Tenascin Mr 220,000 isoform expression correlates with corneal cell migration

Development ◽  
1991 ◽  
Vol 112 (2) ◽  
pp. 605-614 ◽  
Author(s):  
A. Kaplony ◽  
D.R. Zimmermann ◽  
R.W. Fischer ◽  
B.A. Imhof ◽  
B.F. Odermatt ◽  
...  
Keyword(s):  
Cell Migration ◽  
Close Correlation ◽  
Basement Membranes ◽  
Embryonic Tissues ◽  
Corneal Fibroblasts ◽  
Sds Page ◽  
Brain Extracts ◽  
Fibronectin Type Iii ◽  
Alternatively Spliced ◽  
Fibronectin Type

The three isoforms of chicken tenascin, an extracellular matrix glycoprotein, are generated by alternatively spliced fibronectin type III domains. The resulting proteins migrate as bands of Mr 220,000 (ten220), Mr 200,000 (ten200) and Mr 190,000 (ten190) on SDS-PAGE. We describe here two monoclonal antibodies, one specific for ten220 (mAb T17) and another that recognizes all isoforms (mAb T16). These were used to examine the differential expression of isoforms during development. Most impressive is the close correlation between ten220 expression and cell migration in the embryonic cornea. Initially (stage 18), ten190/200 can be detected within the corneal epithelium and along the basement membranes of the lens and sclera. Ten220 appears within the primary stroma immediately prior to the invasion by neural-crest-derived cells. This expression is maintained during the subsequent migration of fibroblasts from the conjunctiva into the primary stroma. With the completion of migration and the marked increase in matrix synthesis by corneal fibroblasts, ten220 disappears. Ten190/200 remains in the region adjoining the endothelium, the Bowman's membrane and the adjacent stroma. The cell-migration-associated isoform is isolated from extracts of embryonic tissues as a homohexamer. Low molecular weight forms appeared absent but a new tenascin band of Mr 210,000 could be detected in brain extracts which may be a new isoform. We conclude that the synthesis of tenascin isoforms is under tight developmental control and speculate that a function of the additional domains is to facilitate cell migration.

Novel tenascin variants with a distinctive pattern of expression in the avian embryo

Development ◽  
1994 ◽  
Vol 120 (3) ◽  
pp. 637-647
Author(s):  
R.P. Tucker ◽  
J. Spring ◽  
S. Baumgartner ◽  
D. Martin ◽  
C. Hagios ◽  
...  
Keyword(s):  
Genomic Library ◽  
Chicken Embryo Fibroblast ◽  
Cdna Probe ◽  
Avian Embryo ◽  
Degenerate Primers ◽  
Type Iii ◽  
Embryonic Tissues ◽  
Fibronectin Type Iii ◽  
Fibronectin Type

Previous studies have shown that several forms of the glycoprotein tenascin are present in the embryonic extracellular matrix. These forms are the result of alternative splicing, which generates tenascin variants with different numbers of fibronectin type III repeats. We have used degenerate primers and PCR to isolate a novel tenascin exon from an avian genomic library. Genomic clones contained a sequence encoding a fibronectin type III repeat that corresponds to repeat ‘C’ from the variable domain of human tenascin. To demonstrate that tenascin containing repeat ‘C’ is actually synthesized by avian cells, a monospecific antiserum was raised against a repeat ‘C’ fusion protein. This antiserum recognized a novel high-molecular-weight variant on immunoblots of tenascin isolated from chicken embryo fibroblast-conditioned medium, and stained tendons on frozen sections of chicken embryos. A cDNA probe specific for mRNA encoding repeat ‘C’ was used for in situ hybridization. This probe hybridized in a subset of the embryonic tissues labelled with a universal tenascin probe, including tendons, ligaments and mesenchyme at sites of epithelial-mesenchymal interactions. Finally, we provide evidence that additional fibronectin type III repeats, one corresponding to a recently discovered human repeat as well as one entirely novel sequence, also exists in chicken tenascin mRNA. These data indicate that tenascin is present in the embryonic matrix in a multitude of forms and that these forms have distinctive distributions that may reflect more than one function for tenascin in development.

Domains of tenascin involved in glioma migration

1998 ◽  
Vol 111 (8) ◽  
pp. 1095-1104
Author(s):  
G.R. Phillips ◽  
L.A. Krushel ◽  
K.L. Crossin
Keyword(s):  
Cell Migration ◽  
Matrix Protein ◽  
Glioma Cells ◽  
C6 Glioma ◽  
Extracellular Matrix Protein ◽  
C6 Glioma Cells ◽  
Dependent Manner ◽  
Type Iii ◽  
Fibronectin Type Iii ◽  
Fibronectin Type

Tenascin (TN) is an extracellular matrix protein found in areas of cell migration during development and expressed at high levels in migratory tumor cells. TN was previously shown to support the attachment and migration of glioma cells in culture. To determine the domains responsible for glioma migration and attachment, we produced recombinant fusion proteins that collectively span the majority of the molecule including its epidermal growth factor-like repeats, fibronectin type III repeats and fibrinogen domain. These domains were tested for their ability to support migration of C6 glioma cells in an aggregate migration assay. A recombinant fusion protein including fibronectin type III (FNIII) repeats 2–6 (TNfn2-6) was the only fragment found to promote migration of C6 glioma cells at levels similar to that promoted by intact TN. Evaluation of smaller segments and individual FNIII repeats revealed that TNfn3 promoted migration and attachment of glioma cells and TNfn6 promoted migration but not attachment. While TNfn3 and TNfn6 promoted migration individually, the presence of both TNfn3 and TNfn6 was required for migration on segments of the FNIII region that included TNfn5. TNfn5 inhibited migration in a dose dependent manner when mixed with TNfn3 and also promoted strong attachment and spreading of C6 glioma cells. TNfn3 and TNfn6 promote cell migration and may function cooperatively to overcome the inhibitory activity of TNfn5. Additional cell attachment studies suggested that both beta1 integrins and heparin may differentially influence the attachment of glioma cells to TN fragments. Together, these findings show that C6 glioma cells integrate their response upon binding to at least three domains within TN.

scholarly journals Tenascin promotes cerebellar granule cell migration and neurite outgrowth by different domains in the fibronectin type III repeats.

1992 ◽  
Vol 116 (6) ◽  
pp. 1475-1486 ◽  
Author(s):  
K Husmann ◽  
A Faissner ◽  
M Schachner
Keyword(s):  
Cell Migration ◽  
Neurite Outgrowth ◽  
Granule Cell ◽  
Soluble Form ◽  
Cerebellar Granule ◽  
Type Iii ◽  
The Third ◽  
Extracellular Matrix Molecule ◽  
Fibronectin Type Iii ◽  
Fibronectin Type

The extracellular matrix molecule tenascin has been implicated in neuron-glia recognition in the developing central and peripheral nervous system and in regeneration. In this study, its role in Bergmann glial process-mediated neuronal migration was assayed in vitro using tissue explants of the early postnatal mouse cerebellar cortex. Of the five mAbs reacting with nonoverlapping epitopes on tenascin, mAbs J1/tn1, J1/tn4, and J1/tn5, but not mAbs J1/tn2 and J1/tn3 inhibited granule cell migration. Localization of the immunoreactive domains by EM of rotary shadowed tenascin molecules revealed that the mAbs J1/tn4 and J1/tn5, like the previously described J1/tn1 antibody, bound between the third and fifth fibronectin type III homologous repeats and mAb J1/tn3 bound between the third and fifth EGF-like repeats. mAb J1/tn2 had previously been found to react between fibronectin type III homologous repeats 10 and 11 of the mouse molecule (Lochter, A., L. Vaughan, A. Kaplony, A. Prochiantz, M. Schachner, and A. Faissner. 1991. J. Cell Biol. 113:1159-1171). When postnatal granule cell neurons were cultured on tenascin adsorbed to polyornithine, both the percentage of neurite-bearing cells and the length of outgrowing neurites were increased when compared to neurons growing on polyornithine alone. This neurite outgrowth promoting effect of tenascin was abolished only by mAb J1/tn2 or tenascin added to the culture medium in soluble form. The other antibodies did not modify the stimulatory or inhibitory effects of the molecule. These observations indicate that tenascin influences neurite outgrowth and migration of cerebellar granule cells by different domains in the fibronectin type III homologous repeats.

scholarly journals Tenascin-C contains distinct adhesive, anti-adhesive, and neurite outgrowth promoting sites for neurons.

1996 ◽  
Vol 132 (4) ◽  
pp. 681-699 ◽  
Author(s):  
B Götz ◽  
A Scholze ◽  
A Clement ◽  
A Joester ◽  
K Schütte ◽  
...  
Keyword(s):  
Neurite Outgrowth ◽  
Hippocampal Neurons ◽  
Neural Cells ◽  
Cell Binding ◽  
Tenascin C ◽  
Functional Studies ◽  
Distinct Cell ◽  
Fibronectin Type Iii ◽  
Alternatively Spliced ◽  
Fibronectin Type

The glia-derived extracellular matrix glycoprotein tenascin-C (TN-C) is transiently expressed in the developing CNS and may mediate neuron-glia interactions. Perturbation experiments with specific monoclonal antibodies suggested that TN-C functions for neural cells are encoded by distinct sites of the glycoprotein (Faissner, A., A. Scholze, and B. Götz. 1994. Tenascin glycoproteins in developing neural tissues--only decoration? Persp. Dev. Neurobiol. 2:53-66). To characterize these further, bacterially expressed recombinant domains were generated and used for functional studies. Several short-term-binding sites for mouse CNS neurons could be assigned to the fibronectin type III (FNIII) domains. Of these, the alternatively spliced insert TNfnA1,2,4,B,D supported initial attachment for both embryonic day 18 (E18) rat and postnatal day 6 (P6) mouse neurons. Only TNfn1-3 supported binding and growth of P6 mouse cerebellar neurons after 24 h, whereas attachment to the other domains proved reversible and resulted in cell detachment or aggregation. In choice assays on patterned substrates, repulsive properties could be attributed to the EGF-type repeats TNegf, and to TNfnA1,2,4. Finally, neurite outgrowth promoting properties for E18 rat hippocampal neurons and P0 mouse DRG explants could be assigned to TNfnB,D, TNfnD,6, and TNfn6. The epitope of mAb J1/tn2 which abolishes the neurite outgrowth inducing effect of intact TN-C could be allocated to TNfnD. These observations suggest that TN-C harbors distinct cell-binding, repulsive, and neurite outgrowth promoting sites for neurons. Furthermore, the properties of isoform-specific TN-C domains suggest functional significance of the alternative splicing of TN-C glycoproteins.

scholarly journals Large and small splice variants of collagen XII: differential expression and ligand binding.

1995 ◽  
Vol 130 (4) ◽  
pp. 1005-1014 ◽  
Author(s):  
M Koch ◽  
B Bohrmann ◽  
M Matthison ◽  
C Hagios ◽  
B Trueb ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Splice Variants ◽  
Expression Patterns ◽  
Collagen I ◽  
Heparin Binding ◽  
Fibronectin Type Iii ◽  
Alternatively Spliced ◽  
Collagenous Domain ◽  
Fibronectin Type

Collagen XII has a short collagenous tail and a very large, three-armed NC3 domains consisting primarily of fibronectin type III repeats. Differential splicing within this domain gives rise to a large (320 kD) and a small (220 kD) subunit; the large but not the small can carry glycosaminoglycan. To investigate whether collagen XII variants have distinct expression patterns and functions, we generated antibody and cDNA probes specific for the alternatively spliced domain. We report here that the large variant has a more restricted expression in embryonic tissue than the small. For example, whereas the small variant is widespread in the dermis, the large is limited to the base of feather buds. Distinct proportions of mRNA for the two variants were detected depending on the tissue. Monoclonal antibodies allowed us to separate collagen XII variants, and to show that homo- and heterotrimers exist. Collagen XII variants differ in ligand binding. Small subunits interact weakly with heparin via their COOH-terminal domain. Large subunits have additional, stronger heparin-binding site(s) in their NH2-terminal extra domain. In vivo, both large and small collagen XII are associated with interstitial collagen. Here we show biochemically and ultrastructurally that collagen XII can be incorporated into collagen I fibrils when it is present during, but not after, fibril formation. Removal of the collagenous domain of collagen XII reduces its coprecipitation with collagen I. Our results indicate that collagen XII is specifically associated with fibrillar collagen, and that the large variant has binding sites for extracellular ligands not present in the small variant.

scholarly journals Site-specific HNK-1 epitope on alternatively spliced fibronectin type-III repeats in tenascin-C promotes neurite outgrowth of hippocampal neurons through contactin-1

PLoS ONE ◽  
2019 ◽  
Vol 14 (1) ◽  
pp. e0210193 ◽  
Author(s):  
Ayasa Nakamura ◽  
Jyoji Morise ◽  
Keiko Yabuno-Nakagawa ◽  
Yuki Hashimoto ◽  
Hiromu Takematsu ◽  
...  
Keyword(s):  
Neurite Outgrowth ◽  
Hippocampal Neurons ◽  
Site Specific ◽  
Tenascin C ◽  
Type Iii ◽  
Fibronectin Type Iii ◽  
Alternatively Spliced ◽  
Fibronectin Type

Expression of tenascin-C by human endometrial adenocarcinoma and stroma cells: heterogeneity of splice variants and induction by TGF- b

1997 ◽  
Vol 75 (6) ◽  
pp. 759-769 ◽  
Author(s):  
Günter Vollmer ◽  
Marselina I Tan ◽  
Winfried Wünsche ◽  
Kirsten Frank
Keyword(s):  
Cell Culture ◽  
Splice Variant ◽  
Splice Variants ◽  
Endometrial Adenocarcinoma ◽  
Polymerase Chain Reaction Analysis ◽  
Tenascin C ◽  
Type Iii ◽  
Fibronectin Type Iii ◽  
Alternatively Spliced ◽  
Fibronectin Type

Localization of tenascin-C in vivo and cell culture experiments in vitro have provided evidence for stromal production of tenascin-C in malignant tumors of a variety of organs. Here we raised the question of whether the mesenchymal stroma in the case of endometrial adenocarcinoma is the unique source of tenascin-C. Therefore, the expression of tenascin-C mRNA by human endometrial adenocarcinoma cells and endometrial stroma cells was investigated. Several preparations of endometrial stroma cells produced tenascin-C mRNA. Using a serum-free defined cell culture medium, production of tenascin-C mRNA could be increased by adding either serum or 20 ng TGF- beta /mL to the cell culture medium. Reverse transcriptase polymerase chain reaction analysis revealed that five out of six endometrial adenocarcinoma cell lines produced tenascin-C mRNA. Northern blot experiments and ribonuclease protection assays provided evidence that the number of copies of tenascin-C mRNA was small. Analysis of expressed splice variants by reverse transcriptase polymerase chain reaction analysis revealed the abundance of one major splice variant that lacked all potential alternatively spliced fibronectin type-III-like repeats. Regarding larger splice variants, all fragment sizes that could theoretically originate from seven alternatively spliced fibronectin type-III-like repeats were observed. Evaluating relative signal intensities, the splice variants containing a single fibronectin type-III-like repeat and the variant possessing all but one alternatively spliced repeats were most frequent. In summary, evidence is provided that tenascin-C can originate from both tissue compartments of the human endometrium stroma and (tumor) epithelium. Splice variant analysis revealed a high number of splice variants and a relative high proportion of variants that have so far been regarded as minor constituents of expressed tenascin-C. Key words: gene expression, splice variant analysis, extracellular matrix, endometrial cancer, growth factors.

scholarly journals LAR tyrosine phosphatase receptor: alternative splicing is preferential to the nervous system, coordinated with cell growth and generates novel isoforms containing extensive CAG repeats.

1995 ◽  
Vol 128 (3) ◽  
pp. 415-431 ◽  
Author(s):  
J S Zhang ◽  
F M Longo
Keyword(s):  
Nervous System ◽  
Alternative Splicing ◽  
Cell Growth ◽  
Neural Development ◽  
Tyrosine Phosphatase ◽  
Cag Repeats ◽  
Fibronectin Type Iii ◽  
Alternatively Spliced ◽  
Fibronectin Type

Receptor-linked tyrosine phosphatases regulate cell growth by dephosphorylating proteins involved in tyrosine kinase signal transduction. The leukocyte common antigen-related (LAR) tyrosine phosphatase receptor has sequence similarity to the neural cell adhesion molecule N-CAM and is located in a chromosomal region (1p32-33) frequently altered in neuroectodermal tumors. To understand the function of receptor-linked tyrosine phosphatases in neural development, we sought to identify LAR isoforms preferentially expressed in the nervous system and cellular processes regulating LAR alternative splicing. We report here the isolation of a series of rat LAR cDNA clones arising from complex combinatorial alternative splicing, not previously demonstrated for the tyrosine phosphatase-receptor gene family in general. Isoforms included: (a) deletions of the fourth, sixth and seventh fibronectin type III-like domains; (b) an alternatively spliced novel cassette exon in the fifth fibronectin type III-like domain; (c) two alternatively spliced novel cassette exons in the juxtamembrane region; (d) a retained intron in the extracellular region with in-frame stop codons predicting a secreted LAR isoform; and (e) an LAR transcript including an alternative 3' untranslated region containing multiple stretches of tandem CAG repeats up to 21 repeats in length. This number of repeats was in the range found in normal alleles of genes in which expansions of repeats are associated with neurodegenerative disease and the genetic phenomenon of anticipation. RT-PCR and Northern analysis demonstrated that LAR alternative splicing occurred preferentially in neuromuscular tissue in vivo and in neurons compared to astrocytes in vitro and was developmentally regulated. Alternative splicing was also regulated in PC12 cells by NGF, in 3T3 fibroblasts by cell confluence and in sciatic nerve and muscle subsequent to nerve transection. Western blot analysis demonstrated that alternatively spliced cassette exons result in the presence of corresponding amino acid segments of LAR protein in vivo. These studies suggest specialized functions of LAR isoforms in the nervous system and support our hypothesis that LAR-like tyrosine phosphatase receptors play a role in neural development and regeneration.

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