Analysis of cellular expression of gangliosides by gene transfection I: GD3 expression in  -transfected and transformed 3Y1 correlates with anchorage-independent growth activity

Nakaishi, Hitoshi; Sanai, Yutaka; Shiroki, Kazuko; Nagai, Yoshitaka

doi:10.1016/0006-291x(88)90456-1

Cited by 20 publications

(2 citation statements)

References 13 publications

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“…Similar results were presented by Berra et al (1991). Nakaishi et al (1988) showed an increased concentration of GD3 in c-myc transfected rat fibroblastic 3Y 1 cell line, and recently The extraction and determination of gangliosides are described in Materials and Methods. Specimens 5a and 5b as well as 6a and 6b represent two tumor specimens from the same patient.…”

Section: Discussionmentioning

confidence: 99%

Expression of Gangliosides GD3 and 3′‐isoLM1 in Autopsy Brains from Patients with Malignant Tumors

Fredman

Holst

et al. 1993

Journal of Neurochemistry

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Three autopsy brains from patients who succumbed to malignant gliomas have been analyzed in various regions with regard to their ganglioside content. The study focused on the gangliosides GD3 and 3'-isoLM1, which in a previous study of biopsies were found to be associated with these tumors. In particular, 3'-isoLM1, was suggested to be a marker for malignant gliomas. The highest concentrations (200-1,000 nmol of sialic acid/g wet weight) of GD3 was found in specimens of macroscopically pure tumor, where the proportion of GD3 was, at the most, 78% (range, 11-78%) of the total ganglioside sialic acid compared with < 10% in normal brain tissue. The proportion of the total ganglioside sialic acid made up by GD3 was also elevated in the periphery of the tumor and in the same region in the opposite hemisphere, where no tumor cells were detected. In four of eight brain metastases of various carcinomas, GD3 was > 10% of the total ganglioside sialic acid (range, 3-37%). The ganglioside 3'-isoLM1, as determined by TLC-enzyme-linked immunosorbent assay using a specific monoclonal antibody (SL-50), was not present at detectable levels in any of the macroscopically homogenous tumor areas. It was, however, found in the periphery of the tumor, in the corpus callosum, and at highest concentrations in the region of the opposite hemisphere corresponding to the tumor. The concentration varied between 0.1 and 6.0 nmol/g wet weight of tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

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Section: Discussionmentioning

confidence: 99%

Expression of Gangliosides GD3 and 3′‐isoLM1 in Autopsy Brains from Patients with Malignant Tumors

Fredman

Holst

et al. 1993

Journal of Neurochemistry

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“…GD3 is known as tumor-associated antigen of rat 3Y1 cells transformed with adenovirus EJA gene and myc (49)(50)(51). A high level of GD3 is expressed in human melanoma (24,52).…”

Section: Discussionmentioning

confidence: 99%

Expression cloning of a CMP-NeuAc:NeuAc alpha 2-3Gal beta 1-4Glc beta 1-1'Cer alpha 2,8-sialyltransferase (GD3 synthase) from human melanoma cells.

Nara

Watanabe

Maruyama

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

111

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Using an expression cloning approach, we have isolated a cDNA encoding GD3 synthase (CMPNeuAc:NeuAca2-3GalJ1-4Glcf8l-1'Cer a2,8-sialyltransferase, EC 2.4.99.8), which is a key regulatory enzyme determining the prominence of the gangloide biosynthesis pathway. The cloned cDNA encodes a 341-amino acid protein containing a single transmembrane domain at its N-terminal region, suggesting that the protein has a type H transmembrane topology. The sequence of a2,8-sialyltransferase showed a high level ofsimilarity with other sialyltransferases at two conserved regions typical in the sialyltransferase family. Transfected cells containing the cloned cDNA expressed GD3 ganglioside on the cell surface, which was detectable with specific anti-GD3 antibody by immunofluorescence and immunotaining after separation of isolated glycolipids on thin-layer chromatography. The cDNA hybridized to a single mRNA species of 2.4 kb in melanoma cells. This sialyltransferase is distinctive in catalyzing the formation of the a2-8 linkage of sialic acids.important to isolate the gene coding for a2,8-sialyltransferase for understanding of the biological roles of sialic acid in glycoconjugates.To clone the cDNA of a2,8-sialyltransferase responsible for synthesis of GD3 ganglioside, which is the first ganglioside in the synthetic pathway having sialyl-a2, Gangliosides are membrane-bound glycosphingolipids containing sialic acids that are found in high concentrations on the central nervous system. The carbohydrate moieties of gangliosides undergo profound changes during mammalian development, differentiation, and malignant transformation, suggesting that they may play fundamental roles in these processes (for reviews see refs. 2 and 3 and references therein). In particular, the GD3 ganglioside has been shown to be important for cell adhesion and growth of cultured malignant cells (4, 5) and the inductive epithelial-mesenchymal interaction in the kidney development (6). Although these ganglioside biological phenomena have been known, the mechanism regulating ganglioside expression remains unclear. To address this question, cloned genes that determine ganglioside expression are essential tools.Ganglioside biosynthesis takes place in the Golgi apparatus, where glucosylceramide is glycosylated by sequential addition of galactose, N-acetylgalactosamine, and N-acetylneuraminic acid (7). These reactions are catalyzed by specific glycosyltransferases. Recently, several glycosyltransferase cDNAs responsible for glycolipids and glycoproteins have been cloned (for reviews, see refs. 8 and 9), and the regulation of cell-type-specific expression of glycosyltransferases is being intensively investigated (9). With regard to sialyltransferases, there is a family of more than 12 different enzymes showing different substrate specificities (9). To date, cDNAs of 5 different sialyltransferases have been cloned (10-16), but a sialyltransferase responsible for the sialyl-a2,8-sialyl linkage, which appears in poly(sialic acid) in glycoconjugates, has not yet b...

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Mitogenic response of near‐diploid mouse cell line m5S/1M induced by epidermal growth factor

Watanabe

Umeda

Nomura

et al. 1990

Journal Cellular Physiology

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A nonmalignant near-diploid cell line m5s/1M, established by Sasaki and Kodama (J. Cell. Res., 131:114-122, 1987), was shown to respond to the epidermal growth factor (EGF). The m5s/1M cells showed high sensitivity to post-confluence inhibition of cell division and formed a uniform monolayer after the cells had become confluent. The addition of EGF resulted in loss of contact-dependent inhibition of growth and caused a massive piling up of a multilayered array of cells after they had become confluent. When EGF was removed from the medium, the cell number decreased rapidly, and the cells formed a uniform monolayer at the density observed in the absence of EGF. m5S/1M cells have high- and low-affinity receptors for EGF (approximately 40,000 receptors per cell), and the apparent dissociation constants of the EGF-binding reactions were 3.3 nM and 0.15 nM, respectively. The effect of EGF on the intracellular mobilization of Ca2+ and the formation of inositol phosphates was studied by using the calcium-sensitive fluorescent indicator fura 2 and [3H]inositol. EGF had no effect either on the mobilization of cytosolic free calcium [( Ca2+]i) or on the formation of inositol phosphates in m5s/1M cells, whereas bradykinin induced a rapid increase in both [Ca2+]i and inositol phosphates. Analysis of the glycosphingolipid (GSL) composition of m5S/1M cells showed that globotriaosylceramide (Gb3Cer), which is known to be a Burkitt lymphoma-associated antigen, is specifically expressed in the EGF-treated cells. The expression of Gb3Cer is dependent on the presence of EGF, with a reversible shift in GSL composition being observed in the presence or absence of EGF.

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Analysis of cellular expression of gangliosides by gene transfection I: GD3 expression in -transfected and transformed 3Y1 correlates with anchorage-independent growth activity

Cited by 20 publications

References 13 publications

Expression of Gangliosides GD3 and 3′‐isoLM1 in Autopsy Brains from Patients with Malignant Tumors

Expression of Gangliosides GD3 and 3′‐isoLM1 in Autopsy Brains from Patients with Malignant Tumors

Expression cloning of a CMP-NeuAc:NeuAc alpha 2-3Gal beta 1-4Glc beta 1-1'Cer alpha 2,8-sialyltransferase (GD3 synthase) from human melanoma cells.

Mitogenic response of near‐diploid mouse cell line m5S/1M induced by epidermal growth factor

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