Biomedical subjects

K Saigo

Publications and source records attributed to K Saigo.

At least 91 records · Page 5Linked to original sources

PubMed1994

Induction of a mirror-image duplication of anterior wing structures by localized hedgehog expression in the anterior compartment of Drosophila melanogaster wing imaginal discs.

The segment polarity gene hedgehog (hh) encodes a secretory protein involved in cell-cell communication in Drosophila melanogaster. The hh gene is expressed in the posterior compartment and is essential for the establishment and maintenance of the anterior/posterior-compartment boundary of each embryonic parasegment [Ingham, P.W., Nature 366 (1993) 560-562]. To clarify possible hh functions in adult appendage formation, we isolated a fly line (h9D) associated with a wing malformation from among fly lines with an hh transgene whose expression is under the control of trapped enhancers. In h9D flies, the ectopic expression of hh occurred in the anterior edge of wing pouch in the wing disc. This abnormal hh expression resulted in not only a mirror-image duplication and ectopic outgrowth in the anterior wing compartment, but also the ectopic expression of patched and decapentaplegic, strongly suggesting that the hh product serves as a morphogen or an inducer essential for wing development, including the proximal/distal axis formation.

Animals↗

PubMed1994

Calpain localization changes in coordination with actin-related cytoskeletal changes during early embryonic development of Drosophila.

Calpain, a calcium-dependent intracellular protease, was identified in Drosophila melanogaster. Drosophila calpain has an amino acid sequence highly homologous to those of mammalian calpains and exhibits a distinct domain structure consisting of cysteine protease and calcium-binding domains. Specific antibodies raised against a recombinant calpain fragment were used to identify the localization of calpain in developing Drosophila embryos. Calpain was first detected near the anterior pole and in posterior region of the embryo just after fertilization. The anterior calpain disappeared during the cleavage cycles. On the other hand, the posterior calpain moved to the posterior pole when polar buds were formed, and condensed just below the pole cells. At cleavage cycles 8 and 9, when nuclei reached the egg surface, calpain was localized between the nuclei at the surface beneath the precleavage furrows. Co-staining experiments with anti-actin antibody revealed that calpain condenses specifically at the edge of and between actin caps that underlie the plasma membrane immediately above each nucleus. These results indicate that calpain is involved in the dynamic changes in the embryonic cytoskeleton, especially actin-related structures, during early embryogenesis prior to cellularization.

Actins↗

PubMed1994

Drosophila phospholipase C-gamma expressed predominantly in blastoderm cells at cellularization and in endodermal cells during later embryonic stages.

A Drosophila gene encoding a gamma-type isozyme of phosphoinositide-specific phospholipase C (PLC) was isolated and characterized. The gene, termed plc-gamma d, was mapped at position 14B-C of the X chromosome. The encoded protein, termed PLC-gamma D, contains X and Y regions, common to all known PLC isozymes. The two regions are split by a Z region that comprises two src homology 2 and one src homology 3 domains and is characteristic of gamma-type mammalian PLC (PLC-gamma 1 and -gamma 2). The deduced amino acid sequence of PLC-gamma D shows overall similarity to mammalian PLC-gamma s; no large deletion was observed except the short C-terminal extended region. In particular, the two split catalytic domains (X and Y regions) and the regulatory Z region including the src homology 2 and src homology 3 domains are well conserved. The mRNA is expressed throughout development, but expression is relatively higher during the embryonic stage, suggesting fundamental and important roles in both cell proliferation and differentiation. Distribution of the mRNA during embryogenesis, as analyzed by whole amount in situ hybridization, revealed that the mRNA emerges and reaches maximum levels at the cellular blastoderm stage and then decreases rapidly to a lower level. In later embryonic stages, invaginated anterior and posterior midgut primordia show high levels of mRNA expression, and fused midgut also maintains a high level of expression. In other tissues and cells, the mRNA was detected at lower levels. These results indicate that Drosophila PLC-gamma may be involved in universal cellular processes mediated possibly by receptor tyrosine kinases during embryogenesis and may also play specific roles during cellularization and midgut differentiation.

Amino Acid Sequence↗

PubMed1994

Platelet release reaction during EDTA-induced platelet agglutinations and inhibition of EDTA-induced platelet agglutination by anti-glycoprotein II b/III a complex monoclonal antibody.

To characterize the nature of EDTA-induced platelet agglutination, the spontaneous release of beta-thromboglobulin (beta-TG) and platelet factor 4 (PF4) was examined during EDTA-induced platelet agglutinations. A slight release of beta-TG and PF4 was observed when EDTA-anticoagulated whole blood from cases with EDTA-induced platelet agglutination was kept for 60 minutes, whereas a high spontaneous release of these proteins was found from normal blood anticoagulated with EDTA. These findings imply that EDTA-dependent platelet agglutinin may stabilize the platelet membrane surfaces. Secondly, we found that pretreatment of fresh blood with anti-glycoprotein (GP) II b/III a complex monoclonal antibody dramatically reduced EDTA-induced platelet agglutinations. This study indicated that the binding sites of EDTA-dependent antibody might be GP II b/III a complex. The use of an anti-GP II b/III a complex monoclonal antibody may be useful in avoiding analytical errors in some cases with EDTA-induced pseudothrombocytopenia.

Adenosine Diphosphate↗

PubMed1994

Extramedullary tumor as presentation of leukemia: establishment of a new human GPIIb- and GPIIIa-positive leukemia cell line.

A 25-year-old man noted swelling of the right cervical lymph nodes in October 1983. Diagnosis of malignant lymphoma was made on the basis of pathological examination of biopsies. Despite both chemotherapy and irradiation treatment, blast cells appeared in the peripheral blood and bone marrow in April 1984. Immunophenotypic analysis demonstrated that the blasts in the patient's peripheral blood expressed CD13, CD33, CD41a, and no markers for T or B lymphocytes, suggesting that he had been suffering from megakaryocytic sarcoma. We established a new cell line derived from the blasts in the peripheral blood, designated KH184. KH184 cells expressed glycoprotein (GP) Ib (CD42b) and GPIIb/IIIa (CD41a), while platelet peroxidase (PPO) activity was negative in an ultrastructural study. Both Northern blot and flow cytometric analysis of surface antigens and DNA content revealed that treatment with 12-O-tetradecanoylphorbol 13-acetate (TPA) did not induce the maturation of these cells. Various cytokines such as interleukin 3 (IL-3), interleukin 6 (IL-6), and leukemia inhibitory factor (LIF) had no effect in promoting the growth of KH184 cells. KH184 cells expressing CD41a seem to possess unusual characteristics. KH184 cells, human GPIIb- and GPIIIa-positive leukemia cells, which lack response to TPA-induced differentiation, provide a new and unique model for the characterization of factors that are implicated in the terminal differentiation of megakaryocytes, and should aid in studies of the mechanism underlying the occurrence of megakaryocytic sarcoma.

Adult↗

PubMed1994

A Drosophila homolog of human proto-oncogene ret transiently expressed in embryonic neuronal precursor cells including neuroblasts and CNS cells.

We have identified a Drosophila gene encoding a putative receptor tyrosine kinase by screening a genomic DNA library with a DNA probe for a Drosophila homolog of fibroblast growth factor receptors. The newly isolated gene codes for a transmembrane protein most similar in sequence to a mammalian proto-oncogene ret; thus, the gene was termed Dret. Dret mRNA is transcribed in very small amounts in the embryonic, larval, and pupal stages. Whole mount in situ hybridization experiments revealed that the mRNA is transiently expressed in neuroblasts in early embryos. In late embryos, Dret mRNA was detected in subpopulations of differentiating CNS and PNS cells. In addition, Dret expression was affected in neurogenic mutants. These results suggest that Dret can be considered as a functional homolog of mammalian ret and should play important roles in neurogenesis.

Amino Acid Sequence↗

PubMed1994

Identification of a novel Drosophila gene encoding a Cdc2-related protein kinase.

We have identified a novel gene encoding a putative protein kinase from a Drosophila genomic library. The gene, about 2 kbp in length, consists of four exons and codes for a protein of 349 amino acid residues. The deduced sequence shows significant similarity to various kinases, especially to a subgroup of Ser/Thr kinases related to Cdc2 kinase; thus, the gene was termed Dcdrk (Drosophila cdc2-related kinase gene). Among the kinases examined, mammalian galactosyltransferase-associated 58 kDa protein kinase showed the highest homology (about 50% identity in the kinase domain) to Dcdrk kinase. Northern blot analysis revealed that the Dcdrk mRNA is expressed throughout development in nearly constant amounts. Moreover, a whole mount in situ hybridization experiment showed that the Dcdrk mRNA is ubiquitously distributed in almost all embryonic cells and tissues, suggesting a universal function of Dcdrk, possibly in cell cycle regulation.

Amino Acid Sequence↗

PubMed1994

Identification of the gene encoding the major NAD(P)H-flavin oxidoreductase of the bioluminescent bacterium Vibrio fischeri ATCC 7744.

The gene encoding the major NAD(P)H-flavin oxidoreductase (flavin reductase) of the luminous bacterium Vibrio fischeri ATCC 7744 was isolated by using synthetic oligonucleotide probes corresponding to the N-terminal amino acid sequence of the enzyme. Nucleotide sequence analysis suggested that the major flavin reductase of V. fischeri consisted of 218 amino acids and had a calculated molecular weight of 24,562. Cloned flavin reductase expressed in Escherichia coli was purified virtually to homogeneity, and its basic biochemical properties were examined. As in the major flavin reductase in crude extracts of V. fischeri, cloned flavin reductase showed broad substrate specificity and served well as a catalyst to supply reduced flavin mononucleotide (FMNH2) to the bioluminescence reaction. The major flavin reductase of V. fischeri not only showed significant similarity in amino acid sequence to oxygen-insensitive NAD(P)H nitroreductases of Salmonella typhimurium, Enterobacter cloacae, and E. coli but also was associated with a low level of nitroreductase activity. The major flavin reductase of V. fischeri and the nitroreductases of members of the family Enterobacteriaceae would thus appear closely related in evolution and form a novel protein family.

Amino Acid Sequence↗

PubMed1994

Identification of the genes encoding NAD(P)H-flavin oxidoreductases that are similar in sequence to Escherichia coli Fre in four species of luminous bacteria: Photorhabdus luminescens, Vibrio fischeri, Vibrio harveyi, and Vibrio orientalis.

Genes encoding NAD(P)H-flavin oxidoreductases (flavin reductases) similar in both size and sequence to Fre, the most abundant flavin reductase in Escherichia coli, were identified in four species of luminous bacteria, Photorhabdus luminescens (ATCC 29999), Vibrio fischeri (ATCC 7744), Vibrio harveyi (ATCC 33843), and Vibrio orientalis (ATCC 33934). Nucleotide sequence analysis showed Fre-like flavin reductases in P. luminescens and V. fischeri to consist of 233 and 236 amino acids, respectively. As in E. coli Fre, Fre-like enzymes in luminous bacteria preferably used riboflavin as an electron acceptor when NADPH was used as an electron donor. These enzymes also were good suppliers of reduced flavin mononucleotide (FMNH2) to the bioluminescence reaction. In V. fischeri, the Fre-like enzyme is a minor flavin reductase representing < 10% of the total FMN reductase. That the V. fischeri Fre-like enzyme has no appreciable homology in amino acid sequence to the major flavin reductase in V. fischeri, FRase I, indicates that at least two different types of flavin reductases supply FMNH2 to the luminescence system in V. fischeri. Although Fre-like flavin reductases are highly similar in sequence to luxG gene products (LuxGs), Fre-like flavin reductases and LuxGs appear to constitute two separate groups of flavin-associated proteins.

Amino Acid Sequence↗

PubMed1994

Diagnosis of post-transfusion graft-versus-host disease after formalin-fixation.

A 72-year-old woman with multiple recurrence of gallbladder cancer was treated by intrahepatic-arterial infusion of doxorubicin using an extracorporeal system of direct hemoperfusion with venovenous bypass. During this treatment, the patient received 600 ml of fresh whole blood and 30 units of platelet concentrate from five unrelated donors. Thereafter, high fever, skin rash over the whole body, and watery diarrhea developed, followed by leukopenia progressing to a fatal sepsis. Post-transfusion graft-versus-host disease (PT-GVHD) was suspected by the clinical manifestations and postmortem pathologic findings. To establish the diagnosis of PT-GVHD, polymerase chain reaction (PCR) amplification of DNA polymorphism associated with length variation in dinucleotide or trinucleotide microsatellite repeats at the loci of D6S89, int-2 protooncogene, and human growth factor with each of the different primer sets was performed using DNA from blood drawn from the patient with clinically established PT-GVHD of a donor origin and formalin-fixed pancreas of recipient origin. Genetic analysis revealed the changes in the patient's lymphocytes from that of the patient to that of donor origin. The present finding that formalin-fixed tissues can be used as a material of patient origin may contribute to accurate diagnosis of PT-GVHD after autopsy.

Aged↗

PubMed1993

Molecular cloning of cDNAs for two Xenopus proteasome subunits and their expression in adult tissues.

Proteasome, a large protein complex with ATP-dependent protease activities, is composed of non-identical but closely related multi-subunits. Using cDNAs for rat proteasome subunits as probes, we obtained three cDNA clones for two Xenopus proteasome subunits from ovary cDNA library. The primary structures of the three cDNAs showed high homology to the corresponding proteasome subunits of other mammalian species (above 90%) and also considerable homology to those of Drosophila and yeast. These results indicate that the sequences of proteasome subunits are well conserved during evolution. Northern blot hybridization revealed that RNAs for the newly isolated subunits (XC8 and XC9) and the previously isolated subunit (XC3) occur at very high levels in testis and ovary, at moderately high levels in lung, skin kidney and spleen, and at low levels in liver, stomach and muscle. It was also shown that relative amounts of the mRNAs for the three subunits are similar in all the adult tissues examined. From these results, we concluded that the expression of the genes for the three subunits (XC3 XC8 and XC9-1) takes place in a roughly coordinated manner in different adult tissues.

Amino Acid Sequence↗

PubMed1993

Distinct expression of two Drosophila homologs of fibroblast growth factor receptors in imaginal discs.

The expression of two Drosophila homologs of FGF receptors (DRF1 and DRF2) in imaginal discs was studied. DFR1 mRNA was observed in several imaginal discs, whereas DFR2 mRNA was not detected. DFR1 expression in the wing and leg discs took place in probable myoblasts in a pattern similar to that of twist, a mesodermal gene. The mRNA was also detected in the morphogenetic furrow and its posterior region of the eye disc and around the proliferation center of the brain. These results suggest that DFR1 is involved in the development of mesodermal and neuronal cells constituting the adult body.

Animals↗

PubMed1993

Chimeric reverse transcriptase of Moloney murine leukaemia virus, having the YXDD box of a Drosophila retrotransposon, 17.6.

All reverse transcriptases (RTs) so far examined are known to possess in common a stretch of 7 amino acids called the YXDD box. Various mutations were introduced in vitro into the nucleotide sequence coding for this box of Moloney murine leukaemia virus (Mo-MuLV) RT. The YXDD box of Mo-MuLV could be replaced by the counterpart of a Drosophila retrotransposon, 17.6, without loss of activity, while virtually complete loss of activity was detected in the case of replacement of the YXDD box of Rous sarcoma virus RT. Amino-acid substitution at position X resulted in significant change in divalent metal ion preference and enzyme activity, thus indicating that the YXDD box possibly constitutes a part of the active center of RT through metal ion binding.

Amino Acid Sequence↗

PubMed1993

Structure and expression of hedgehog, a Drosophila segment-polarity gene required for cell-cell communication.

The complete nucleotide sequence of the coding region of hedgehog (hh), a segment-polarity gene in Drosophila melanogaster, was determined. The gene was found to include three exons which would encode a 421- (or 471-) amino acid (aa) polypeptide with a long hydrophobic stretch. The hh mRNA was about 2.3 kb long and expressed throughout development. The hh expression in an embryo occurred in stripes, while that in imaginal discs occurred in the posterior compartment. As a whole, the spatial expression pattern of hh mRNA was very similar to that of engrailed (en), a homeobox gene required for the formation of the anterior-posterior compartment boundary. Unlike en, no hh expression was observed in the central nervous system.

Animals↗

PubMed1993

The Drosophila forked gene encodes two major RNAs, which, in gypsy or springer insertion mutants, are partially or completely truncated within the 5'-LTR of the inserted retrotransposon.

Mutations in the forked (f) gene of Drosophila cause deformation of bristles and hairs. Our molecular analysis showed the f gene to span more than 30 kb, and to encode two major RNAs, 6.0 and 2.5 kb long, both of which are prematurely terminated in gypsy and springer insertion mutants. These truncated RNAs were polyadenylated using putative polyadenylation signals within the 5'-LTR of the inserted retrotransposon. No evidence was found for effects of the retrotransposon insertions on the promoters for transcription of the 6.0 and 2.5 kb RNAs. In f1 and fx, a single gypsy element was found to be inserted at identical sites in the second intron of region encoding the 2.5 kb f RNA and both truncated and wild-type sized RNAs were detected. Recessive mutations at suppressor of forked (su(f)) increased the fraction of wild-type sized RNAs considerably, suggesting that the wild-type su(f) product either stimulates premature termination at the gypsy LTR or inhibits normal splicing. In f36a, a springer element inserted in the third exon of the region encoding the 2.5 kb f RNA completely suppressed the formation of apparently wild-type transcripts.

Amino Acid Sequence↗

PubMed1993

Mechanism of induction of Bar-like eye malformation by transient overexpression of Bar homeobox genes in Drosophila melanogaster.

The Bar locus of Drosophila is known to be a small complex consisting of two similar homeobox genes, BarH1 and BarH2. Using egr as an ommatidium marker, possible mechanisms of formation of malformed eyes were examined. As in the case of BarH1, overexpression of BarH2 was found to be capable of inducing Bar-like eye malformation. It was suggested that suppression of the anterior progression of the morphogenetic furrow and inhibition of reinitiation of normal ommatidial differentiation were mandatory to formation of the reduced eye morphology in Bar mutants.

Animals↗

PubMed1993

Two FGF-receptor homologues of Drosophila: one is expressed in mesodermal primordium in early embryos.

The fibroblast growth factor (FGF)/receptor system is thought to mediate various developmental events in vertebrates. We examined molecular structures and expression of DFR1 and DFR2, two Drosophila genes closely related to vertebrate FGF-receptor genes. DFR1 and DFR2 proteins contain two and five immunoglobulin-like domains, respectively, in the extracellular region, and a split tyrosine kinase domain in the intracellular region. In early embryos, DFR1 RNA expression, requiring both twist and snail proteins, is specific to mesodermal primordium and invaginated mesodermal cells. At later stages, putative muscle precursor cells and cells in the central nervous system (CNS) express DFR1. DFR2 expression occurs in endodermal precursor cells, CNS midline cells and certain ectodermal cells such as those of trachea and salivary duct. FGF-receptor homologues in Drosophila would thus appear essential for generation of mesodermal and endodermal layers, invaginations of various types of cells, and CNS formation.

Amino Acid Sequence↗

PubMed1993

[Histidine-rich glycoprotein in diabetics].

Histidine-rich glycoprotein (HRGP) has been believed to have anti-fibrinolytic activity through its binding to plasminogen. Diabetic complications such as retinopathy or nephropathy are considered to be related to impaired coagulation-fibrinolysis system. Therefore, we studied the serum levels of HRGP in diabetics. There were no significant alterations of serum HRGP levels in comparison with normal controls, nor was there a mutual relation between HRGP levels and HbA1 values. Moreover, the molecular weight of HRGP was not deviated from control. These results suggest that HRGP might not have critical role on fibrinolytic system in diabetics.

Adult↗