Biallelic NAA60 variants with impaired n-terminal acetylation capacity cause autosomal recessive primary familial brain calcifications.

Primary familial brain calcification (PFBC) is characterized by calcium deposition in the brain, causing progressive movement disorders, psychiatric symptoms, and cognitive decline. PFBC is a heterogeneous disorder currently linked to variants in six different genes, but most patients remain genetically undiagnosed. Here, we identify biallelic NAA60 variants in ten individuals from seven families with autosomal recessive PFBC. The NAA60 variants lead to loss-of-function with lack of protein N-terminal (Nt)-acetylation activity. We show that the phosphate importer SLC20A2 is a substrate of NAA60 in vitro. In cells, loss of NAA60 caused reduced surface levels of SLC20A2 and a reduction in extracellular phosphate uptake. This study establishes NAA60 as a causal gene for PFBC, provides a possible biochemical explanation of its disease-causing mechanisms and underscores NAA60-mediated Nt-acetylation of transmembrane proteins as a fundamental process for healthy neurobiological functioning.

A co-opted endogenous retroviral envelope promotes cell survival by controlling CTR1-mediated copper transport and homeostasis.

Copper is a critical element for eukaryotic life involved in numerous cellular functions, including redox balance, but is toxic in excess. Therefore, tight regulation of copper acquisition and homeostasis is essential for cell physiology and survival. Here, we identify a different regulatory mechanism for cellular copper homeostasis that requires the presence of an endogenous retroviral envelope glycoprotein called Refrex1. We show that cells respond to elevated extracellular copper by increasing the expression of Refrex1, which regulates copper acquisition through interaction with the main copper transporter CTR1. Downmodulation of Refrex1 results in intracellular copper accumulation leading to reactive oxygen species (ROS) production and subsequent apoptosis, which is prevented by copper chelator treatment. Our results show that Refrex1 has been co-opted for its ability to regulate copper entry through CTR1 in order to limit copper excess, redox imbalance, and ensuing cell death, strongly suggesting that other endogenous retroviruses may have similar metabolic functions among vertebrates.

Identification of Copper Transporter 1 as a Receptor for Feline Endogenous Retrovirus ERV-DC14.

Vertebrates harbor hundreds of endogenous retroviral (ERV) sequences in their genomes, which are considered signs of past infections that occurred during evolution. On rare occasions, ERV genes like env are maintained and coopted by hosts for physiological functions, but they also participate in recombination events with exogenous retroviruses to generate rearranged viruses with novel tropisms. In domestic cats, feline leukemia virus type D (FeLV-D) has been described as a recombinant virus between the infectious FeLV-A and likely the ERV-DC14 env gene that resulted in an extended tropism due to the usage of a new uncharacterized retroviral receptor. Here, we report the identification of SLC31A1 encoding the copper transporter 1 (CTR1) as a susceptibility gene for ERV-DC14 infection. Expression of human CTR1 into nonpermissive cells was sufficient to confer sensitivity to ERV-DC14 pseudotype infection and to increase the binding of an ERV-DC14 Env ligand. Moreover, inactivation of CTR1 by genome editing or cell surface downmodulation of CTR1 by a high dose of copper dramatically decreased ERV-DC14 infection and binding, while magnesium treatment had no effect. We also investigated the role of CTR1 in the nonpermissivity of feline and hamster cells. While feline CTR1 was fully functional for ERV-DC14, we found that binding was strongly reduced upon treatment with conditioned medium of feline cells, suggesting that the observed resistance to infection was a consequence of CTR1 saturation. In contrast, hamster CTR1 was inactive due to the presence of a N-linked glycosylation site at position 27, which is absent in the human ortholog. These results provide evidence that CTR1 is a receptor for ERV-DC14. Along with chimpanzee endogenous retrovirus type 2, ERV-DC14 is the second family of endogenous retrovirus known to have used CTR1 during past infections of vertebrates. IMPORTANCE Receptor usage is an important determinant of diseases induced by pathogenic retroviruses. In the case of feline leukemia viruses, three subgroups (A, B, and C) based on their ability to recognize different cell host receptors, respectively, the thiamine transporter THTR1, the phosphate transporter PiT1, and the heme exporter FLVCR1, are associated with distinct feline diseases. FeLV-A is horizontally transmitted and found in all naturally infected cats, while FeLV-B and FeLV-C have emerged from FeLV-A, respectively, by recombination with endogenous retroviral env sequences or by mutations in the FeLV-A env gene, both leading to a switch in receptor usage and in subsequent in vivo tropism. Here, we set up a genetic screen to identify the retroviral receptor of ERV-DC14, a feline endogenous provirus whose env gene has been captured by infectious FeLV-A to give rise to FeLV-D in a process similar to FeLV-B. Our results reveal that the copper transporter CTR1 was such a receptor and provide new insights into the acquisition of an expanded tropism by FeLV-D.

Haploinsufficiency of the Primary Familial Brain Calcification Gene SLC20A2 Mediated by Disruption of a Regulatory Element.

OBJECTIVE: Primary familial brain calcification (PFBC) is a rare cerebral microvascular calcifying disorder with diverse neuropsychiatric expression. Five genes were reported as PFBC causative when carrying pathogenic variants. Haploinsufficiency of SLC20A2, which encodes an inorganic phosphate importer, is a major cause of autosomal-dominant PFBC. However, PFBC remains genetically unexplained in a proportion of patients, suggesting the existence of additional genes or cryptic mutations. We analyzed exome sequencing data of 71 unrelated, genetically unexplained PFBC patients with the aim to detect copy number variations that may disrupt the expression of core PFBC-causing genes. METHODS: After the identification of a deletion upstream of SLC20A2, we assessed its consequences on gene function by reverse transcriptase droplet digital polymerase chain reaction (RT-ddPCR), an ex vivo inorganic phosphate uptake assay, and introduced the deletion of a putative SLC20A2 enhancer mapping to this region in human embryonic kidney 293 (HEK293) cells by clustered regularly interspaced short palindromic repeats (CRISPR) - CRISPR-associated protein 9 (Cas9). RESULTS: The 8p11.21 deletion, segregating with PFBC in a family, mapped 35 kb upstream of SLC20A2. The deletion carriers/normal controls ratio of relative SLC20A2 mRNA levels was 60.2% (P < 0.001). This was comparable with that of patients carrying an SLC20A2 premature stop codon (63.4%; P < 0.001). The proband exhibited a 39.3% decrease of inorganic phosphate uptake in blood (P = 0.015). In HEK293 cells, we observed a 39.8% decrease in relative SLC20A2 mRNA levels after normalization on DNA copy number (P < 0.001). DISCUSSION: We identified a deletion of an enhancer of SLC20A2 expression, with carriers showing haploinsufficiency in similar ranges to loss-of-function alleles, and we observed reduced mRNA levels after deleting this element in a cellular model. We propose a 3-step strategy to identify and easily assess the effect of such events. © 2020 International Parkinson and Movement Disorder Society.

Interplay between primary familial brain calcification-associated SLC20A2 and XPR1 phosphate transporters requires inositol polyphosphates for control of cellular phosphate homeostasis.

Solute carrier family 20 member 2 (SLC20A2) and xenotropic and polytropic retrovirus receptor 1 (XPR1) are transporters with phosphate uptake and efflux functions, respectively. Both are associated with primary familial brain calcification (PFBC), a genetic disease characterized by cerebral calcium-phosphate deposition and associated with neuropsychiatric symptoms. The association of the two transporters with the same disease suggests that they jointly regulate phosphate fluxes and cellular homeostasis, but direct evidence is missing. Here, we found that cross-talk between SLC20A2 and XPR1 regulates phosphate homeostasis, and we identified XPR1 as a key inositol polyphosphate (IP)-dependent regulator of this process. We found that overexpression of WT SLC20A2 increased phosphate uptake, as expected, but also unexpectedly increased phosphate efflux, whereas PFBC-associated SLC20A2 variants did not. Conversely, SLC20A2 depletion decreased phosphate uptake only slightly, most likely compensated for by the related SLC20A1 transporter, but strongly decreased XPR1-mediated phosphate efflux. The SLC20A2-XPR1 axis maintained constant intracellular phosphate and ATP levels, which both increased in XPR1 KO cells. Elevated ATP levels are a hallmark of altered inositol pyrophosphate (PP-IP) synthesis, and basal ATP levels were restored after phosphate efflux rescue with WT XPR1 but not with XPR1 harboring a mutated PP-IP-binding pocket. Accordingly, inositol hexakisphosphate kinase 1-2 (IP6K1-2) gene inactivation or IP6K inhibitor treatment abolished XPR1-mediated phosphate efflux regulation and homeostasis. Our findings unveil an SLC20A2-XPR1 interplay that depends on IPs such as PP-IPs and controls cellular phosphate homeostasis via the efflux route, and alteration of this interplay likely contributes to PFBC.

Characterization of XPR1/SLC53A1 variants located outside of the SPX domain in patients with primary familial brain calcification.

Primary familial brain calcification (PFBC) is a rare neurological disease characterized by deposits of calcium phosphate in the basal ganglia and other regions of the brain. Pathogenic variants in the XPR1/SLC53A1 gene, which encodes the only known inorganic phosphate exporter, cause an autosomal dominant form of PFBC. These variants are typically located in the SPX N-terminal domain of the protein. Here, we characterize three XPR1 variants outside of SPX in three PFBC patients with an apparently sporadic presentation: c.1375C > T p.(R459C), c.1855A > G p.(N619D) and c.1886T > G p.(I629S), with the latter identified as the first XPR1/SLC53A1 de novo mutation to occur in a PFBC proband. When tested in an in vitro physiological complementation assay, the three XPR1 variants were impaired in phosphate export function, although they were normally expressed at the cell surface and could serve as functional receptors for retrovirus entry. Moreover, peripheral blood cells from the p.N619D patient could be assayed ex vivo and displayed significantly impaired phosphate export. Our results establish for the first time the clinical and molecular characteristics of XPR1 variants located outside the SPX domain and assert a direct link between these variants, deficient phosphate export, and PFBC. Moreover, we unveiled new structural features in XPR1 C-terminal domain that play a role in phosphate export and disease.

Cyclophilins and nucleoporins are required for infection mediated by capsids from circulating HIV-2 primary isolates.

HIV-2 groups have emerged from sooty mangabey SIV and entered the human population in Africa on several separate occasions. Compared to world pandemic HIV-1 that arose from the chimpanzee SIVcpz virus, the SIVsm-derived HIV-2, largely confined to West Africa, is less replicative, less transmissible and less pathogenic. Here, we evaluated the interactions between host cellular factors, which control HIV-1 infection and target the capsid, and HIV-2 capsids obtained from primary isolates from patients with different disease progression status. We showed that, like HIV-1, all HIV-2 CA we tested exhibited a dependence on cyclophilin A. However, we observed no correlation between HIV-2 viremia and susceptibility to hu-TRIM5alpha or dependence to CypA. Finally, we found that all CA from HIV-2 primary isolates exploit Nup358 and Nup153 for nucleus transposition. Altogether, these findings indicate that the ability to use the two latter nucleoporins is essential to infection of human cells for both HIV-1 and HIV-2. This dependence provides another molecular target that could be used for antiviral strategies against both HIV-1 and 2, based on both nucleoporins.

XPR1 mutations are a rare cause of primary familial brain calcification.

Mutations in XPR1, a gene encoding an inorganic phosphate exporter, have recently been identified in patients with primary familial brain calcification (PFBC). Using Sanger sequencing, we screened XPR1 in 18 unrelated patients with PFBC and no SLC20A2, PDGFB, or PDGFRB mutation. XPR1 variants were tested in an in vitro physiological complementation assay and patient blood cells were assessed ex vivo for phosphate export. We identified a novel c.260T > C, p.(Leu87Pro) XPR1 variant in a 41-year-old man complaining of micrographia and dysarthria and demonstrating mild parkinsonism, cerebellar ataxia and executive dysfunction. Brain (123)I-Ioflupane scintigraphy showed marked dopaminergic neuron loss. Peripheral blood cells from the patient exhibited decreased phosphate export. XPR1 in which we introduced the mutation was not detectable at the cell surface and did not lead to phosphate export. These results confirm that loss of XPR1-mediated phosphate export function causes PFBC, occurring in less than 8 % of cases negative for the other genes, and may be responsible for parkinsonism.

Cytostatic Effect of Repeated Exposure to Simvastatin: A Mechanism for Chronic Myotoxicity Revealed by the Use of Mesodermal Progenitors Derived from Human Pluripotent Stem Cells.

Statin treatment of hypercholesterolemia can lead to chronic myotoxicity which is, in most cases, alleviated by drug withdrawal. Cellular and molecular mechanisms of this adverse effect have been elusive, in particular because of the lack of in vitro models suitable for long-term exposures. We have taken advantage of the properties of human pluripotent stem cell-derived mesodermal precursors, that can be maintained unaltered in vitro for a long period of time, to develop a model of repeated exposures to simvastatin during more than 2 weeks. This approach unveiled major differences, both in functional and molecular terms, in response to single versus repeated-dose exposures to simvastatin. The main functional effect of the in vitro simvastatin-induced long-term toxicity was a loss of proliferative capacity in the absence of concomitant cell death, revealing that cytostatic effect could be a major contributor to statin-induced myotoxicity. Comparative analysis of molecular modifications induced by simvastatin short-term versus prolonged exposures demonstrated powerful adaptive cell responses, as illustrated by the dramatic decrease in the number of differentially expressed genes, distinct biological pathway enrichments, and distinct patterns of nutrient transporters expressed at the cell surface. This study underlines the potential of derivatives of human pluripotent stem cells for developing new approaches in toxicology, in particular for chronic toxicity testing.

Mutations in XPR1 cause primary familial brain calcification associated with altered phosphate export.

Primary familial brain calcification (PFBC) is a neurological disease characterized by calcium phosphate deposits in the basal ganglia and other brain regions and has thus far been associated with SLC20A2, PDGFB or PDGFRB mutations. We identified in multiple families with PFBC mutations in XPR1, a gene encoding a retroviral receptor with phosphate export function. These mutations alter phosphate export, implicating XPR1 and phosphate homeostasis in PFBC.

Glucose and glutamine metabolism regulate human hematopoietic stem cell lineage specification.

The metabolic state of quiescent hematopoietic stem cells (HSCs) is an important regulator of self-renewal, but it is unclear whether or how metabolic parameters contribute to HSC lineage specification and commitment. Here, we show that the commitment of human and murine HSCs to the erythroid lineage is dependent upon glutamine metabolism. HSCs require the ASCT2 glutamine transporter and active glutamine metabolism for erythroid specification. Blocking this pathway diverts EPO-stimulated HSCs to differentiate into myelomonocytic fates, altering in vivo HSC responses and erythroid commitment under stress conditions such as hemolytic anemia. Mechanistically, erythroid specification of HSCs requires glutamine-dependent de novo nucleotide biosynthesis. Exogenous nucleosides rescue erythroid commitment of human HSCs under conditions of limited glutamine catabolism, and glucose-stimulated nucleotide biosynthesis further enhances erythroid specification. Thus, the availability of glutamine and glucose to provide fuel for nucleotide biosynthesis regulates HSC lineage commitment under conditions of metabolic stress.

Heterogeneous susceptibility of circulating SIV isolate capsids to HIV-interacting factors.

BACKGROUND: Many species of non-human primates in Africa are naturally infected by simian immunodeficiency viruses (SIV) and humans stand at the forefront of exposure to these viruses in Sub-Saharan Africa. Cross-species transmission and adaptation of SIV to humans have given rise to human immunodeficiency viruses (HIV-1 and HIV-2) on twelve accountable, independent occasions. However, the determinants contributing to a simian-to-human lasting transmission are not fully understood. Following entry, viral cores are released into the cytoplasm and become the principal target of host cellular factors. Here, we evaluated cellular factors likely to be involved in potential new SIV cross-species transmissions. We investigated the interactions of capsids from naturally circulating SIV isolates with both HIV-1 restricting (i.e. TRIM5 proteins) and facilitating (i.e. cyclophilin A and nucleopore-associated Nup358/RanBP2 and Nup153) factors in single-round infectivity assays that reproduce early stages of the viral life-cycle. RESULTS: We show that human TRIM5α is unlikely to prevent cross-species transmission of any SIV we tested and observed that the SIV CA-CypA interaction is a widespread but not a universal feature. Moreover, entry in the nucleus of different SIV appeared to follow pathways that do not necessarily recruit Nup358/RanBP2 or Nup153, and this regardless of their interaction with CypA. Nevertheless, we found that, like HIV-1, human-adapted HIV-2 infection was dependent on Nup358/RanBP2 and Nup153 interactions for optimal infection. Furthermore, we found that, unlike HIV CA, SIV CA did not require a direct interaction with the Cyp-like domain of Nup358/RanBP2 to carry out successful infection. CONCLUSIONS: Circulating SIV present a variety of phenotypes with regard to CA-interacting restricting or facilitating factors. Altogether, we unveiled unidentified pathways for SIV CA, which could also be exploited by HIV in different cellular contexts, to drive entry into the nucleus. Our findings warrant a closer evaluation of other potential defenses against circulating SIV.

Inorganic phosphate export by the retrovirus receptor XPR1 in metazoans.

Inorganic phosphate uptake is a universal function accomplished by transporters that are present across the living world. In contrast, no phosphate exporter has ever been identified in metazoans. Here, we show that depletion of XPR1, a multipass membrane molecule initially identified as the cell-surface receptor for xenotropic and polytropic murine leukemia retroviruses (X- and P-MLV), induced a decrease in phosphate export and that reintroduction of various XPR1 proteins, from fruit fly to human, rescued this defect. Inhibition of phosphate export was also obtained with a soluble ligand generated from the envelope-receptor-binding domain of X-MLV in all human cell lines tested, as well as in diverse stem cells and epithelial cells derived from renal proximal tubules, the main site of phosphate homeostasis regulation. These results provide new insights on phosphate export in metazoans and the role of Xpr1 in this function.

Metabolic adaptation of neutrophils in cystic fibrosis airways involves distinct shifts in nutrient transporter expression.

Inflammatory conditions can profoundly alter human neutrophils, a leukocyte subset generally viewed as terminally differentiated and catabolic. In cystic fibrosis (CF) patients, neutrophils recruited to CF airways show active exocytosis and sustained phosphorylation of prosurvival, metabolic pathways. Because the CF airway lumen is also characterized by high levels of free glucose and amino acids, we compared surface expression of Glut1 (glucose) and ASCT2 (neutral amino acids) transporters, as well as that of PiT1 and PiT2 (inorganic phosphate transporters), in blood and airway neutrophils, using specific retroviral envelope-derived ligands. Neither nutrient transporter expression nor glucose uptake was altered on blood neutrophils from CF patients compared with healthy controls. Notably, however, airway neutrophils of CF patients had higher levels of PiT1 and Glut1 and increased glucose uptake compared with their blood counterparts. Based on primary granule exocytosis and scatter profiles, CF airway neutrophils could be divided into two subsets, with one of the subsets characterized by more salient increases in Glut1, ASCT2, PiT1, and PiT2 expression. Moreover, in vitro exocytosis assays of blood neutrophils suggest that surface nutrient transporter expression is not directly associated with primary (or secondary) granule exocytosis. Although expression of nutrient transporters on CF blood or airway neutrophils was not altered by genotype, age, gender, or Pseudomonas aeruginosa infection, oral steroid treatment decreased Glut1 and PiT2 levels in blood neutrophils. Thus, neutrophils recruited from blood into the CF airway lumen display augmented cell surface nutrient transporter expression and glucose uptake, consistent with metabolic adaptation.

Optimization of tumor xenograft dissociation for the profiling of cell surface markers and nutrient transporters.

Metabolic adaptations and changes in the expression of nutrient transporters are known to accompany tumorigenic processes. Nevertheless, in the context of solid tumors, studies of metabolism are hindered by a paucity of tools allowing the identification of cell surface transporters on individual cells. Here, we developed a method for the dissociation of human breast cancer tumor xenografts combined with quantification of cell surface markers, including metabolite transporters. The expression profiles of four relevant nutrient transporters for cancer cells' metabolism, Glut1, ASCT2, PiT1 and PiT2 (participating to glucose, glutamine and inorganic phosphate, respectively), as detected by new retroviral envelope glycoprotein-derived ligands, were distinctive of each tumor, unveiling underlying differences in metabolic pathways. Our tumor dissociation procedure and nutrient transporter profiling technology provides opportunities for future basic research, clinical diagnosis, prognosis and evaluation of therapeutic responses, as well as for drug discovery and development.

Glut1-mediated glucose transport regulates HIV infection.

Cell cycle entry is commonly considered to positively regulate HIV-1 infection of CD4 T cells, raising the question as to how quiescent lymphocytes, representing a large portion of the viral reservoir, are infected in vivo. Factors such as the homeostatic cytokine IL-7 have been shown to render quiescent T cells permissive to HIV-1 infection, presumably by transiently stimulating their entry into the cell cycle. However, we show here that at physiological oxygen (O(2)) levels (2-5% O(2) tension in lymphoid organs), IL-7 stimulation generates an environment permissive to HIV-1 infection, despite a significantly attenuated level of cell cycle entry. We identify the IL-7-induced increase in Glut1 expression, resulting in augmented glucose uptake, as a key factor in rendering these T lymphocytes susceptible to HIV-1 infection. HIV-1 infection of human T cells is abrogated either by impairment of Glut1 signal transduction or by siRNA-mediated Glut1 down-regulation. Consistent with this, we show that the susceptibility of human thymocyte subsets to HIV-1 infection correlates with Glut1 expression; single-round infection is markedly higher in the Glut1-expressing double-positive thymocyte population than in any of the Glut1-negative subsets. Thus, our studies reveal the Glut1-mediated metabolic pathway as a critical regulator of HIV-1 infection in human CD4 T cells and thymocytes.

Xpr1 is an atypical G-protein-coupled receptor that mediates xenotropic and polytropic murine retrovirus neurotoxicity.

Xenotropic murine leukemia virus-related virus (XMRV) was first identified in human prostate cancer tissue and was later found in a high percentage of humans with chronic fatigue syndrome (CFS). While exploring potential disease mechanisms, we found that XMRV infection induced apoptosis in SY5Y human neuroblastoma cells, suggesting a mechanism for the neuromuscular pathology seen in CFS. Several lines of evidence show that the cell entry receptor for XMRV, Xpr1, mediates this effect, and chemical cross-linking studies show that Xpr1 is associated with the Gß subunit of the G-protein heterotrimer. The activation of adenylate cyclase rescued the cells from XMRV toxicity, indicating that toxicity resulted from reduced G-protein-mediated cyclic AMP (cAMP) signaling. Some proteins with similarity to Xpr1 are involved in phosphate uptake into cells, but we found no role of Xpr1 in phosphate uptake or its regulation. Our results indicate that Xpr1 is a novel, atypical G-protein-coupled receptor (GPCR) and that xenotropic or polytropic retrovirus binding can disrupt the cAMP-mediated signaling function of Xpr1, leading to the apoptosis of infected cells. We show that this pathway is also responsible for the classic toxicity of the polytropic mink cell focus-forming (MCF) retrovirus in mink cells. Although it now seems clear that the detection of XMRV in humans was the result of sample contamination with a recombinant mouse virus, our findings may have relevance to neurologic disease induced by MCF retroviruses in mice.

Regional characterization of energy metabolism in the brain of normal and MPTP-intoxicated mice using new markers of glucose and phosphate transport.

The gibbon ape leukemia virus (GALV), the amphotropic murine leukemia virus (AMLV) and the human T-cell leukemia virus (HTLV) are retroviruses that specifically bind nutrient transporters with their envelope glycoproteins (Env) when entering host cells. Here, we used tagged ligands derived from GALV, AMLV, and HTLV Env to monitor the distribution of their cognate receptors, the inorganic phosphate transporters PiT1 and PiT2, and the glucose transporter GLUT1, respectively, in basal conditions and after acute energy deficiency. For this purpose, we monitored changes in the distribution of PiT1, PiT2 and GLUT1 in the cerebellum, the frontal cortex, the corpus callosum, the striatum and the substantia nigra (SN) of C57/BL6 mice after administration of 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridinium (MPTP), a mitochondrial complex I inhibitor which induces neuronal degeneration in the striato-nigral network.The PiT1 ligand stained oligodendrocytes in the corpus callosum and showed a reticular pattern in the SN. The PiT2 ligand stained particularly the cerebellar Purkinje cells, while GLUT1 labelling was mainly observed throughout the cortex, basal ganglia and cerebellar gray matter. Interestingly, unlike GLUT1 and PiT2 distributions which did not appear to be modified by MPTP intoxication, PiT1 immunostaining seemed to be more extended in the SN. The plausible reasons for this change following acute energy stress are discussed.These new ligands therefore constitute new metabolic markers which should help to unravel cellular adaptations to a wide variety of normal and pathologic conditions and to determine the role of specific nutrient transporters in tissue homeostasis.

Cell surface expression of the bovine leukemia virus-binding receptor on B and T lymphocytes is induced by receptor engagement.

Bovine leukemia virus (BLV), one of the most common infectious viruses of cattle, is endemic in many herds. Approximately 30-40% of adult cows in the United States are infected by this oncogenic C-type retrovirus and 1-5% of animals will eventually develop a malignant lymphoma. BLV, like the human and simian T cell leukemia viruses, is a deltaretrovirus but, in contrast with the latter, the BLV receptor remains unidentified. In this study, we demonstrate that the amino-terminal 182 residues of the BLV envelope glycoprotein surface unit encompasses the receptor-binding domain. A bona fide interaction of this receptor-binding domain with the BLV receptor was demonstrated by specific interference with BLV, but not human T cell leukemia virus, envelope glycoprotein-mediated binding. We generated a rabbit Ig Fc-tagged BLV receptor-binding domain construct and ascertained that the ligand binds the BLV receptor on target cells from multiple species. Using this tool, we determined that the BLV-binding receptor is expressed on differentiating pro/pre-B cells in mouse bone marrow. However, the receptor was not detected on mature/quiescent B cells but was induced upon B cell activation. Activation of human B and T lymphocytes also induced surface BLV-binding receptor expression and required de novo protein synthesis. Receptor levels were down-regulated as activated lymphocytes returned to quiescence. In the human thymus, BLV-binding receptor expression was specifically detected on thymocytes responding to the IL-7 cytokine. Thus, expression of the BLV-binding receptor is a marker of enhanced metabolic activity in B cells, T cells, and thymocytes.