lewis-x-antigen and Teratocarcinoma

Treatment of mouse teratocarcinoma F9 cells with all-trans-retinoic acid (RA) causes a 9-fold increase in steady-state levels of mRNA for UDP-Gal:beta-D-Gal alpha1,3-galactosyltransferase (alpha1,3GT) beginning at 36 h. Enzyme activity rises in a similar fashion, which also parallels the induction of laminin and type IV collagen. Nuclear run-on assays indicate that this increase in alpha1,3GT in RA-treated F9 cells, like that of type IV collagen, is transcriptionally regulated. Differentiation also results in increased secretion of soluble alpha1,3GT activity into the growth media. The major alpha-galactosylated glycoprotein present in the media of RA-treated F9 cells, but not of untreated cells, was identified as laminin. Differentiation of F9 cells is accompanied by an increase in alpha-galactosylation of membrane glycoproteins and a decrease in expression of the stage-specific embryonic antigen, SSEA-1 (also known as the Lewis X antigen or LeX), which has the structure Galbeta1-4(Fucalpha1-3)GlcNAcbeta1-R. However, flow cytometric analyses with specific antibodies and lectins, following treatment of cells with alpha-galactosidase, demonstrate that differentiated cells contain LeX antigens that are masked by alpha-galactosylation. Thus, RA induces alpha1,3GT at the transcriptional level, resulting in major alterations in the surface phenotype of the cells and masking of LeX antigens.

Topics: Animals; Bucladesine; Carbohydrate Conformation; Carbohydrate Sequence; Carcinoma, Embryonal; Cell Differentiation; Cell Nucleus; Flow Cytometry; Galactosyltransferases; Gene Expression Regulation, Neoplastic; Glycosylation; Kinetics; Laminin; Lewis X Antigen; Membrane Glycoproteins; Mice; Molecular Sequence Data; Raffinose; RNA, Messenger; Teratocarcinoma; Time Factors; Transcription, Genetic; Tretinoin; Tumor Cells, Cultured

Human papillomaviruses (HPVs) are known to infect human keratinocytes and cause alterations in epithelial differentiation. We showed in this study that expression of the HPV-16 genome was able to interfere with the in vitro differentiation of a human simple-epithelial cell type, the 2102Ep teratocarcinoma cell line. Stable HPV-16 genome-expressing 2102Ep cell lines were generated, and subsequent alterations in differentiation were analyzed in comparison with parental 2102Ep cells. We found that in 2102Ep cells phorbol ester-induced differentiation led to changes in the expression of SSEA antigens, whereas in HPV-transfected cell lines only minor changes were observed.

Topics: Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Cell Differentiation; DNA, Viral; Gene Expression; Genes, Viral; Glycosphingolipids; Humans; Lewis X Antigen; Male; Papillomaviridae; Stage-Specific Embryonic Antigens; Teratocarcinoma; Testicular Neoplasms; Tetradecanoylphorbol Acetate; Transfection; Tumor Cells, Cultured

The efficiency of membrane fusion between reconstituted Sendai viral envelopes containing only the fusion protein (F-virosomes) and the plasma membrane of mouse teratocarcinoma cells (F9) in culture was assessed using an assay based on the relief of self-quenching of a lipid probe incorporated in the F-virosomes. The potential of F-virosomes was also evaluated for a targeted cytosolic delivery of lysozyme to F9 cells. [125I]Lysozyme entrapped into F-virosomes was taken to examine its fusion-mediated transfer to the F9 cells. Target specificity of the F-virosomes was confirmed by the interaction between the terminal Le(x) moiety (Gal beta 1-->4(Fuc alpha 1-->3)GlcNAc) of F protein and the Le(x) determinant on the membrane of F9 cells. Incubation of the loaded F-virosomes with cells led to fusion-mediated delivery, as inferred from the ability of cells to internalize lysozyme in the presence of azide (a potent inhibitor of endocytosis). These results suggest that carbohydrate-carbohydrate interaction is strong enough for target cell recognition followed by phospholipid bilayer melding induced by fusion glycoprotein of Sendai virus.

Topics: Animals; Biological Transport; Carbohydrate Sequence; Cell Membrane; Endocytosis; Kinetics; Lewis X Antigen; Membrane Fusion; Mice; Molecular Sequence Data; Muramidase; Parainfluenza Virus 1, Human; Subcellular Fractions; Teratocarcinoma; Tumor Cells, Cultured; Viral Fusion Proteins

Retinoic acid (RA), a well-known inducer of differentiation, has been shown to regulate its own receptor gene expression in F9 teratocarcinoma cells. The homologous regulation of receptors by RA might be critical for RA-induced F9 cell differentiation. F9 cell lines from two different laboratories, named F9-1 and F9-2, were compared for retinoic acid receptor (RAR) and retinoid x receptor (RXR) gene expression in response to RA. The data show that both F9-1 and F9-2 cell lines are embryonal carcinoma cells, but of different phenotypes and different sensitivity to RA. In F9-1 cells, RA regulates all three RARs (alpha, beta, and gamma), two RXRs (alpha and gamma), two activin receptors (ActR II and IIB), and tissue-specific plasminogen activator (t-PA) gene expression. In F9-2 cells RA regulates only the RAR beta, RXR alpha, and t-PA genes. The induction of mRNA levels was much higher in F9-1 than in F9-2 cells. Different basal RAR gamma and RXR gamma mRNA levels were also noted. In these two cell lines F9-2 cells expressed greater amounts of RAR gamma 1, gamma 2, and gamma 3 mRNA isoforms, but lacked RXR gamma mRNA compared with F9-1 cells. Since RAR gamma 1 has been shown to exert an antagonistic effect on other types of RA receptors, the decreased sensitivity of F9-2 cells to RA might be due to its high level of RAR gamma 1 and/or low level of RXR gamma. This notion was in part supported by gel shift assay which demonstrated constitutive binding of RAR gamma to a RA responsive element (RAR beta E) in F9-2 cells. Further, the binding of nuclear protein to RAR beta E was increased upon RA treatment in F9-1 cells, but not in F9-2 cells. These differences in the regulation of RA receptors might determine the sensitivity of the two substrains of F9 cells to RA.

Topics: Animals; Cell Differentiation; Keratins; Lewis X Antigen; Mice; Receptors, Retinoic Acid; Retinoid X Receptors; Teratocarcinoma; Transcription Factors; Tretinoin; Tumor Cells, Cultured