Elevated endogenous retrovirus (ERV) transcription and anti-ERV antibody reactivity are implicated in lupus pathogenesis. Overproduction of non-ecotropic ERV (NEERV) envelope glycoprotein gp70 and ...resultant nephritis occur in lupus-prone mice, but whether NEERV mis-expression contributes to lupus etiology is unclear. Here we identified suppressor of NEERV (Snerv) 1 and 2, Krüppel-associated box zinc-finger proteins (KRAB-ZFPs) that repressed NEERV by binding the NEERV long terminal repeat to recruit the transcriptional regulator KAP1. Germline Snerv1/Snerv2 deletion increased activating chromatin modifications, transcription, and gp70 expression from NEERV loci. F1 crosses of lupus-prone New Zealand Black (NZB) and 129 mice to Snerv1/Snerv2−/− mice failed to restore NEERV repression, demonstrating that loss of SNERV underlies the lupus autoantigen gp70 overproduction that promotes nephritis in susceptible mice and that SNERV encodes for Sgp3 (in NZB mice) and Gv-1 loci (in 129 mice). Increased ERV expression in lupus patients inversely correlated with three putative ERV-suppressing KRAB-ZFPs, suggesting that loss of KRAB-ZFP-mediated ERV control may contribute to human lupus pathogenesis.
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•We identify the suppressor of non-ecotropic (NE) endogenous retroviruses (Snerv)•SNERV1 and SNERV2 are KRAB-ZFPs that bind to the NEERV LTR and recruit KAP1•Loss of SNERV1/SNERV2 underlies the lupus autoantigen gp70-associated loci Sgp3 and Gv1•Elevated ERV in SLE patients’ blood cells correlates with KRAB-ZFP dysregulation
Treger et al. identify Snerv as the genes, encoding Krüppel-associated box zinc-finger proteins (KRAB-ZFPs), that underlie the Sgp3 and Gv1 lupus-susceptibility loci in mice. SNERV represses expression of non-ecotropic endogenous retroviruses (ERVs). Elevated ERVs in lupus patients correlate with KRAB-ZFP down-regulation, suggesting a central role for dysfunctional ERV regulation in human lupus.
Endogenous retroviruses (ERV) are found throughout vertebrate genomes, and failure to silence their activation can have deleterious consequences on the host. Mutation and subsequent disruption of ERV ...loci is therefore an indispensable component of the cell-intrinsic defenses that maintain the integrity of the host genome. Abundant
and
evidence have revealed that APOBEC3 cytidine-deaminases, including human APOBEC3G (hA3G), can potently restrict retrotransposition; yet,
data demonstrating such activity is lacking, since no replication-competent human ERV have been identified. In mice deficient for Toll-like receptor 7 (TLR7), transcribed ERV loci can recombine and generate infectious ERV. In this study, we show that ectopic expression of hA3G can prevent the emergence of replication-competent, infectious ERV in
mice. Mice encode one copy of
in their genome. ERV reactivation in
mice was comparable in the presence or absence of
In contrast, expression of a human
transgene abrogated emergence of infectious ERV in the
background. No ERV RNA was detected in the plasma of hA3G
mice, and infectious ERV virions could not be amplified through coculture with permissive cells. These data reveal that hA3G can potently restrict active ERV
and suggest that expansion of the
locus in primates may have helped to provide for the continued restraint of ERV in the human genome.
Although APOBEC3 proteins are known to be important antiviral restriction factors in both mice and humans, their roles in the restriction of endogenous retroviruses (ERV) have been limited to
studies. Here, we report that human APOBEC3G expressed as a transgene in mice prevents the emergence of infectious ERV from endogenous loci. This study reveals that APOBEC3G can powerfully restrict active retrotransposons
and demonstrates how transgenic mice can be used to investigate host mechanisms that inhibit retrotransposons and reinforce genomic integrity.
Iron studies are critical for diagnosing iron deficiency and hemochromatosis. We present a case exhibiting macrocytic anemia with perplexingly high plasma iron concentrations.
The initial clinical ...presentation with significantly elevated iron results raised concerns for hemochromatosis. However, inconsistent results in dilution studies suggested the presence of an interfering substance. Inspection of the reaction curves from the instrument revealed very high background absorption in the 800 nm channel. This, coupled with the observation of an insoluble precipitate upon mixing the acid buffer reagent with the patient’s serum, as well as the patient’s high total protein and low albumin levels, suggested immunoglobulin overproduction. Serum protein electrophoresis confirmed a monoclonal gammopathy with a subsequent diagnosis of multiple myeloma.
Excessive monoclonal immunoglobulins can precipitate in acidic buffers and interfere with spectrophotometric measurements in iron testing. Although challenging, investigating an interference and determining its cause can uncover underlying diseases that have yet to be diagnosed.
Abstract
Objectives
To evaluate the real-world performance and reference intervals of the Binding Site Freelite serum free light chain (SFLC) assay (Thermo Fisher Scientific), a global standard for ...diagnosis, prognostication, and response assessment for monoclonal gammopathies.
Methods
An informatics-based approach was used to retrospectively evaluate concordance between SFLC and the orthogonal Sebia HYDRASYS immunofixation assay results in a large clinical data set consecutively reported between 2010 and 2020.
Results
Among patients with monoclonal-negative results by both SFLC and Sebia HYDRASYS immunofixation assays, 25% (1226/5057) had κ/λ ratios (KLRs) outside the manufacturer-defined and International Myeloma Working Group–cited normal reference interval of 0.26 to 1.65. These results were consistent over the study period and were not affected by sex, age, impaired kidney function, or assay antisera lot variation. Assay drift, in addition to other potential factors, affected the KLR distribution. Using International Statistical Classification of Diseases (ICD) codes, kidney function data, and the central 95% of KLR values generated on the Optilite platform (Thermo Fisher Scientific), we derived a new reference interval of 0.67 to 2.13, reducing the KLR false-positive rate to 8%. However, normal KLR persisted among 16% (14/85) of samples with free λ chains by immunofixation, warranting caution during interpretation.
Conclusions
Our analysis indicated that revision of Freelite SFLC reference intervals improves assay interpretation and should prompt reconsideration of Freelite reference intervals worldwide.
Endogenous retroviruses (ERVs) are genomic sequences that originated from retroviruses and are present in most eukaryotic genomes. Both beneficial and detrimental functions are attributed to ERVs, ...but whether ERVs contribute to antiviral immunity is not well understood. Here, we used herpes simplex virus type 2 (HSV-2) infection as a model and found that Toll-like receptor 7 (
) deficient mice that have high systemic levels of infectious ERVs are protected from intravaginal HSV-2 infection and disease, compared to wildtype C57BL/6 mice. We deleted the endogenous ecotropic murine leukemia virus (Emv2) locus on the
background (
) and found that
mice lose protection against HSV-2 infection. Intravaginal application of purified ERVs from
mice prior to HSV-2 infection delays disease in both wildtype and highly susceptible interferon-alpha receptor-deficient (
) mice. However, intravaginal ERV treatment did not protect
mice from HSV-2 disease, suggesting that the protective mechanism mediated by exogenous ERV treatment may differ from that of constitutively and systemically expressed ERVs in
mice. We did not observe enhanced type I interferon (IFN-I) signaling in the vaginal tissues from Tlr7-/- mice, and instead found enrichment in genes associated with extracellular matrix organization. Together, our results revealed that constitutive and/or systemic expression of ERVs protect mice against vaginal HSV-2 infection and delay disease.
Autoantibodies that bind self-antigens are a hallmark of autoimmune diseases, but can also be present in healthy individuals. Clinical assays that detect and titer antigen-specific autoantibodies are ...an important component of the diagnosis and monitoring of autoimmune diseases. Autoantibodies may contribute to disease pathogenesis via effector functions that are dictated by both the antigen-binding site and constant domain.
In this review, we discuss features of antibodies, in addition to antigen-binding specificity, which determine effector function. These features include class, subclass, allotype, and glycosylation. We discuss emerging data indicating that analysis of these antibody features may be informative for diagnosis and monitoring of autoimmune diseases. We also consider methodologies to interrogate these features and consider how they could be implemented in the clinical laboratory.
Future autoantibody assays may incorporate assessment of additional antibody features that contribute to autoimmune disease pathogenesis and provide added clinical value.
Endogenous retroviruses (ERV) are transposable retroelements that form ~10% of the murine genome and whose family members are differentially expressed throughout embryogenesis. However, precise ...regulation of ERV in germ cells remains unclear. To investigate ERV expression in oocytes, we adapted a single-cell mRNA-sequencing library preparation method to generate bulk sequencing libraries from growing oocytes in a time- and cost-efficient manner. Here, we present a modified Smart-seq2 protocol that yields full-length cDNA libraries from purified RNA obtained from low numbers of pooled immature or mature oocytes. Using this method, RNA-sequencing libraries can be generated from any rare or difficult-to-isolate populations for subsequent sequencing and retroelement expression analysis.
Abstract
Endogenous retroviruses (ERVs), comprising a substantial portion of the vertebrate genome, are remnants of ancient genetic invaders. Although multiple layers of cell-intrinsic control are ...utilized to prevent retroviral reactivation, ERVs with intact coding potential are able to reactivate in immunodeficient mice. While previous studies have indicated that B cells are indispensable for preventing ERV reactivation, it is not yet clear which B cell population mediates the blockade of ERV emergence to prevent subsequent damage in the host. Here, we employed direct labeling of B cells reactive with emerged ERV particles to characterize the B cell population and clonal repertoire responsible for recognition of ERV, and to study the mechanism by which B cells provide protection against ERV emergence. We found that ERV-reactive B cells are enriched in innate-like B-1 cell compartment that predominantly reside in peritoneal and pleural cavities. B-1 cells specificity is biased toward conserved epitopes shared by bacterial- and self-antigens due to distinct developmental pathways in fetal haematopoiesis. We identified ERV-reactive antibodies in unimmunized mice, the level of which further increases upon innate sensor stimulation. B cell receptor repertoire profiling of ERV-reactive B-1 cells revealed increased usage of Igh VH genes that give rise to glycan-specific antibodies targeting glycan structures exhibited by bacterial and viral antigens. We demonstrated that these glycan-specific natural antibodies engage complement pathway to facilitate clearance of reactivated ERV particles. In conclusion, we elucidated the role of glycan-specific B-1 cells and secreted natural antibodies in mediating blockade of ERV emergence.
Abstract
Endogenous retroviruses (ERVs) are genomic sequences that originated from retroviruses and are present in most eukaryotic genomes. Both beneficial and detrimental functions are attributed to ...ERVs, but whether ERVs contribute to antiviral immunity is not known. Here we used herpes simplex virus type 2 (HSV-2) infection and found that mice deficient in Toll-like receptor 7 (Tlr7−/−) that have high systemic levels of infectious ERVs are resistant to intravaginal HSV-2 infection compared with wildtype C57BL/6 (B6) mice. We deleted the endogenous ecotropic murine leukemia virus (Emv2) locus on the Tlr7−/− background (Emv2−/−Tlr7−/−) and found that Emv2−/−Tlr7−/− mice are no longer protected against HSV-2. Intravaginal application of purified ERVs prior to HSV-2 infection in both B6 and highly susceptible interferon-alpha receptor-deficient (Ifnar1−/−) mice delayed disease course. We did not observe enhanced type I interferon (IFN-I) signaling in the vaginal tissues from Tlr7−/− mice or B6 mice treated with purified ERVs. Instead, we observed enhanced expression of epithelial tight junction protein, E-cadherin, in the vaginal epithelium of ERV-treated B6 mice. Similar increase in tight junction proteins was observed in Tlr7−/− but not in the Emv2−/−Tlr7−/− mice. Together, our results showed that IFN-independent modulation of the vaginal epithelium by ERVs protects mice against vaginal HSV-2 infection and lowers disease burden.
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