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ABSTRACT: Engelbreth-Holm-Swarm (EHS) tumors produce large amounts of basement membrane (BM) components that are widely used as cell culture substrates mimicking BM functions. To delineate the tissue/organ origin of the tumor and the mechanisms operating in the BM overproduction, a genome-wide expression profile of EHS tumor was analyzed using RIKEN cDNA microarrays containing approximately 40,000 mouse cDNA clones. Expression profiles of F9 embryonal carcinoma cells that produce laminin-1 and other BM components upon differentiation into parietal endoderm-like cells (designated F9-PE) were also analyzed. Hierarchical clustering analysis showed that the gene expression profiles of EHS and F9-PE were the most similar among 49 mouse tissues/organs in the RIKEN Expression Array Database, suggesting that EHS tumor is parietal endoderm-derived. Quantitative PCR analysis confirmed that not only BM components but also the machineries required for efficient production of BM components, such as enzymes involved in post-translational modification and molecular chaperones, were highly expressed in both EHS and F9-PE. Pairs of similar transcription factor isoforms, such as Gata4/Gata6, Sox7/Sox17, and Cited1/Cited2, were also highly expressed in both EHS tumor and F9-PE. Time course analysis of F9 differentiation showed that up-regulation of the transcription factors was associated with those of BM components, suggesting their involvement in parietal endoderm specification and overproduction of the BM components.
Journal of Biological Chemistry 01/2004; 278(50):50691-701. · 4.77 Impact Factor
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ABSTRACT: Laminins, the major basement membrane glycoproteins, are composed of three subunits. We identified a splice variant of the human laminin alpha4 subunit transcript containing 21 extra nucleotides. A heptapeptide sequence, MDCPTIS, was inserted close to the two cysteine residues possibly involved in the intersubunit disulfide bonds. Both the authentic alpha4 subunit (alpha4A) and the variant with the heptapeptide insertion (alpha4B) were readily secreted as laminin-8 trimers (alpha4Abeta1gamma1 or alpha4Bbeta1gamma1) upon cotransfection with expression vectors for the beta1 and gamma1 subunits. The purified recombinant laminin-8 containing the alpha4B subunit was more potent in promoting cell spreading than that containing alpha4A, raising the possibility that the alternative splicing of the alpha4 subunit transcript regulates the cell-adhesive activity of laminin-8. Since both alpha4A and alpha4B transcripts were detected by RT-PCR in several human cell lines, these two isoforms of laminin-8 with differing cell-adhesive activities are present in the basement membranes of human tissues.
Biochemical and Biophysical Research Communications 01/2003; 299(3):498-504. · 2.48 Impact Factor
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ABSTRACT: Laminins, the major basement membrane glycoproteins, are composed of three subunits. We identified a splice variant of the human laminin α4 subunit transcript containing 21 extra nucleotides. A heptapeptide sequence, MDCPTIS, was inserted close to the two cysteine residues possibly involved in the intersubunit disulfide bonds. Both the authentic α4 subunit (α4A) and the variant with the heptapeptide insertion (α4B) were readily secreted as laminin-8 trimers (α4Aβ1γ1 or α4Bβ1γ1) upon cotransfection with expression vectors for the β1 and γ1 subunits. The purified recombinant laminin-8 containing the α4B subunit was more potent in promoting cell spreading than that containing α4A, raising the possibility that the alternative splicing of the α4 subunit transcript regulates the cell-adhesive activity of laminin-8. Since both α4A and α4B transcripts were detected by RT-PCR in several human cell lines, these two isoforms of laminin-8 with differing cell-adhesive activities are present in the basement membranes of human tissues.
Biochemical and Biophysical Research Communications 299(3):498-504. · 2.48 Impact Factor
