PubMed Health⌕ Search

Biomedical subjects

Yusuke Yanagi

Publications and source records attributed to Yusuke Yanagi.

At least 19 recordsLinked to original sources
PubMed2007

SAP regulation of follicular helper CD4 T cell development and humoral immunity is independent of SLAM and Fyn kinase.

Mutations in SH2D1A resulting in lack of SLAM-associated protein (SAP) expression cause the human genetic immunodeficiency X-linked lymphoproliferative disease. A severe block in germinal center development and lack of long-term humoral immunity is one of the most prominent phenotypes of SAP(-) mice. We show, in this study, that the germinal center block is due to an essential requirement for SAP expression in Ag-specific CD4 T cells to develop appropriate follicular helper T cell functions. It is unknown what signaling molecules are involved in regulation of SAP-dependent CD4 T cell help functions. SAP binds to the cytoplasmic tail of SLAM, and we show that SLAM is expressed on resting and activated CD4 T cells, as well as germinal center B cells. In addition, SAP can recruit Fyn kinase to SLAM. We have now examined the role(s) of the SLAM-SAP-Fyn signaling axis in in vivo CD4 T cell function and germinal center development. We observed normal germinal center development, long-lived plasma cell development, and Ab responses in SLAM(-/-) mice after a viral infection (lymphocytic choriomeningitis virus). In a separate series of experiments, we show that SAP is absolutely required in CD4 T cells to drive germinal center development, and that requirement does not depend on SAP-Fyn interactions, because CD4 T cells expressing SAP R78A are capable of supporting normal germinal center development. Therefore, a distinct SAP signaling pathway regulates follicular helper CD4 T cell differentiation, separate from the SLAM-SAP-Fyn signaling pathway regulating Th1/Th2 differentiation.

Animals↗
PubMed2006

Multiple amino acid substitutions in hemagglutinin are necessary for wild-type measles virus to acquire the ability to use receptor CD46 efficiently.

Measles virus (MV) possesses two envelope glycoproteins, namely, the receptor-binding hemagglutinin (H) and fusion proteins. Wild-type MV strains isolated in B-lymphoid cell lines use signaling lymphocyte activation molecule (SLAM), but not CD46, as a cellular receptor, whereas MV vaccine strains of the Edmonston lineage use both SLAM and CD46 as receptors. Studies have shown that the residue at position 481 of the H protein is critical in determining the use of CD46 as a receptor. However, the wild-type IC-B strain with a single N481Y substitution in the H protein utilizes CD46 rather inefficiently. In this study, a number of chimeric and mutant H proteins, and recombinant viruses harboring them, were generated to determine which residues of the Edmonston H protein are responsible for its efficient use of CD46. Our results show that three substitutions (N390I and E492G plus N416D or T446S), in addition to N481Y, are necessary for the IC-B H protein to use CD46 efficiently as a receptor. The N390I, N416D, and T446S substitutions are present in the H proteins of all strains of the Edmonston lineage, whereas the E492G substitution is found only in the H protein of the Edmonston tag strain generated from cDNAs. The T484N substitution, found in some of the Edmonston-lineage strains, resulted in a similar effect on the use of CD46 to that caused by the E492G substitution. Thus, multiple residues in the H protein that have not previously been implicated have important roles in the interaction with CD46.

Amino Acid Substitution↗
PubMed2006

Measles virus infection of SLAM (CD150) knockin mice reproduces tropism and immunosuppression in human infection.

The human signaling lymphocyte activation molecule (SLAM, also called CD150), a regulator of antigen-driven T-cell responses and macrophage functions, acts as a cellular receptor for measles virus (MV), and its V domain is necessary and sufficient for receptor function. We report here the generation of SLAM knockin mice in which the V domain of mouse SLAM was replaced by that of human SLAM. The chimeric SLAM had an expected distribution and normal function in the knockin mice. Splenocytes from the SLAM knockin mice permitted the in vitro growth of a virulent MV strain but not that of the Edmonston vaccine strain. Unlike in vitro infection, MV could grow only in SLAM knockin mice that also lacked the type I interferon receptor (IFNAR). After intraperitoneal or intranasal inoculation, MV was detected in the spleen and lymph nodes throughout the body but not in the thymus. Notably, the virus appeared first in the mediastinal lymph node after intranasal inoculation. Splenocytes from MV-infected IFNAR(-/-) SLAM knockin mice showed suppression of proliferative responses to concanavalin A. Thus, MV infection of SLAM knockin mice reproduces lymphotropism and immunosuppression in human infection, serving as a useful small animal model for measles.

Animals↗
PubMed2006

Translational inhibition and increased interferon induction in cells infected with C protein-deficient measles virus.

In addition to the phosphoprotein, the P gene of measles virus (MV) also encodes the V and C proteins by an RNA editing process and by alternative initiation of translation in a different reading frame, respectively. Although the MV C protein is required for efficient MV replication in vivo and in some cultured cells, its exact functions in virus infection are currently unclear. Here, we report that a recombinant MV lacking the C protein (MVDeltaC) grew poorly in a human cell line possessing the intact interferon (IFN) pathway and that this growth defect was associated with reduced viral translation and genome replication. The translational inhibition was correlated with phosphorylation of the alpha subunit of eukaryotic translation initiation factor 2. Moreover, increased IFN induction was observed in MVDeltaC-infected cells. The NS1 protein of influenza virus, which binds to double-stranded RNA (dsRNA) and consequently inhibits IFN induction and dsRNA-dependent protein kinase activation, complemented the growth defect of MVDeltaC. These results indicate that the MV C protein inhibits IFN induction and modulates host antiviral responses, thereby ensuring MV growth in host cells.

Animals↗
PubMed2006

Rescue system for measles virus from cloned cDNA driven by vaccinia virus Lister vaccine strain.

A rescue system for measles virus from cloned cDNA was established using CHO/hSLAM cells (Chinese hamster ovary cells expressing a measles virus receptor, signaling lymphocyte activation molecule), LO-T7-1 virus (the Lister vaccine strain of vaccinia virus expressing the T7 RNA polymerase under the control of the early/late p7.5 promoter), and caspase inhibitor. LO-T7-1 drove efficiently the T7 promoter in CHO/hSLAM cells. Rescue efficiency with LO-T7-1 was not as high as that with the vTF7-3 strain based on a neurovirulent vaccinia virus, but was increased by using a caspase inhibitor to block apoptosis of CHO/hSLAM cells induced by LO-T7-1. These modifications resulted in a measles virus rescue efficiency that was even higher than that of previous systems. This safer and more efficient system will facilitate further the genetic manipulation of measles virus in basic research and virus vector development.

Animals↗
PubMed2006

Recombinant wild-type measles virus containing a single N481Y substitution in its haemagglutinin cannot use receptor CD46 as efficiently as that having the haemagglutinin of the Edmonston laboratory strain.

Signalling lymphocyte activation molecule (SLAM) acts as a cellular receptor for Measles virus (MV). The recombinant MV, based on a SLAM-using clinical isolate in which asparagine at position 481 of the haemagglutinin was replaced with tyrosine, was generated. Characterization of this recombinant virus revealed that the N481Y substitution in the haemagglutinin allowed it to utilize CD46 as an alternative receptor, but that its ability to use CD46 was rather low in CD46+ SLAM- cell lines compared with that of the recombinant virus possessing the haemagglutinin of the Edmonston laboratory strain. Thus, an N481Y substitution alone may not be sufficient to make SLAM-using MVs use CD46 efficiently, suggesting that further substitutions in the haemagglutinin are required for them to grow efficiently in CD46+ cells like the Edmonston strain. This may be a reason why few CD46-using MVs are detected in vivo.

Adaptation, Physiological↗
PubMed2006

Measles virus: cellular receptors, tropism and pathogenesis.

Measles virus (MV), a member of the genus Morbillivirus in the family Paramyxoviridae, is an enveloped virus with a non-segmented, negative-strand RNA genome. It has two envelope glycoproteins, the haemagglutinin (H) and fusion proteins, which are responsible for attachment and membrane fusion, respectively. Human signalling lymphocyte activation molecule (SLAM; also called CD150), a membrane glycoprotein of the immunoglobulin superfamily, acts as a cellular receptor for MV. SLAM is expressed on immature thymocytes, activated lymphocytes, macrophages and dendritic cells and regulates production of interleukin (IL)-4 and IL-13 by CD4+ T cells, as well as production of IL-12, tumour necrosis factor alpha and nitric oxide by macrophages. The distribution of SLAM is in accord with the lymphotropism and immunosuppressive nature of MV. Canine distemper virus and Rinderpest virus, other members of the genus Morbillivirus, also use canine and bovine SLAM as receptors, respectively. Laboratory-adapted MV strains may use the ubiquitously expressed CD46, a complement-regulatory molecule, as an alternative receptor through amino acid substitutions in the H protein. Furthermore, MV can infect SLAM- cells, albeit inefficiently, via the SLAM- and CD46-independent pathway, which may account for MV infection of epithelial, endothelial and neuronal cells in vivo. MV infection, however, is not determined entirely by the H protein-receptor interaction, and other MV proteins can also contribute to its efficient growth by facilitating virus replication at post-entry steps. Identification of SLAM as the principal receptor for MV has provided us with an important clue for better understanding of MV tropism and pathogenesis.

Humans↗
PubMed2006

Generation of measles virus with a segmented RNA genome.

Viruses classified in the order Mononegavirales have a single nonsegmented RNA molecule as the genome and employ similar strategies for genome replication and gene expression. Infectious particles of Measles virus (MeV), a member of the family Paramyxoviridae in the order Mononegavirales, with two or three RNA genome segments (2 seg- or 3 seg-MeV) were generated using a highly efficient reverse genetics system. All RNA segments of the viruses were designed to have authentic 3' and 5' self-complementary termini, similar to those of negative-stranded RNA viruses that intrinsically have multiple RNA genome segments. The 2 seg- and 3 seg-MeV were viable and replicated well in cultured cells. 3 seg-MeV could accommodate up to six additional transcriptional units, five of which were shown to be capable of expressing foreign proteins efficiently. These data indicate that the MeV genome can be segmented, providing an experimental insight into the divergence of the negative-stranded RNA viruses with nonsegmented or segmented RNA genomes. They also illustrate a new strategy to develop mononegavirus-derived vectors harboring multiple additional transcriptional units.

Animals↗
PubMed2006

[Cellular tropism and adaptation of the measles virus].

Measles virus (MV) is a member of the genus Morbillivirus in the family Paramyxoviridae. Clinical isolates of MV use signaling lymphocyte activating molecule (SLAM) as a cellular receptor. SLAM is mainly expressed on immune cells such as immature thymocytes, activated lymphocytes and mature dendritic cells. This distribution of SLAM can account for the lymphotropism of MV. On the other hand, laboratory strains of MV use CD46 as an alternative receptor, through amino acid change(s) in the receptor binding hemagglutinin protein. Recently, several reports imply the existence of the cellular receptor(s) other than SLAM and CD46. In this review, we discuss the receptor usage of MV and its adaptation to cultured cells.

Adaptation, Biological↗
PubMed2006

Measles virus receptors and tropism.

Measles virus (MV) is a member of the Morbillivirus genus in the Paramyxoviridae family. Human signaling lymphocyte activation molecule (SLAM) acts as a cellular receptor for MV. SLAM is expressed on immature thymocytes, activated lymphocytes, macrophages and mature dendritic cells. This distribution of SLAM is in accord with lymphotropism and immunosuppressive nature of MV. Canine distemper and rinderpest viruses, other members of the Morbillivirus genus, also use SLAM as receptors. Laboratory-adapted MV strains often use ubiquitously expressed CD46 as an alternative receptor through the amino acid change(s) in the receptor-binding hemagglutinin. Furthermore, MV can infect various cultured cells, albeit with very low efficiency, via SLAM- and CD46-independent pathway, which may also account for MV infection of SLAM(-) cells in vivo.

Animals↗
PubMed2005

Induction of broadly neutralizing antibodies against measles virus mutants using a polyepitope vaccine strategy.

Chimeric molecules expressing multiple copies of the loop-forming hemagglutinin noose epitope (designated as "L"; aa386-400), a protective B cell epitope of the measles virus were generated by recombinant technology. The recombinant polyepitope [L4T4]2 combining two sets of four repeats of the L epitope and with two sets of four repeats of the human promiscuous T cell epitope of tetanus toxoid ("T", tt830-844) was produced in transgenic carrot plants. After intraperitoneal immunization of mice with plant membrane extract, sera neutralized all wild-type viruses. In a modified plaque reduction neutralization assay based on CD150-transfected Vero cells anti-[L4T4]2 sera neutralized all field isolates, irrespective of mutations in the L epitope. Even viruses with a mutation in the contact residues of a neutralizing L-specific monoclonal antibody or two mutations in other positions of the epitope were equally sensitive to neutralization. These results suggest that the multiple copies of the L epitope fold into different conformations that induce a repertoire of B cells diverse enough to overcome the genetic diversity of field viruses.

Amino Acid Sequence↗
PubMed2005

Efficient rescue of measles virus from cloned cDNA using SLAM-expressing Chinese hamster ovary cells.

We here report a highly efficient reverse genetics system for measles virus (MeV), using Chinese hamster ovary cells constitutively expressing a MeV receptor human signaling lymphocyte activation molecule (CHO/hSLAM cells). The recombinant vaccinia virus vTF7-3 that encodes the T7 RNA polymerase under the control of the early/late promoter was used in the system. Replication of vTF7-3 was highly restricted in CHO/hSLAM cells, but the virus could still drive the T7 promoter, allowing us to recover MeV from the transfected cDNA efficiently. With this system the number of infectious centers, in which MeV replication cycles are initiated from transfected cDNAs, was approximately 100 times higher than that with the previous system (. J. Virol. 74, 6643-6647), and the recovery rate was 100%. The wild-type MeV that encodes the lac-Z gene of approximately 3.2kb in length, was easily generated with this CHO/hSLAM system, while such virus could not be recovered with the previous system. Since SLAM acts as a cellular receptor for both MeV vaccine and wild-type strains, the Edmonston vaccine strain was also recovered with this system more efficiently than with any other systems reported previously. Thus, the CHO/hSLAM-based system would expand applications of the MeV reverse genetics by allowing productions of mutant MeVs that have been difficult to generate with less efficient systems.

Animals↗
PubMed2005

Long untranslated regions of the measles virus M and F genes control virus replication and cytopathogenicity.

Measles is still a major cause of mortality mainly in developing countries. The causative agent, measles virus (MeV), is an enveloped virus having a nonsegmented negative-sense RNA genome, and belongs to the genus Morbillivirus of the family Paramyxoviridae. One feature of the moribillivirus genomes is that the M and F genes have long untranslated regions (UTRs). The M and F mRNAs of MeV have 426-nucleotide-long 3' and 583-nucleotide-long 5' UTRs, respectively. Though these long UTRs occupy as much as approximately 6.4% of the virus genome, their function remains unknown. To elucidate the role of the long UTRs in the context of virus infection, we used the reverse genetics based on the virulent strain of MeV, and generated a series of recombinant viruses having alterations or deletions in the long UTRs. Our results showed that these long UTRs per se were not essential for MeV replication, but that they regulated MeV replication and cytopathogenicity by modulating the productions of the M and F proteins. The long 3' UTR of the M mRNA was shown to have the ability to increase the M protein production, promoting virus replication. On the other hand, the long 5' UTR of the F mRNA was found to possess the capacity to decrease the F protein production, inhibiting virus replication and yet greatly reducing cytopathogenicity. We speculate that the reduction in cytopathogenicity may be advantageous for MeV fitness and survival in nature.

Animals↗
PubMed2005

Contributions of matrix and large protein genes of the measles virus edmonston strain to growth in cultured cells as revealed by recombinant viruses.

The Edmonston strain of measles virus (MV) was obtained by sequential passages of the original isolate in various cultured cells. Although attenuated in vivo, it grows efficiently in most primate cell lines. Previous studies have revealed that MV tropism cannot be solely explained by the use of CD150 and/or CD46 as a cellular receptor. In order to evaluate the contributions of individual genes of the Edmonston strain to growth in cultured cells, we generated a series of recombinant viruses in which part of the genome of the clinical isolate IC-B (which uses CD150 as a receptor) was replaced with the corresponding sequences of the Edmonston strain. The recombinant virus possessing the Edmonston hemagglutinin (H) gene (encoding the receptor-binding protein) grew as efficiently in Vero cells as the Edmonston strain. Those viruses having either the matrix (M) or large (L) protein gene from the Edmonston strain could also replicate well in Vero cells, although they entered them at low efficiencies. P64S and E89K substitutions were responsible for the ability of the M protein to make virus grow efficiently in Vero cells, while the first half of the Edmonston L gene was important for better replication. Despite efficient growth in Vero cells, the recombinant viruses with these mutations had growth disadvantage in CD150-positive lymphoid B95a cells. Thus, not only the H gene but also the M and L genes contribute to efficient replication of the Edmonston strain in some cultured cells.

Animals↗
PubMed2004

Genetic evidence linking SAP, the X-linked lymphoproliferative gene product, to Src-related kinase FynT in T(H)2 cytokine regulation.

SAP is an adaptor mutated in X-linked lymphoproliferative disease. It plays a critical role in T helper 2 (T(H)2) cytokine production. This function was suggested to reflect the capacity of SAP to associate with SLAM family receptors and enable tyrosine phosphorylation signaling by these receptors through SAP-mediated recruitment of Src-related kinase FynT. Here, we addressed by genetic means the importance of the SAP-FynT interaction in normal T cell functions. By creating a mouse in which the FynT binding site of SAP was inactivated in the germ line (sap(R78A) mouse) and by analyzing mice lacking SAP, FynT or SLAM, evidence was obtained that the SAP-FynT cascade is indeed crucial for normal T(H)2 functions in vitro and in vivo. These data imply that SAP is necessary for T(H)2 cytokine regulation primarily as a result of its capacity to recruit FynT. They also establish a previously unappreciated role for FynT in SAP-dependent T(H)2 cytokine regulation.

Animals↗
PubMed2004

Absence of tumour necrosis factor facilitates primary and recurrent herpes simplex virus-1 infections.

Tumour necrosis factor (TNF) is an important cytokine in the innate immune response against various infections, including herpes simplex virus (HSV) infection. It has recently become a molecular target of anti-cytokine treatment in certain inflammatory diseases. TNF depletion resulted in a more rapid emergence of infectious HSV-1 in the explant cultures of latently infected trigeminal ganglia (TG), compared with controls. To further evaluate the importance of TNF in the host's defence responses against HSV-1, TNF-knockout mice were challenged via scarified cornea. These mice were more susceptible to primary acute corneal HSV-1 infection than controls, as manifested by an increased mortality rate and higher infectious virus titres in the eyes and TG, indicating that TNF is critical for defence during acute HSV infection. These results imply that the administration of anti-inflammatory TNF antagonists might facilitate the propagation of infectious HSV, resulting in an exacerbation of primary and recurrent acute lesions.

Animals↗
PubMed2004

Cell tropism of wild-type measles virus is affected by amino acid substitutions in the P, V and M proteins, or by a truncation in the C protein.

Two nucleotide differences in the P/C/V and M genes between B95a cell- and Vero cell-isolated wild-type measles viruses (MV) have previously been found from the same patient. The nucleotide difference in the P/C/V gene resulted in an amino acid difference (M175I) in the P and V proteins and a 19 aa deletion in the C protein. The nucleotide difference in the M gene resulted in an amino acid difference (P64H) in the M protein. To verify this result and to examine further whether the amino acid difference or truncation is important for MV cell tropism, recombinant MV strains containing one of the two nucleotide substitutions, or both, were generated. It was found that the P64H substitution in the M protein was important for efficient virus growth and dissemination in Vero cells and that the M175I substitution in the P and V protein or truncation of the C protein was required for optimal growth.

Amino Acid Substitution↗