Search PubMed⌕ Search

Biomedical subjects

K Shimotohno

Publications and source records attributed to K Shimotohno.

At least 181 records · Page 10Linked to original sources
PubMed1990

A structural protein of hepatitis C virus expressed in E. coli facilitates accurate detection of hepatitis C virus.

A putative core protein derived from hepatitis C virus was expressed in E. coli. More than 5% of the total protein expressed in the bacteria after induction by isopropylthio-beta-D-galactoside was shown to be the expected protein. Western blotting with this E. coli lysate proved to be more efficient than ELISA with a non-structural viral protein, C100, to detect infection of hepatitis C virus in the sera of patients with non-A, non-B chronic hepatitis, hepatocellular carcinoma as well as in sera from healthy persons.

Base Sequence↗
PubMed1990

Generation and characterization of monoclonal antibodies against multiple epitopes on the C-terminal half of envelope gp46 of human T-cell leukemia virus type-I (HTLV-I).

In order to study the antigenicity of envelope 46 kDa glycoprotein (gp46) of human T-cell leukemia virus type-I (HTLV-1), we have generated monoclonal anti-gp46 antibodies (MAbs), REY-7, REY-11, REY-16, REY-30, MET-2 and MET-3 from rats and mice. Immunoblot and immunofluorescence assays showed that these MAbs recognize gp46 and its related antigens, and specifically stained HTLV-I-bearing cells. All MAbs reacted with a recombinant gp46 antigen, N147, expressing the 147 amino acids in the C-terminal half of gp46. By using various synthetic peptides corresponding to the gp46 sequence, epitopes recognized by REY-7 and MET-3, REY-11 and REY-16, and REY-30 were mapped to regions corresponding to the amino acids 175-199, 253-282 and 288-312, respectively. MET-2 did not react with any of the peptides used. These results indicate that the present MABs are directed against at least 4 distinct epitopes expressed on the C-terminal half of gp46. The binding of these MAbs to gp46 was specifically inhibited by sera from HTLV-I-infected individuals, but none of these MAbs inhibited the cell fusion activity of HTLV-I.

Animals↗
PubMed1990

Molecular cloning of the human hepatitis C virus genome from Japanese patients with non-A, non-B hepatitis.

The nucleotide sequence of the Japanese type of hepatitis C virus (HCV-J) genome, consisting of 9413 nucleotides, was determined by analyses of cDNA clones from plasma specimens from Japanese patients with chronic hepatitis. HCV-J genome contains a long open reading frame that can encode a sequence of 3010 amino acid residues. Comparison of HCV-J with the American isolate of HCV showed 22.6% difference in nucleotide sequence and 15.1% difference in amino acid sequence. Thus HCV-J and the American isolate of HCV are probably different subtypes of HCV. The relationship of HCV-J with other animal RNA virus families and the putative organization of the HCV-J genome are discussed.

Amino Acid Sequence↗
PubMed1990

Expression of HTLV-I envelope protein fused to hydrophobic amino-terminal peptide of baculovirus polyhedrin in insect cells and its application for serological assays.

The envelope of human T-cell leukemia virus type I (HTLV-I) consists of two glycoproteins gp46 and p20E. Recombinant envelope proteins were produced by using an expression vector derived from insect baculovirus, Bombyx mori nuclear polyhedrosis virus. Polyhedrin fusion proteins C182, N147, and N287 contained whole region p20E, C-terminal half of gp46, and almost whole region gp46, respectively. N147 and N287 were suggested to be processed forms resulting from internal cleavage by cellular enzymes. In cultured cells and the insect larvae, C182 and N147 were produced abundantly enough to be purified to homogeneity; however, N287 was produced poorly and not purified. The purified proteins were recognized by HTLV-I-infected human sera and shown to be highly specific antigens for blood screening systems.

Amino Acid Sequence↗
PubMed1990

A structural protein encoded by the 5' region of the hepatitis C virus genome efficiently detects viral infection.

A structural region of the hepatitis C virus genome was molecularly cloned. A protein expressed in vitro by transcription followed by translation was precipitated immunologically by sera from patients with chronic hepatitis or hepatocellular carcinoma that were positive for antibody against the non-structural protein, C100, of hepatitis C virus, but not by sera from healthy persons. Thus, this structural protein should be useful for detection of infection with this virus.

Amino Acid Sequence↗
PubMed1990

Repressive effect of the nef cDNA of human immunodeficiency virus type 1 on the promoter activity of the viral long terminal repeat.

The nef gene product of human immunodeficiency virus type 1 (HIV-1) has been implicated as a negative factor for viral replication and is suspected to play an important role in the maintenance of viral latency. However, there seems to be evidence both for and against the negative effect of nef gene product. In the present report, we reevaluated the function of the nef gene by means of transient CAT assays with two human T cell lines. In most of the experiments, carefully controlled triplicate studies were carried out. We observed that not only the nef-expression plasmid, but also an effector plasmid containing the nef cDNA sequence in a reverse orientation, not expressing the Nef protein, showed a similar extent of repression of the HIV-1 promoter activity. We also examined the repressive effect of the nef cDNA with deletion mutants of HIV-1 long terminal repeat and heterologous promoters. The results led us to conclude that the apparent "repressor"-like action of the nef cDNA itself could be explained by competition for certain transcription factors required for HIV-1 gene expression by identical sequences also present in the nef cDNA.

Binding, Competitive↗
PubMed1990

lck suppresses gene expression from various promoters including human T-cell leukemia virus type I promoter.

The T-lymphocyte-specific tyrosine kinase gene, lck, is expressed in T-lymphocyte cell lines, except for several human T-cell leukemia virus type I(HTLV-I)-transformed T-lymphocyte cell lines, which produce HTLV-I. By introducing an lck-expression vector, we found that lck product suppresses gene expression from HTLV-I promoter in a transient assay. Moreover, various other promoters of cellular genes or viruses were found to have their transcriptional activity repressed by lck.

Chloramphenicol O-Acetyltransferase↗
PubMed1990

Molecular cloning and characterization of a cDNA for a novel phorbol-12-myristate-13-acetate-responsive gene that is highly expressed in an adult T-cell leukemia cell line.

To identify gene products that might be involved in leukemogenesis of adult T-cell leukemia (ATL), we constructed a cDNA library from an ATL tumor cell line named IKD. By differential plaque hybridization using [32P]cDNAs of poly(A)+ RNA from IKD cells and a human T-lymphotropic virus type I-infected T-cell line (C91/PL) as probes and RNA blot analysis, we obtained a single cDNA clone of a gene that is highly expressed in IKD cells. Expression of this gene was also detected in fresh peripheral blood mononuclear cells of several ATL patients but not in those of healthy donors. Sequence analysis showed that the cDNA was that of a previously undescribed gene. On structural analysis of the cDNA (1,897 base pairs), a short open reading frame encoding a polypeptide of 54 amino acid residues was found. Exposure of human peripheral blood mononuclear cells, a T-cell lymphoma cell line (Jurkat), and quiescent human embryonic lung cells to phorbol-12-myristate-13-acetate resulted in rapid, transient expression of 2.0-kilobase mRNA of this gene. This induction of the gene was not inhibited by an inhibitor of protein synthesis, cycloheximide. From these findings, we suggest that this gene, named APR, is a member of the cellular immediate-early-response genes.

Amino Acid Sequence↗
PubMed1990

Human proviral mRNAs down regulated in choriocarcinoma encode a zinc finger protein related to Krüppel.

RNA transcripts of the HERV-R (ERV3) human provirus that are abundant in placenta but absent in choriocarcinoma contain nonproviral genomic sequences at their 3' ends. We report here the isolation of cDNA clones of these genomic sequences. The transcripts encode a Krüppel-related zinc finger protein consisting of a unique leader region and more than 12 28-amino-acid finger motifs.

Amino Acid Sequence↗
PubMed1990

Sequence diversity of hepatitis C viral genomes.

The nucleotide sequences of cDNAs (275 base-pairs) in the non-structural protein 5 regions of Japanese isolates of hepatitis C virus (HCV-J) from the plasma of 11 patients with non-A, non-B hepatitis and the livers of five patients with hepatocellular carcinoma were analyzed. Approximately 14 to 17% of nucleotide sequences of the HCV-Js examined differed from that of the original isolate in the United States (HCV-US). Furthermore, 2.5 to 11% sequence diversity was found among the HCV-Js. The nucleotide sequences of the HCV-Js showed characteristic common differences from that of HCV-US, although they also showed some random substitutions. Plural HCV-J genomes were found in two of the cDNAs derived from liver specimens, and a deletion of 102 nucleotides was found in the cDNA derived from one plasma specimen. These results suggest that HCV-J is a strain different from the HCV-US and that mutation of the viral genome occurs at as high a frequency as in that of the human immunodeficiency virus.

Amino Acid Sequence↗
PubMed1989

Detection of mRNA for the tax1/rex1 gene of human T-cell leukemia virus type I in fresh peripheral blood mononuclear cells of adult T-cell leukemia patients and viral carriers by using the polymerase chain reaction.

Expression of human T-cell leukemia virus type I (HTLV-I) is not detectable by immunofluorescence analysis or RNA blot analysis in most fresh peripheral blood mononuclear cells of patients with adult T-cell leukemia or of asymptomatic HTLV-I carriers. However, in this work, mRNA for the HTLV-I tax1/rex1 genes was detected in fresh peripheral blood mononuclear cells of adult T-cell leukemia patients and asymptomatic HTLV-I carriers by using reverse transcription followed by the polymerase chain reaction. By using fresh peripheral blood mononuclear cells, the expression of tax1/rex1 mRNA was detected in five of the six adult T-cell leukemia patients and four of the eight HTLV-I carriers examined. The amounts of tax1/rex1 mRNA detected corresponded to approximately 10(5) to 10(6) times less than that in the HTLV-I-infected MT-2 cell line. These results indicate that, in some individuals infected with HTLV-I, the provirus in circulating blood cells is transcribed in vivo. Thus the expression of viral antigens in circulating blood cells in vivo is suggested.

Adult↗