Yuzuru Ikehara

α1,3‐Fucoslytransferase IX (Fuc‐TIX) is very highly conserved between human and mouse; molecular cloning, characterization and tissue distribution of human Fuc‐TIX

The amino acid sequence of Fuc‐TIX is very highly conserved between mouse and human. The number of non‐synonymous nucleotide substitutions of the Fuc‐TIX gene between human and mouse was strikingly low, and almost equivalent to that of the α‐actin gene. This indicates that Fuc‐TIX is under a strong selective pressure of preservation during evolution. The human Fuc‐TIX (hFuc‐TIX) showed a unique characteristics, i.e. hFuc‐TIX was not activated by Mn2+ and Co2+, whereas hFuc‐TIV and hFuc‐TVI were activated by the cations. The hFuc‐TIX transcripts were abundantly expressed in brain and stomach, and interestingly were detected in spleen and peripheral blood leukocytes.

Up‐regulation of Lewis enzyme (Fuc‐TIII) and plasma‐type α1,3Fucosyltransferase (Fuc‐TVI) expression determines the augmented expression of sialyl Lewis x antigen in non‐small cell lung cancer

Sialyl Lewis a and x antigens are well‐known tumor‐associated antigens expressed in many cancer tissues. The expression of the genes encoding 5 α1,3fucosyltransferases, which are able to synthesize the sialyl Lewis antigens, was examined in normal and cancerous lung tissues of patients with non‐small cell lung carcinoma. In all 20 cases examined, the transcripts only for the Lewis gene, encoding the Lewis enzyme (α1,3/4fucosyltransferase, Fuc‐TIII), were abundantly expressed in lung tissue, and interestingly they were markedly up‐regulated in the lung cancer tissues of all 20 cases in comparison with normal lung tissues. Myeloid‐type α1,3fucosyltransferase (Fuc‐TIV) was expressed at an intermediate level but was not up‐regulated in lung cancer tissues. The transcripts for plasma‐type α1,3fucosyltransferase (Fuc‐TVI) gene were detected at a very low level but were apparently up‐regulated in cancer tissues. Fuc‐TVI was found to exhibit stronger relative activity for sialyl Lewis x synthesis (almost 6.4‐fold that of Fuc‐TIII). The amount of sialyl Lewis x antigen on mucins in the lung cancer tissues was found to be determined by both enzymes, the Lewis enzyme (Fuc‐TIII) and Fuc‐TVI. However, the amount of the sialyl Lewis a antigens was not determined by any of the α1,3‐fucosyltransferases, although the expression of sialyl Lewis a antigens definitely required the Lewis enzyme. Int. J. Cancer 83:70–79, 1999.

Determination of ABO genotypes with DNA extracted from formalin-fixed, paraffin-embedded tissues.

SummaryThe gene encoding the specific glycosyltransferases which catalyze the conversion of the H antigen to A or B antigens shows a slight but distinct variation in its allelic nucleotide sequence and can be divided into 6 genotypes when digested with specific restriction enzymes. We extracted DNA from formalin-fixed, paraffin-embedded tissues using SDS/proteinase K treatment followed by phenol/chloroform extraction. The sequence of nucleotides for the A, B and O genes was amplified by the polymerase chain reaction (PCR). DNA fragments of 128 by and 200 by could be amplified in the second round of PCR, using an aliquot of the first round PCR product as template. Degraded DNA from paraffin blocks stored for up to 10.7 years could be successfully typed. The ABO genotype was deduced from the digestion patterns with an appropriate combination of restriction enzymes and was compatible with the phenotype obtained from the blood sample.ZusammenfassungDas Gen wird für die spezifischen Glykosyltransferasen kodiert, welche die Konversion des H-Antigens in das A- oder B-Antigen katalysieren, weist eine leichte, aber definierte Variation seiner allelischen Nukleotid-Sequenz auf. Auf diese Weise lassen sich 6 Genotypen unterscheiden, nach Spaltung mit spezifischen Restriktionsenzymen. Wir haben DNA von formalinfixierten, paraffineingebetteten Geweben extrahiert, indem wir SDS-Proteinase-K-Behandlung mit nachfolgender Phenol-Chloroform-Extraktion anwandten. Die Sequenz der Nukleotide für die A-, B- und O-Gene wurde amplifiziert mit Hilfe der Polymerase-Kettenreaktion (PCR). DNA-Fragmente mit 128 Bp und 200 Bp konnten in der zweiten PCR-Phase amplifiziert werden, indem ein Aliquot der ersten PCR-Phase eingesetzt wurde. Degradierte DNA von Paraffin-Blöcken, welche bis zu 10,7 Jahren gelagert waren, konnte erfolgreich typisiert werden. Der ABO-Genotyp wurde dann abgeleitet von den Mustern nach Spaltung mit einer geeigneten Kombination von Restriktionsenzyneu und war kompatibel mit dem Phänotyp, der von der Blutprobe erhalten wurde.

An immunohistochemical study of β1,4-galactosyltransferase in human skin tissue

An immunohistochemical investigation of beta1,4-galactosyltransferase (beta1,4-GalT) on human skin tissue was performed on formalin-fixed paraffin-embedded sections using a monoclonal antibody, MAb8628, which specifically recognizes a protein moiety of human beta1,4-GalT. Distribution of the galactose beta1,4-N-acetylglucosamine (Gal beta1,4GlcNAc)-R epitope was also detected by staining with Ricinus communis agglutinin (RCA) 120. The beta1,4-GalT was observed to be localized at the perinuclear region of epidermal keratinocytes. The fine localization was also observed at the supranuclear region in the cells of apocrine glands, eccrine ducts and glands. The positive staining with RCA 120 was well colocalized with the cells expressing the beta1,4-GalT. An electron microscopic study revealed that positive signals of beta1,4-GalT definitely reside in the Golgi apparatus. No immunoreactivity was observed in any other intracellular structure or on the cell surface. These findings strongly indicated that the beta1,4-GalT is the major enzyme responsible for the Gal beta1,4GlcNAc-R epitope synthesis in human skin tissue.

Lectin- and Immuno-Histochemistry on Mucous Substances of the Taste Buds and Lingual Glands in Some Mammals

The ABO blood group antigens were discovered at the beginning of this century. Although the biochemical and molecular-biochemical nature of the ABH and related antigens is now quite well characterized and it is presumed that they are concerned with oncodevelopmental and/or cell recognition phenomena(1), no clear explanation of the biological significance of these antigens has been proposed. Characterization of cells with regard to their surface and secretory glycoconjugates is important in order to understand the basis of their physiological and interactive behaviour. The taste buds of vertebrates are covered by mucous substances and it has been suggested that the carbohydrate residues of the mucous substances are essential for chemoreceptor processes in the function of taste buds. Therefore, in the present study, we examined the distribution of different types of carbohydrate antigens in the taste buds cells and lingual glands using monoclonal antibodies(MoAb) against blood group antigens and lectins conjugated with horseradish peroxidase(HRP).

Comparative Lectin and Immuno-Histochemistry on Antigen Expression in Blood Group A1 and A2 Individuals

Blood group A has been divided into the major subgroups, A1 and A2, since it was first described by von Dungern and Hirszfeld in 19111).