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J Schneider-Schaulies

Publications and source records attributed to J Schneider-Schaulies.

At least 19 recordsLinked to original sources
PubMed2006

A mouse model of persistent brain infection with recombinant Measles virus.

Measles virus (MV) nucleocapsids are present abundantly in brain cells of patients with subacute sclerosing panencephalitis (SSPE). This invariably lethal brain disease develops years after acute measles as result of a persistent MV infection. Various rodent models for MV infection of the central nervous system (CNS) have been described in the past, in which the detection of viral antigens is based on histological staining procedures of paraffin embedded brains. Here, the usage of a recombinant MV (MV-EGFP-CAMH) expressing the haemagglutinin (H) of the rodent-adapted MV-strain CAM/RB and the enhanced green fluorescent protein (EGFP) is described. In newborn rodents the virus infects neurons and causes an acute lethal encephalitis. From 2 weeks on, when the immune system of the genetically unmodified animal is maturating, intracerebral (i.c.) infection is overcome subclinically, however, a focal persistent infection in groups of neurons remains. The complete brain can be analysed in 50 or 100 microm slices, and infected autofluorescent cells are readily detected. Seven and 28 days post-infection (p.i.) 86 and 81% of mice are infected, respectively, and virus persists for more than 50 days p.i. Intraperitoneal immunization with MV 1 week before infection, but not after infection, protects and prevents persistence. The high percentage of persistence demonstrates that this is a reliable and useful model of a persistent CNS infection in fully immunocompetent mice, which allows the investigation of determinants of the immune system.

Age Factors↗
PubMed2001

Measles virus interactions with cellular receptors: consequences for viral pathogenesis.

Although CNS complications occurring early and late after acute measles are a serious problem and often fatal, the transient immunosuppression lasting for several weeks after the rash is the major cause of measles-related morbidity and mortality worldwide. This review is focused on the interactions of measles virus (MV) with cellular receptors on neural and lymphoid cells which are important elements in viral pathogenesis. First, the cognate MV receptors, CD46 and CD150, are important components of viral tropism by mediating binding and entry. Second, however, additional unknown cellular surface molecules may (independently of viral uptake) after interaction with the MV glycoprotein complex act as signaling molecules and thereby modulate cellular survival, proliferation, and specific functions.

Antigens, CD↗
PubMed2001

CD150 (SLAM) is a receptor for measles virus but is not involved in viral contact-mediated proliferation inhibition.

Measles virus (MV) interacts with cellular receptors on the surface of peripheral blood lymphocytes (PBL) which mediate virus binding and uptake. Simultaneously, the direct contact of the viral glycoproteins with the cell surface induces a negative signal blocking progression to the S phase of the cell cycle, resulting in a pronounced proliferation inhibition. We selected a monoclonal antibody (MAb 5C6) directed to the surface of highly MV-susceptible B cells (B95a), which inhibits binding to and infection of cells with MV wild-type and vaccine strains. By screening a retroviral cDNA library from human splenocytes (ViraPort; Stratagene) with this antibody, we cloned and identified the recognized molecule as signaling lymphocytic activation molecule (SLAM; CD150), which is identical to the MV receptor recently found by H. Tatsuo et al. (Nature 406:893-897, 2000). After infection of cells, and after surface contact with MV envelope proteins, SLAM is downregulated from the cell surface of activated PBL and cell lines. Although anti-SLAM and/or anti-CD46 antibodies block virus binding, they do not interfere with the contact-mediated proliferation inhibition. In addition, the cell-type-specific expression of SLAM does not correlate with the sensitivity of cells for proliferation inhibition. The data indicate that proliferation inhibition induced by MV contact is independent of the presence or absence of the virus-binding receptors SLAM and CD46.

Animals↗
PubMed2001

Recombinant measles viruses expressing altered hemagglutinin (H) genes: functional separation of mutations determining H antibody escape from neurovirulence.

Measles virus (MV) strain CAM/RB, which was adapted to growth in the brain of newborn rodents, is highly neurovirulent. It has been reported earlier that experimentally selected virus variants escaping from the monoclonal antibodies (MAbs) Nc32 and L77 to hemagglutinin (H) preserved their neurovirulence, whereas mutants escaping MAbs K71 and K29 were found to be strongly attenuated (U. G. Liebert et al., J. Virol. 68:1486-1493, 1994). To investigate the molecular basis of these findings, we have generated a panel of recombinant MVs expressing the H protein from CAM/RB and introduced the amino acid substitutions thought to be responsible for antibody escape and/or neurovirulence. Using these recombinant viruses, we identified the amino acid changes conferring escape from the MAbs L77 (377R-->Q and 378M-->K), Nc32 (388G-->S), K71 (492E-->K and 550S-->P), and K29 (535E-->G). When the corresponding recombinant viruses were tested in brains of newborn rodents, we found that the mutations mediating antibody escape did not confer differential neurovirulence. In contrast, however, replacement of two different amino acids, at positions 195G-->R and 200S-->N, which had been described for the escape mutant set, caused the change in neurovirulence. Thus, antibody escape and neurovirulence appear not to be associated with the same structural alterations of the MV H protein.

Animals↗
PubMed2001

Measles virus induced immunosuppression: targets and effector mechanisms.

A profound, transient suppression of immune functions during and after the acute infection is the major cause of more than one million cases of infant deaths associated with measles worldwide. Concommittant with the generation of an efficient measles virus (MV) specific immunity, immune responses towards other pathogens are strongly impaired and provide the basis for the establishment and severe course of opportunistic infections. The molecular basis for MV-induced immunosuppression has not been resolved as yet. Similar to other immunosuppressive viruses, MV is lymphotropic and viral nucleic acid and proteins are detectable in peripheral blood mononuclear cells (PBMC). It is considered central to MV-induced immunosuppression that PBMC isolated from patients largely fail to proliferate in response to antigen specific and polyclonal stimulation. The low abundancy of MV-infected PBMC suggests that MV-induced immunosuppression is not directly caused by infection-mediated cell loss or fusion, but rather by indirect mechanisms such as deregulation of cytokines or surface contact-mediated signaling which may lead to apoptosis or impair the proliferative response of uninfected PBMC. Evidence for a role of any of these mechanisms was obtained in vitro, however, much has still to be learned about the tropism of MV and its interactions with particular host cells such as dendritic cells in vivo.

Animals↗
PubMed2000

CD46 transgene expression in pig peripheral blood mononuclear cells does not alter their susceptibility to measles virus or their capacity to downregulate endogenous and transgenic CD46.

CD46 (or membrane cofactor protein) protects autologous cells from complement-mediated lysis and has been expressed as a transgene in pigs to overcome complement-mediated hyperacute rejection of porcine organs upon transplantation into primates. Since CD46 has been identified as a receptor for measles virus (MV), the susceptibility of CD46-transgenic (tg) pig peripheral blood mononuclear cells (PBMC) to infection with MV strains which do and do not use CD46 as receptor was investigated. Surprisingly, it was found that MV vaccine strains (e.g. Edmonston) bound to tg as well as non-tg pig PBMC. Phytohaemagglutinin-stimulated CD46-tg and non-tg pig PBMC were equally well infected with MV vaccine strains irrespective of CD46 expression. Upon infection, tg CD46 was downregulated from the cell surface. In contrast, the binding capacity for MV wild-type strains to pig and human PBMC was low, irrespective of CD46 expression. These MV strains did not infect tg or non-tg pig cells. Expression of endogenous pig CD46 was detected with polyclonal sera against human CD46. After infection of pig PBMC with MV strain Edmonston, endogenous pig CD46 was also downregulated. This suggests an interaction between MV Edmonston and pig CD46. However, polyclonal CD46 sera did not inhibit infection with MV Edmonston indicating that CD46 may not exclusively act as a receptor for MV on these cells. Interestingly, similar results were observed using human PBMC. Data suggest that CD46 downregulation after interaction with MV may also occur in porcine organs which express endogenous and/or human CD46 as a means of protection against complement-mediated damage.

Animals↗
PubMed2000

Antibodies to CD9, a tetraspan transmembrane protein, inhibit canine distemper virus-induced cell-cell fusion but not virus-cell fusion.

Canine distemper virus (CDV) causes a life-threatening disease in several carnivores including domestic dogs. Recently, we identified a molecule, CD9, a member of the tetraspan transmembrane protein family, which facilitates, and antibodies to which inhibit, the infection of tissue culture cells with CDV (strain Onderstepoort). Here we describe that an anti-CD9 monoclonal antibody (MAb K41) did not interfere with binding of CDV to cells and uptake of virus. In addition, in single-step growth experiments, MAb K41 did not induce differences in the levels of viral mRNA and proteins. However, the virus release of syncytium-forming strains of CDV, the virus-induced cell-cell fusion in lytically infected cultures, and the cell-cell fusion of uninfected with persistently CDV-infected HeLa cells were strongly inhibited by MAb K41. These data indicate that anti-CD9 antibodies selectively block virus-induced cell-cell fusion, whereas virus-cell fusion is not affected.

Animals↗
PubMed1999

Measles virus spread by cell-cell contacts: uncoupling of contact-mediated receptor (CD46) downregulation from virus uptake.

CD46, which serves as a receptor for measles virus (MV; strain Edmonston), is rapidly downregulated from the cell surface after contact with viral particles or infected cells. We show here that the same two CD46 complement control protein (CCP) domains responsible for primary MV attachment mediate its downregulation. Optimal downregulation efficiency was obtained with CD46 recombinants containing CCP domains 1 and 2, whereas CCP 1, alone and duplicated, induced a slight downregulation. Using persistently infected monocytic/promyelocytic U937 cells which release very small amounts of infectious virus, and uninfected HeLa cells as contact partners, we then showed that during contact the formation of CD46-containing patches and caps precedes CD46 internalization. Nevertheless, neither substances inhibiting capping nor the fusion-inhibiting peptide Z-D-Phe-L-Phe-Gly-OH (FIP) blocked CD46 downregulation. Thus, CD46 downregulation can be uncoupled from fusion and subsequent virus uptake. Interestingly, in that system cell-cell contacts lead to a remarkably efficient infection of the target cells which is only partially inhibited by FIP. The finding that the contact of an infected with uninfected cells results in transfer of infectious viral material without significant (complete) fusion of the donor with the recipient cell suggests that microfusion events and/or FIP-independent mechanisms may mediate the transfer of MV infectivity from cell to cell.

Animals↗
PubMed1999

A recombinant measles vaccine virus expressing wild-type glycoproteins: consequences for viral spread and cell tropism.

Wild-type, lymphotropic strains of measles virus (MV) and tissue culture-adapted MV vaccine strains possess different cell tropisms. This observation has led to attempts to identify the viral receptors and to characterize the functions of the MV glycoproteins. We have functionally analyzed the interactions of MV hemagglutinin (H) and fusion (F) proteins of vaccine (Edmonston) and wild-type (WTF) strains in different combinations in transfected cells. Cell-cell fusion occurs when both Edmonston F and H proteins are expressed in HeLa or Vero cells. The expression of WTF glycoproteins in HeLa cells did not result in syncytia, yet they fused efficiently with cells of lymphocytic origin. To further investigate the role of the MV glycoproteins in virus cell entry and also the role of other viral proteins in cell tropism, we generated recombinant vaccine MVs containing one or both glycoproteins from WTF. These viruses were viable and grew similarly in lymphocytic cells. Recombinant viruses expressing the WTFH protein showed a restricted spread in HeLa cells but spread efficiently in Vero cells. Parental WTF remained restricted in both cell types. Therefore, not only differential receptor usage but also other cell-specific factors are important in determining MV cell tropism.

Animals↗
PubMed1999

The H gene of rodent brain-adapted measles virus confers neurovirulence to the Edmonston vaccine strain.

Molecular determinants of neuropathogenesis have been shown to be present in the hemagglutinin (H) protein of measles virus (MV). An H gene insertion vector has been generated from the Edmonston B vaccine full-length infectious clone of MV. Using this vector, it is possible to insert complete H open reading frames into the parental (Edtag) background. The H gene from a rodent brain-adapted MV strain (CAM/RB) was inserted into this vector, and a recombinant virus (EdtagCAMH) was rescued by using a modified vaccinia virus which expresses T7 RNA polymerase (MVA-T7). The recombinant virus grew at an equivalent rate and to similar titers as the CAM/RB and Edtag parental viruses. Neurovirulence was assayed in a mouse model for MV encephalitis. Viruses were injected intracerebrally into the right cortex of C57/BL/6 suckling mice. After infection mice inoculated with the CAM/RB strain developed hind limb paralysis and ataxia. Clinical symptoms were never observed with an equivalent dose of Edtag virus or in sham infections. Immunohistochemistry (IHC) was used to detect viral antigen in formalin-fixed brain sections. Measles antigen was observed in neurons and neuronal processes of the hippocampus, frontal, temporal, and olfactory cortices and neostriatum on both sides of symmetrical structures. Viral antigen was not detected in mice infected with Edtag virus. Mice infected with the recombinant virus, EdtagCAMH, became clinically ill, and virus was detected by IHC in regions of the brain similar to those in which it was detected in animals infected with CAM/RB. The EdtagCAMH infection had, however, progressed much less than the CAM/RB virus at 4 days postinfection. It therefore appears that additional determinants are encoded in other regions of the MV genome which are required for full neurovirulence equivalent to CAM/RB. Nevertheless, replacement of the H gene alone is sufficient to cause neuropathology.

Adaptation, Biological↗
PubMed1998

Differential receptor usage by measles virus strains.

Recently, we demonstrated that infection of cells with all measles virus (MV) strains tested was inhibited by antibodies against CD46, although not all strains caused downregulation of the MV receptor CD46 from the surface of human cells. We now show that infection of cells with MV strain WTFb, a variant of wild-type isolate WTF which has been isolated and propagated on human BJAB cells, is not inhibited by antibodies against CD46. In contrast, infection of cells with the closely related strain WTFv, a Vero cell-adapted variant of WTF, is inhibited by antibodies against CD46. This observation led us to investigate the interaction of these viruses and the vaccine strain Edmonston (Edm) with CD46 and target cells. Cellular receptors with high affinity binding for WTFb are present on BJAB cells, but not on transfected CD46-expressing CHO cells. In contrast to the Edm strain, virus particles and solubilized envelope glycoproteins of WTFb have a very limited binding capacity to CD46. Furthermore, we show that recombinant soluble CD46 either does not bind, or binds very weakly, to WTFb glycoproteins expressed on the cell surface. Our findings indicate that wild-type MV strain WTFb and vaccine strain Edm use different binding sites on human cells. In addition, the results suggest that MV strains may alternatively use CD46 and an unknown molecule as receptors, and that the degree of usage of both receptors may be MV strain-specific.

Animals↗
PubMed1998

Receptor (CD46)- and replication-mediated interleukin-6 induction by measles virus in human astrocytoma cells.

A major source of inflammatory cytokines in the measles virus (MV)-infected brain are astrocytes, which produce a variety of soluble mediators including interferons-alpha/beta (IFN-alpha/beta), interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF-alpha), and interleukin-6 (IL-6). Using the MV-strain Edmonston (ED) and the recombinant MV-strain MGV in which the MV-envelope proteins H and F have been replaced by the vesicular stomatitis virus (VSV) envelope protein G, we investigated IL-6 induction in human U-251 astrocytoma cells in the presence and absence of a MV-specific receptor (CD46) interaction. The CD46-MV interaction did not inhibit the induction of cytokines. Similar multiplicities of infection of MGV induced generally lower levels of IL-6 than MV-ED. UV-inactivated replication-incompetent MV-ED induced low levels of IL-6. In contrast, MGV did not induce IL-6 after inactivation with UV light, indicating that the MV-ED-receptor interaction or the uptake of viral particles by membrane fusion induced IL-6, whereas interaction with the VSV-G receptor and uptake of viral particles by endocytosis did not induce IL-6. Crosslink of the MV-receptor CD46 with antibodies and treatment of cells with purified viral glycoproteins led to the induction of small but significant amounts of IL-6. Our data suggest that triggering of CD46 and associated protein kinases can lead to the induction of low levels of IL-6, whereas the replication of the negative strand RNA virus constitutes the major stimulus leading to the synthesis of high levels of IL-6 in astrocytes.

Antigens, CD↗
PubMed1997

Mapping of the primary binding site of measles virus to its receptor CD46.

The measles virus (MV) hemagglutinin binds to the complement control protein (CCP) CD46 primarily through the two external modules, CCP-I and -II. To define the residues involved in binding, 40 amino acids predicted to be solvent-exposed on the CCP-I-II module surface were changed to either alanine or serine. Altered proteins were expressed on the cell surface, and their abilities to bind purified MV particles, a soluble form of hemagglutinin (sH) and nine CD46-specific antibodies competing to different levels with sH attachment, were measured. All proteins retained, at least in part, MV and sH binding, but some completely lost binding to certain antibodies. Amino acids essential for binding of antibodies weakly or moderately competing with sH attachment are situated in the membrane-distal tip of CCP-I, whereas residues involved in binding of strongly sH competing antibodies cluster in the center of CCP-I (Arg-25, Asp-27) or in CCP-II (Arg-69, Asp-70). Both clusters face the same side of CCP-I-II and map close to amino acid exchanges impairing sH binding (E11A, R29A, P39A, and D70A) or MV binding (D70A and E84A) and to a six-amino acid loop, previously shown to be necessary for sH binding.

Amino Acid Sequence↗
PubMed1997

Absence of measles virus receptor (CD46) in lesions of subacute sclerosing panencephalitis brains.

In this study we investigated pathological changes of the expression of the measles virus (MV) receptor, CD46, in subacute sclerosing panencephalitis (SSPE) brains. We analyzed CD46 expression in lesions of brain specimens from five SSPE patients in comparison to uninfected regions of the same brains and to normal human brains. The correlation between CD46 and MV infection, in individual cells in SSPE brains, was analyzed by double-staining procedures using monoclonal antibodies (mAbs) and in situ hybridization to detect MV-specific mRNAs. We found that CD46 was expressed at relatively low levels by neurons and astrocytes in normal brains in comparison to neuroblastoma and astrocytoma cell lines. Within heavily infected (MV-positive) brain lesions of all five SSPE cases, CD46 was either not detected or was expressed to a lesser degree by neural cells, irrespective of whether MV antigens were detectable or not. In contrast, normal levels of CD46 were found in SSPE brain tissue distant from the lesion. Using in situ hybridization, mRNAs of both MV nucleocapsid and MV hemagglutinin (MV-H) were detected in all SSPE lesions, while no or only small amounts of MV-H protein were detected. MV-infected neurons were never found to express CD46. Although a strict correlation between levels of the MV-H protein and the absence CD46 could not be seen, these findings suggest that the CD46 expression is reduced by the MV infection in lesions of SSPE brains.

Adolescent↗
PubMed1997

Specific binding of HIV-1 envelope protein gp120 to the structural membrane proteins ezrin and moesin.

The observation that HIV in vitro can infect CD4- and Gal-C-negative brain cell lines has stimulated this study to identify alternative gp120-binding proteins on brain cells. HIV-1 gp120 binding proteins of the CD4-negative and Gal-C-negative, non-productively infectable human glioblastoma cell line D54 were purified by affinity chromatography over a gp120-conjugated sepharose column and identified by peptide microsequencing. The binding capacity and specificity of this column was controlled using extracts of CD4-positive cells. Two of seven prominent proteins eluted from the gp120 affinity column specifically bound gp120 in Western blot overlay experiments and were identified by subsequent immunoblotting and microsequencing as ezrin and moesin, members of the ERM (ezrin, radixin, moesin) family of cellular structural membrane proteins. Antibodies to ezrin and moesin specifically recognized the eluted gp120 binding proteins confirming their identification. Ezrin and moesin are structural proteins binding to the cellular membrane and to several cytoskeletal and transmembrane proteins. Our results suggest that ezrin and moesin might play a role as gp160/gp120 binding proteins during the uptake, the assembly or the budding of HIV.

Chromatography, Affinity↗
PubMed1997

CD9, a tetraspan transmembrane protein, renders cells susceptible to canine distemper virus.

Canine distemper virus (CDV), a lymphotropic and neurotropic negative-stranded RNA virus of the Morbillivirus genus, causes a life-threatening disease in several carnivores, including domestic dogs. To identify the cellular receptor(s) involved in the uptake of CDV by susceptible cells, we isolated a monoclonal antibody (MAb K41) which binds to the cell surface and inhibits the CDV infection of several cell lines from various species. Pretreatment of cells with MAb K41 reduces the number of infectious centers and the size of the syncytia. Using affinity chromatography with MAb K41, we purified from HeLa and Vero cell extracts a 26-kDa protein which contained the amino acid sequence TKDEPQRETLK of human CD9, a member of the tetraspan transmembrane or transmembrane 4 superfamily of cell surface proteins. Transfection of NIH 3T3 or MDBK cells with a CD9 expression plasmid rendered these cells permissive for viral infection and raised virus production by a factor of 10 to 100. The mechanism involved is still unclear, since we were unable to detect direct binding of CDV to CD9 by using immunoprecipitation and a virus overlay protein binding assay. These findings indicate that human CD9 and its homologs in other species are necessary factors for the uptake of CDV by target cells, the formation of syncytia, and the production of progeny virus.

3T3 Cells↗