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Tenascin-C signaling through induction of 14-3-3 tau.

Martin D, Brown-Luedi M, Chiquet-Ehrismann R - J. Cell Biol. (2003)

Bottom Line: We found elevated levels of 14-3-3 tau transcripts and protein in cells grown on tenascin-C.Furthermore, the growth rate on tenascin-C was increased compared with the parental cells.Therefore, we postulate that tenascin-C promotes the growth of tumor cells by causing an increase in the expression of 14-3-3 tau, which in turn has a positive effect on tumor cell adhesion and growth.

View Article: PubMed Central - PubMed

Affiliation: Friedrich Miescher Institute, Novartis Forschungsstiftung, CH-4002 Basel, Switzerland.

ABSTRACT
We searched by a cDNA subtraction screen for differentially expressed transcripts in MCF-7 mammary carcinoma cells grown on tenascin-C versus fibronectin. On tenascin-C, cells had irregular shapes with many processes, whereas on fibronectin they were flat with a cobble stone-like appearance. We found elevated levels of 14-3-3 tau transcripts and protein in cells grown on tenascin-C. To investigate the consequences of an increased level of this phospho-serine/threonine-binding adaptor protein, we transfected MCF-7 cells with a construct encoding full-length 14-3-3 tau protein and selected clones with the highest expression levels. The morphology of these cells on tenascin-C was flat, resembling that of cells on fibronectin. This was reflected by a similar pattern of F-actin staining on either substratum. Furthermore, the growth rate on tenascin-C was increased compared with the parental cells. After transient transfection of HT1080 fibrosarcoma and T98G glioblastoma cells with 14-3-3 tau, only the 14-3-3 tau-expressing cells were able to adhere and survive on tenascin-C, whereas all cells adhered well on fibronectin. Therefore, we postulate that tenascin-C promotes the growth of tumor cells by causing an increase in the expression of 14-3-3 tau, which in turn has a positive effect on tumor cell adhesion and growth.

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Morphological changes of MCF-7 cells grown on a tenascin-C substratum. MCF-7 cells were cultured for 24 h in complete medium on fibronectin-coated (A) and on tenascin-C–coated (B) dishes and photographed. On fibronectin, they adopt a cobble-stone–like epithelial morphology, whereas on tenascin-C they lose the cell–cell contacts and adopt irregular shapes. Bar, 200 μm.
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fig1: Morphological changes of MCF-7 cells grown on a tenascin-C substratum. MCF-7 cells were cultured for 24 h in complete medium on fibronectin-coated (A) and on tenascin-C–coated (B) dishes and photographed. On fibronectin, they adopt a cobble-stone–like epithelial morphology, whereas on tenascin-C they lose the cell–cell contacts and adopt irregular shapes. Bar, 200 μm.

Mentions: MCF-7 cells were grown in medium containing 10% FCS for 24 h on plates coated with fibronectin or tenascin-C, respectively. The cells show completely different morphologies on these two substrates. They grow in epithelial patches on fibronectin but lose their cell–cell contacts and adopt irregular shapes with long processes on a tenascin-C substratum (Fig. 1). This is reminiscent of the results obtained with MCF-7 cells grown on collagen gels in the presence and absence of tenascin-C where the cells in the presence of tenascin-C were found to loose their cell–cell contacts and detached from the substratum (Chiquet-Ehrismann et al., 1989). To identify molecular differences between the cells grown on fibronectin versus tenascin-C, we isolated mRNA from these cells and performed a screen for differentially expressed transcripts. This screen resulted in the identification of a cDNA clone encoding the adaptor protein 14-3-3 tau. 14-3-3 proteins are a family of phospho-serine/threonine–binding proteins with >70 known ligands as diverse as kinases, phosphatases, receptors, structural proteins, and transcription factors (for reviews see Fu et al., 2000; van Hemert et al., 2001; Tzivion and Avruch, 2002). We confirmed the higher level of 14-3-3 tau transcripts in two mRNA batches isolated from independently cultured MCF-7 cells on tenascin-C versus fibronectin substrates by semiquantitative PCR. As shown in Fig. 2 A, transcript levels of GAPDH were equal, whereas 14-3-3 tau was highly increased in the samples from the cells cultured on tenascin-C compared with fibronectin. We next tested whether the level of the 14-3-3 tau protein was also affected. We loaded equal amounts of cellular protein on an SDS-PAGE followed by detection of 14-3-3 tau by a specific antiserum on an immunoblot. As shown in Fig. 2 B, the level of the 14-3-3 protein was also much higher in cell extracts of cells grown on a tenascin-C substratum than on fibronectin.


Tenascin-C signaling through induction of 14-3-3 tau.

Martin D, Brown-Luedi M, Chiquet-Ehrismann R - J. Cell Biol. (2003)

Morphological changes of MCF-7 cells grown on a tenascin-C substratum. MCF-7 cells were cultured for 24 h in complete medium on fibronectin-coated (A) and on tenascin-C–coated (B) dishes and photographed. On fibronectin, they adopt a cobble-stone–like epithelial morphology, whereas on tenascin-C they lose the cell–cell contacts and adopt irregular shapes. Bar, 200 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172641&req=5

fig1: Morphological changes of MCF-7 cells grown on a tenascin-C substratum. MCF-7 cells were cultured for 24 h in complete medium on fibronectin-coated (A) and on tenascin-C–coated (B) dishes and photographed. On fibronectin, they adopt a cobble-stone–like epithelial morphology, whereas on tenascin-C they lose the cell–cell contacts and adopt irregular shapes. Bar, 200 μm.
Mentions: MCF-7 cells were grown in medium containing 10% FCS for 24 h on plates coated with fibronectin or tenascin-C, respectively. The cells show completely different morphologies on these two substrates. They grow in epithelial patches on fibronectin but lose their cell–cell contacts and adopt irregular shapes with long processes on a tenascin-C substratum (Fig. 1). This is reminiscent of the results obtained with MCF-7 cells grown on collagen gels in the presence and absence of tenascin-C where the cells in the presence of tenascin-C were found to loose their cell–cell contacts and detached from the substratum (Chiquet-Ehrismann et al., 1989). To identify molecular differences between the cells grown on fibronectin versus tenascin-C, we isolated mRNA from these cells and performed a screen for differentially expressed transcripts. This screen resulted in the identification of a cDNA clone encoding the adaptor protein 14-3-3 tau. 14-3-3 proteins are a family of phospho-serine/threonine–binding proteins with >70 known ligands as diverse as kinases, phosphatases, receptors, structural proteins, and transcription factors (for reviews see Fu et al., 2000; van Hemert et al., 2001; Tzivion and Avruch, 2002). We confirmed the higher level of 14-3-3 tau transcripts in two mRNA batches isolated from independently cultured MCF-7 cells on tenascin-C versus fibronectin substrates by semiquantitative PCR. As shown in Fig. 2 A, transcript levels of GAPDH were equal, whereas 14-3-3 tau was highly increased in the samples from the cells cultured on tenascin-C compared with fibronectin. We next tested whether the level of the 14-3-3 tau protein was also affected. We loaded equal amounts of cellular protein on an SDS-PAGE followed by detection of 14-3-3 tau by a specific antiserum on an immunoblot. As shown in Fig. 2 B, the level of the 14-3-3 protein was also much higher in cell extracts of cells grown on a tenascin-C substratum than on fibronectin.

Bottom Line: We found elevated levels of 14-3-3 tau transcripts and protein in cells grown on tenascin-C.Furthermore, the growth rate on tenascin-C was increased compared with the parental cells.Therefore, we postulate that tenascin-C promotes the growth of tumor cells by causing an increase in the expression of 14-3-3 tau, which in turn has a positive effect on tumor cell adhesion and growth.

View Article: PubMed Central - PubMed

Affiliation: Friedrich Miescher Institute, Novartis Forschungsstiftung, CH-4002 Basel, Switzerland.

ABSTRACT
We searched by a cDNA subtraction screen for differentially expressed transcripts in MCF-7 mammary carcinoma cells grown on tenascin-C versus fibronectin. On tenascin-C, cells had irregular shapes with many processes, whereas on fibronectin they were flat with a cobble stone-like appearance. We found elevated levels of 14-3-3 tau transcripts and protein in cells grown on tenascin-C. To investigate the consequences of an increased level of this phospho-serine/threonine-binding adaptor protein, we transfected MCF-7 cells with a construct encoding full-length 14-3-3 tau protein and selected clones with the highest expression levels. The morphology of these cells on tenascin-C was flat, resembling that of cells on fibronectin. This was reflected by a similar pattern of F-actin staining on either substratum. Furthermore, the growth rate on tenascin-C was increased compared with the parental cells. After transient transfection of HT1080 fibrosarcoma and T98G glioblastoma cells with 14-3-3 tau, only the 14-3-3 tau-expressing cells were able to adhere and survive on tenascin-C, whereas all cells adhered well on fibronectin. Therefore, we postulate that tenascin-C promotes the growth of tumor cells by causing an increase in the expression of 14-3-3 tau, which in turn has a positive effect on tumor cell adhesion and growth.

Show MeSH
Related in: MedlinePlus