accumulates at the uropod of primary lymphocytes to facilitate viral spreading through the formation of uropod-mediated cell-cell contacts. IMPORTANCE: Viruses have evolved mechanisms to coordinate their assembly and budding with cell polarity to facilitate their spreading. In this study, we demonstrated that the viral determinants for MLV Gag to localize to the uropod in polarized B cells are distinct from the requirements to localize to virological synapses in transformed cell lines. Basic residues in MA that are required for the Gag localization to virological synapses between HEK293 and XC cells are dispensable for Gag localization to the uropod in primary B cells. Rather, plasma membrane association and capsid-driven multimerization of Gag are sufficient to drive MLV Gag to the uropod. MLV-laden uropods also mediate contacts between MLV-infected B cells and uninfected T cells to form virological synapses. Our results indicate that MLV accumulates at the uropod of primary lymphocytes to facilitate viral spreading through the formation of uropod-mediated cell-cell contacts.

Effects of simian betaretrovirus serotype 1 (SRV1) infection on the differentiation of hematopoietic progenitor cells (CD34+) derived from bone marrow of rhesus macaques (Macaca mulatta).

Peripheral blood cytopenias, particularly persistent anemia and neutropenia, are commonly associated with simian betaretrovirus infection of Asian monkeys of the genus Macaca. The pathogenetic mechanisms underlying these hematologic abnormalities are not well understood. The current study investigated the in vitro tropism of simian betaretrovirus (SRV) for both hematopoietic progenitor (CD34(+)) and stromal cells obtained from rhesus macaque bone marrow and assessed the effects of infection on hematopoietic progenitor cell differentiation in vitro. After in vitro exposure, SRV proviral DNA could be demonstrated by real-time PCR in cells and the reverse transcriptase assay in supernatants from SRV-exposed progenitor-associated stroma, but not in differentiated colonies derived from SRV-exposed progenitors. Furthermore, in vitro exposure involving cell-cell contact of uninfected CD34(+) progenitor cells with SRV-infected stromal cells resulted in a statistically significant reduction in granulocyte-macrophage colony formation in absence of detectable SRV-infection of progenitor cells. Reduction in colony formation occurred in a 'dose-dependent' fashion with increasing contact time. No effects on erythroid lineages and RBC differentiation were noted. Our results suggest that hematologic abnormalities observed during SRV disease (natural or experimental) of rhesus macaques may not result from direct effects of viral infection of progenitor cell populations, but rather be (at least in part) a consequence of SRV infection of supportive bone marrow stroma with secondary effects on differentiation of associated progenitor cells.

In situ force mapping of mammary gland transformation.

Tumor progression is characterized by an incremental stiffening of the tissue. The importance of tissue rigidity to cancer is appreciated, yet the contribution of specific tissue elements to tumor stiffening and their physiological significance remains unclear. We performed high-resolution atomic force microscopy indentation in live and snap-frozen fluorescently labeled mammary tissues to explore the origin of the tissue stiffening associated with mammary tumor development in PyMT mice. The tumor epithelium, the tumor-associated vasculature and the extracellular matrix all contributed to mammary gland stiffening as it transitioned from normal to invasive carcinoma. Consistent with the concept that extracellular matrix stiffness modifies cell tension, we found that isolated transformed mammary epithelial cells were intrinsically stiffer than their normal counterparts but that the malignant epithelium in situ was far stiffer than isolated breast tumor cells. Moreover, using an in situ vitrification approach, we determined that the extracellular matrix adjacent to the epithelium progressively stiffened as tissue evolved from normal through benign to an invasive state. Importantly, we also noted that there was significant mechanical heterogeneity within the transformed tissue both in the epithelium and the tumor-associated neovasculature. The vascular bed within the tumor core was substantially stiffer than the large patent vessels at the invasive front that are surrounded by the stiffest extracellular matrix. These findings clarify the contribution of individual mammary gland tissue elements to the altered biomechanical landscape of cancerous tissues and emphasize the importance of studying cancer cell evolution under conditions that preserve native interactions.

Relationships among rhinitis, fibromyalgia, and chronic fatigue.

New information about the pathophysiology of idiopathic nonallergic rhinopathy indicates a high prevalence in chronic fatigue syndrome (CFS). This article shows the relevance of CFS and allied disorders to allergy practice. CFS has significant overlap with systemic hyperalgesia (fibromyalgia), autonomic dysfunction (irritable bowel syndrome and migraine headaches), sensory hypersensitivity (dyspnea; congestion; rhinorrhea; and appreciation of visceral nociception in the esophagus, gastrointestinal tract, bladder, and other organs), and central nervous system maladaptations (central sensitization) recorded by functional magnetic resonance imaging (fMRI). Neurological dysfunction may account for the overlap of CFS with idiopathic nonallergic rhinopathy. Scientific advances are in fMRI, nociceptive sensor expression, and, potentially, infection with xenotropic murine leukemia-related virus provide additional insights to novel pathophysiological mechanisms of the "functional" complaints of these patients that are mistakenly interpreted as allergic syndromes. As allergists, we must accept the clinical challenges posed by these complex patients and provide proper diagnoses, assurance, and optimum care even though current treatment algorithms are lacking.

Microglial response to murine leukemia virus-induced encephalopathy is a good indicator of neuronal perturbations.

The neuronal pathology caused by neonatal infection of rats with the PVC-211 murine leukemia virus (PVC-211 MuLV) and its underlying mechanisms are not well defined even though a loss of neurons and spongiform neurodegeneration has been reported to accompany the disease. Here we sought to identify sites of neurodegeneration using microglial reactivity as an indirect marker and to characterize microglial activation during disease progression. Using a panel of microglial antibodies including Iba1, OX-42, ED1, and anti-ferritin, we have studied the response of microglial cells to neonatal CNS infection with PVC-211 at post-infection survival times 7, 14, 21, and 28 days. We found that microglial activation occurred primarily in the spinal cord and brainstem where it gradually increased in intensity over the time course of this study. Other brain areas were relatively unremarkable in their microglial reaction to viral infection within this time frame. However, the presence of activated microglial cells was not correlated directly with the presence of viral glycoprotein (gp70), which was expressed in endothelial cells throughout the CNS. Although double-labeling of microglia with Iba1 and ED1 revealed numerous actively phagocytic microglia during disease progression, not all activated microglia were ED1-positive. In addition to the intense microglial activation, we found increased ferritin expression sporadically throughout the virus-infected brain. The ferritin-positive cells were mostly microglia that exhibited dystrophic changes and likely represented a degenerating subpopulation of microglial cells. Thus, activated microglia can co-exist with degenerating microglia in the same brain region. We attempted to localize degenerating neurons or neurites using Fluoro-Jade, anti-tau, and anti-alpha synuclein staining, but none of these procedures yielded results to indicate obvious neuronal pathology. We conclude that the visualization of microglial activation is a more sensitive measure of neuronal perturbations than direct detection of neuronal pathology which may be subtle and not produce overt degenerative changes.

Tumour virology--history, status and future challenges.

Viruses enter host cells in order to complete their life cycles and have evolved to exploit host cell structures, regulatory factors and mechanisms. The virus and host cell interactions have consequences at multiple levels, spanning from evolution through disease to models and tools for scientific discovery and treatment. Virus-induced human cancers arise after a long duration of time and are monoclonal or oligoclonal in origin. Cancer is therefore a side effect rather than an essential part of viral infections in humans. Still, 15-20% of all human cancers are caused by viruses. A review of tumour virology shows its close integration in cancer research. Viral tools and experimental models have been indispensible for the progress of molecular biology. In particular, retroviruses and DNA tumour viruses have played major roles in our present understanding of the molecular biology of both viruses and the host. Recently, additional complex relationships due to virus and host co-evolution have appeared and may lead to a further understanding of the overall regulation of gene expression programmes in cancer.

Basic science summary.

The 16th Conference on Retroviruses and Opportunistic Infections featured a strong and balanced program that showcased exciting research into cellular restrictions that defend the cell against viral infection as well as cellular cofactors that regulate central steps in viral replication. Immunopathogenesis presentations continued to reveal some surprises such as evidence for pathogenicity in natural simian immunodeficiency virus infection. The identification of novel cellular restrictions and cellular cofactors of viral replication indicate the possibility of numerous opportunities for the development of novel therapeutic agents for the treatment of HIV and AIDS.

Retrovirus-specificity of regulatory T cells is neither present nor required in preventing retrovirus-induced bone marrow immune pathology.

Chronic viral infections of the hematopoietic system are associated with bone marrow dysfunction, to which both virus-mediated and immune-mediated effects may contribute. Using unresolving noncytopathic Friend virus (FV) infection in mice, we showed that unregulated CD4(+) T cell response to FV caused IFN-gamma-mediated bone marrow pathology and anemia. Importantly, bone marrow pathology was triggered by relative insufficiency in regulatory T (Treg) cells and was prevented by added Treg cells, which suppressed the local IFN-gamma production by FV-specific CD4(+) T cells. We further showed that the T cell receptor (TCR) repertoire of transgenic Treg cells expressing the beta chain of an FV-specific TCR was virtually devoid of FV-specific clones. Moreover, anemia induction by virus-specific CD4(+) T cells was efficiently suppressed by virus-nonspecific Treg cells. Thus, sufficient numbers of polyclonal Treg cells may provide substantial protection against bone marrow pathology in chronic viral infections.

Bridging fundamental RNA biology, retroviral replication, and oncogenesis: Karen Beemon wins the 2007 Retrovirology Prize.

The 2007 M. Jeang Retrovirology Prize has been awarded to Dr. Karen L. Beemon.

Comparative requirements for the restriction of retrovirus infection by TRIM5alpha and TRIMCyp.

The restriction factors, TRIM5alpha in most primates and TRIMCyp in owl monkeys, block infection of various retroviruses soon after virus entry into the host cell. Rhesus monkey TRIM5alpha (TRIM5alpha rh) inhibits human immunodeficiency virus (HIV-1) and feline immunodeficiency virus (FIV) more potently than human TRIM5alpha (TRIM5alpha hu). TRIMCyp restricts infection of HIV-1, simian immunodeficiency virus of African green monkeys (SIV agm) and FIV. Early after infection, TRIMCyp, like TRIM5alpha rh and TRIM5alpha hu, decreased the amount of particulate viral capsid in the cytosol of infected cells. The requirements for the TRIMCyp and TRIM5alpha domains in restricting different retroviruses were investigated. Potent restriction of FIV by TRIMCyp occurred in the complete absence of RING and B-box 2 domains; by contrast, efficient FIV restriction by TRIM5alpha rh required these domains. Variable region 1 of the TRIM5alpha rh B30.2 domain contributed to the potency of HIV-1, FIV and equine infectious anemia virus restriction. Thus, although differences exist in the requirements of TRIMCyp and TRIM5alpha for RING/B-box 2 domains, both restriction factors exhibit mechanistic similarities.

Clinical and virological characterization of persistent human infection with simian foamy viruses.

Persons occupationally exposed to nonhuman primates (NHPs) can be persistently infected with simian foamy virus (SFV). The clinical significance and person-to-person transmissibility of zoonotic SFV infection is unclear. Seven SFV-infected men responded to annual structured interviews and provided whole blood, oral, and urogenital specimens for study. Wives were tested for SFV infection. Proviral DNA was consistently detected by PCR in PBMCs of infected men and inconsistently in oral or urogenital samples. SFV was infrequently cultured from their PBMCs and throat swabs. Despite this and a long period of intimate exposure (median 20 years), wives were SFV negative. Most participants reported nonspecific symptoms and diseases common to aging. However, one of two persons with mild thrombocytopenia had clinically asymptomatic nonprogressive, monoclonal natural killer cell lymphocytosis of unclear relationship to SFV. All participants worked with NHPs before 1988 using mucocutaneous protection inconsistently; 57% described percutaneous injuries involving the infecting NHP species. SFV likely transmits to humans through both percutaneous and mucocutaneous exposures to NHP body fluids. Limited follow-up has not identified SFV-associated illness and secondary transmission among humans.

Increased proinflammatory cytokine and chemokine responses and microglial infection following inoculation with neural stem cells infected with polytropic murine retroviruses.

Proinflammatory cytokines and chemokines are often detected in brain tissue of patients with neurological diseases such as multiple sclerosis (MS), HIV-associated dementia (HAD) and Alzheimer's disease (AD). We have utilized a mouse model of retrovirus-induced neurological disease to examine how these proinflammatory responses contribute to neuropathogenesis. In previous studies with this model, a correlation was found between neurovirulence and cytokine and chemokine expression. However, it was unclear whether the induction of these cytokines and chemokines was in response to specific virus envelope determinants or was regulated by the level of virus infection in the brain. In the current study, we demonstrated that multiple polytropic retroviruses induced cytokine and chemokine mRNA expression following increased virus levels in the brain. Increased virus levels of polytropic viruses also correlated with increased neuropathogenesis. In contrast, the ecotropic retrovirus, FB29, did not induce cytokine or chemokine mRNA expression or neurological disease, despite virus levels either similar to or higher than the polytropic retroviruses. As polytropic and ecotropic viruses utilize different receptors for entry, these receptors may play a critical role in the induction of these innate immune responses in the brain.

Inclusion body disease in snakes: a review and description of three cases in boa constrictors in Belgium.

Inclusion body disease, a fatal disorder in Boidae, is reviewed, and three cases in boa constrictors, the first reported cases in Belgium, are described. The snakes showed nervous signs, and numerous eosinophilic intracytoplasmic inclusions, which are considered to be characteristic of the disease, were found in the liver and pancreas. The disease is suspected to be caused by a retrovirus, but transmission electron microscopic examinations of several tissues from one of the snakes did not reveal particles with a typical retroviral morphology.

Two surface-exposed elements of the B30.2/SPRY domain as potency determinants of N-tropic murine leukemia virus restriction by human TRIM5alpha.

Human TRIM5alpha (TRIM5alpha(hu)) potently restricts N-tropic (N-MLV), but not B-tropic, murine leukemia virus in a manner dependent upon residue 110 of the viral capsid. Rhesus monkey TRIM5alpha (TRIM5alpha(rh)) inhibits N-MLV only weakly. The study of human-monkey TRIM5alpha chimerae revealed that both the v1 and v3 variable regions of the B30.2/SPRY domain contain potency determinants for N-MLV restriction. These variable regions are predicted to be surface-exposed elements on one face of the B30.2 domain. Acidic residues in v3 complement basic residue 110 of the N-MLV capsid. The results support recognition of the retroviral capsid by the TRIM5alpha B30.2 domain.