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In situ localization with digoxigenin-labelled probes of tau-related mRNAs in the rat pancreas

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Summary

Two cDNA probes complementary to fetal rat brain tau cDNA were produced by the polymerase chain reaction (PCR) and labelled by digoxigenin-11-dUTP incorporation during the PCR elongation step. These probes were tested for thein situ localization of tau mRNAs in sections of rat cerebellum. The hybridization signal was consistent with the known localization of brain tau mRNAs, showing the validity of cDNA probes labelled by digoxigenin during the PCR.

Using these probes, anin situ hybridization protocol was established and optimized for the localization of tau-related mRNAs in sections of pancreas. The aim was to determine whether these mRNAs were expressed in the exocrine or the endocrine part of the pancreas. A positive signal was found only in the exocrine part of the pancreas, and was distributed exclusively in the cytoplasm of acinar cells.

The results described here are the first evidence for a specific expression of tau-related proteins in the exocrine pancreas.

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References

  • aizawa,h.,kamijo,m.,ohba,y.,mori,a.,okuhara,k.,kawasaki,h.,murofushi,h.,suzuki,k. &yasuda,h. (1991) Microtubule destabilization by cdc2/H1 histone kinase: phosphorylation of a ‘Pro-rich region’ in the microtubule-binding domain of MAP4.Biochem. Biophys. Res. Commun. 179, 1620–6.

    Article  PubMed  CAS  Google Scholar 

  • aviv,h. &leder,p. (1972) Purification of biologically active globin messenger RNA by chromatography on oligo-thymidylic acid-cellulose.Proc. Natl Acad. Sci. USA 69, 1408–12.

    PubMed  CAS  Google Scholar 

  • borisy,g. g.,marcum,j. g.,olmsted,j. b.,murphy,d. b. &johnson,k. a. (1975) Purification of tubulin and of associated high molecular weight proteins from porcine brain and characterization of microtubule assembly in vitro.Ann. NY Acad. Sci. 253, 107–32.

    PubMed  CAS  Google Scholar 

  • bre,m. h.,pepperkok,r.,hill,a. m.,levilliers,n.,ansorge,w.,steltzer,e. h. k. &karsenti,e. (1990) Regulation of microtubule dynamics and nucleation during polarization in MDCK II cellsJ. Cell. Biol. 111, 3013–21.

    Article  PubMed  CAS  Google Scholar 

  • buendia,b.,bre,m. h.,griffiths,g. &karsenti,e. (1990) Cytoskeletal control of centrioles movement during the establishment of polarity in MDCK cells.J. Cell Biol. 110, 1123–35.

    Article  PubMed  CAS  Google Scholar 

  • bulinski,j. c. &borisy,g. g. (1979) Self-assembly of microtubules in extracts of culture HeLa cells and the identification of HeLa microtubule-associated proteins.Proc. Natl Acad. Sci. USA 76, 293–7.

    PubMed  CAS  Google Scholar 

  • chapin,s. j. &bulinski,j. c. (1991) Non-neuronal 210 × 103 Mr microtubule-associated protein (MAP4) contains a domain homologous to the microtubule-binding domains of neuronal MAP2 and tau.J. Cell Sci. 98, 27–36.

    PubMed  CAS  Google Scholar 

  • chirgwin,j. m.,przybyla,a. e.,mcDonald,r. j. &rutter,w. j. (1979) Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease.Biochem. 18, 5294–9.

    CAS  Google Scholar 

  • cleveland,d. w.,hwo,s. y. &kirschner,m. w. (1977) Purification of tau, a microtubule-associated protein that induces assembly of microtubules from purified tubulin.J. Mol. Biol. 116, 207–25.

    PubMed  CAS  Google Scholar 

  • drubin,d. g. &kirschner,m. w. (1986) Tau protein function in living cell.J. Cell Biol. 103, 2739–46.

    Article  PubMed  CAS  Google Scholar 

  • gilbert,t.,lebivic,a.,quaroni,a. &rodriguez-boulan,e. (1991) Microtubular organization and its involvement in the biogenetic pathway of plasma membrane protein in Caco2 intestinal epithelial cells.J. Cell Biol. 113, 275–88.

    Article  PubMed  CAS  Google Scholar 

  • kanai,y.,takemura,r.,oshima,t.,mori,h.,ihara,y.,yanagisawa,m.,masaki,t. &hirokawa,n. (1989) Expression of multiple tau isoforms and microtubule bundle formation in fibroblasts transfected with a single tau cDNA.J. Cell Biol. 109, 1173–84.

    Article  PubMed  CAS  Google Scholar 

  • kanai,y.,chen,j. &hirokawa,n. (1992) Microtubule bundling by tau proteins in vivo: analysis of functional domains.EMBO J. 11, 3953–61.

    PubMed  CAS  Google Scholar 

  • komminoth,p.,merk,f. b.,leav,i.,wolfe,h. j. &roth,j. (1992) Comparison of35S- and digoxigenin-labeled RNA and oligonucleotide probes forin situ hybridization.Histochem. 98, 217–28.

    Article  CAS  Google Scholar 

  • kurihara,h. &uchida,k. (1987) Distribution of microtubules and microfilaments in exocrine (ventral prostatic epithelial cells and pancreatic exocrine cells) and endocrine (cells of the adenohypophysis and islets of Langerhans). The relationship between cytoskeleton and epithelial cell polarity.Histochem. 87, 223–7.

    Article  CAS  Google Scholar 

  • laemmli,u. k. (1970) Cleavage of structural proteins during assembly of the head of bacteriophage T4.Nature 227, 680–5.

    Article  PubMed  CAS  Google Scholar 

  • launay,j. f.,jellali,a. &vanier,m. t. (1989) Glyceraldehyde-3-phosphate dehydrogenase is a microtubule binding protein in a human colon tumor cell line.Biochim. Biophys. Acta 996, 103–9.

    PubMed  CAS  Google Scholar 

  • lee,g. &rook,s. l. (1992) Expression of tau protein in nonneuronal cells: microtubule binding and stabilization.J. Cell. Sci. 102, 227–37.

    PubMed  CAS  Google Scholar 

  • lehuerou,j.,wicker,c.,guilloteau,p.,toullec,r. &puigserver,a. (1990) Specific regulation of the gene expression of some pancreatic enzymes during postnatal development and weaning in the calf.Biochim. Biophys. Acta 1048, 257–64.

    PubMed  Google Scholar 

  • lewis,s. a.,ivanov,i. e.,lee,g. h. &cowan,n. j. (1989) Organization of microtubules in dendrites and axons is determined by a short hydrophobic zipper in microtubule-associated proteins MAP2 and tau.Nature 342, 498–505.

    PubMed  CAS  Google Scholar 

  • lion,t. &haas,o. (1990) Nonradioactive labeling of probe with digoxigenin by polymerase chain reaction.Anal. Biochem. 188, 335–7.

    Article  PubMed  CAS  Google Scholar 

  • masson,d. &kreis,t. e. (1993) Identification and molecular characterization of E-MAP-115, a novel microtubule-associated protein predominantly expressed in epithelial cells.J. Cell Biol. 123, 357–71.

    Article  PubMed  CAS  Google Scholar 

  • michalik,l.,vanier,m. t. &launay,j. f. (1991) Microtubule affinity chromatography: a new technique for isolating microtubule-binding proteins from rat pancreas.Cell. Mol. Biol. 37, 805–11.

    PubMed  CAS  Google Scholar 

  • michalik,l.,vanier,m. t. &launay,j. f. (1992) Characterization of a 67 kDa microtubule-binding protein in the pancreas from different species.Biochim. Biophys. Acta 1116, 269–73.

    PubMed  CAS  Google Scholar 

  • michalik,l.,neuville,p.,vanier,m. t. &launay,j. f. (1993) Biochemical and immunochemical identification of a microtubule-binding protein from pancreas.Cell. Motil. Cytoskel. 25, 381–90.

    CAS  Google Scholar 

  • michalik, L., Neuville, P., Vanier, M. T. &launay, J. Pancreatic tau related maps: Biochemical and immunofluorescence analysis in a tumoral cell line.Mol. Cell. Biochem. (in press).

  • mori,a.,aizawa,h.,saido,t. c.,kawasaki,h.,mizuno,k.,murofushi,h.,suzuki,k. &sakai,h. (1991) Site-specific phosphorylation by protein kinase C inhibits assembly-promoting activity of microtubule-associated protein 4.Biochem. 30, 9341–6.

    CAS  Google Scholar 

  • murofushi,h.,kotani,s.,aizawa,h.,hisanaga,s.,hirokawa,n. &sakai,h. (1986) Purification and characterization of a 190 kD microtubule associated protein from bovine adrenal cortex.J. Cell Biol. 103, 1911–9.

    Article  PubMed  CAS  Google Scholar 

  • neuville,p.,michalik,l. &launay,j. f. (1993) Identification of a microtubule-binding domain sequence in pancreatic messenger RNAs.Biochim. Biophys. Acta 1181, 317–20.

    PubMed  CAS  Google Scholar 

  • olmsted,j. b. (1986) Microtubule-associated proteins.Annu. Rev. Cell. Biol. 2, 421–57.

    Article  PubMed  CAS  Google Scholar 

  • olmsted,j. b. &lyon,h. d. (1981) A microtubule-associated protein specific to differentiated neuroblastoma cells.J. Biol. Chem. 256, 3507–11.

    PubMed  CAS  Google Scholar 

  • pelletier,g. (1977) Identification of four cell types in the human endocrine pancreas by immunoelectron microscopy.Diabetes 26, 749–56.

    PubMed  CAS  Google Scholar 

  • pepperkok,r.,bre,m. h.,davoust,j. &kreis,t. (1990) Microtubules are stabilized in confluent epithelial cells but not in fibroblasts.J. Cell Biol. 111, 3003–12.

    Article  PubMed  CAS  Google Scholar 

  • rodriguez-boulan,e. &nelson,w. j. (1989) Morphogenesis of the polarized epithelial cell phenotype.Science 245, 718–25.

    PubMed  CAS  Google Scholar 

  • simons,k., &wandinger-ness,a. (1990) Polarized sorting in epithelia.Cell 62, 207–10.

    Article  PubMed  CAS  Google Scholar 

  • takemura,r.,kanai,y. &hirokawa,n. (1991)In situ localization of tau mRNA in developing rat brain.Neurosci. 44, 393–407.

    Article  CAS  Google Scholar 

  • tombes,r. M.,peloquin,j. g. &borisy,g. g. (1991) Specific association of an M-phase kinase with isolated mitotic spindles and identification of two of its substrates as MAP4 et MA1B.Cell. Regul. 2, 861–74.

    PubMed  CAS  Google Scholar 

  • vandre,d. d.,centonze,v. e.,peloquin,j.,tombes,r. m. &borisy,g. g. (1991) Proteins of the mammalian mitotic spindle: phosphorylation/dephosphorylation of MAP4 during mitosis.J. Cell Sci. 98, 577–88.

    PubMed  CAS  Google Scholar 

  • vanier,m. t. &launay,j. f. (1986) In vitro assembly of rat pancreas tubulin in the presence of taxol.Biochim. Biophys. Acta. 871, 71–7.

    Google Scholar 

  • wadsworth,p. &mcGrail,m. (1990) Interphase micro tubule dynamics are cell type specific.J. Cell Sci. 95, 23–32.

    PubMed  Google Scholar 

  • weatherbee,j. a.,sherline,p.,mascardo,r. n.,isant,j. g.,luftig,r. b. &weihing,r. r. (1982) Microtubule-associated proteins of HeLa cells: heat stability of the 200 000 mol wt HeLa MAPs and detection of the presence of MAP2 in HeLa cell extracts and cycled microtubulesJ. Cell Biol. 92, 155–63.

    Article  PubMed  CAS  Google Scholar 

  • west, R. R., Tenbarge, K., Gorman, M., Goldstein, L. S. B. &olmsted, J. B. (1988) Characterization of the MAP4 family.J. Cell Biol. 107, 460a (abstract).

    Google Scholar 

  • west,r. r.,tenbarge,k. m. &olmstedj. b. (1991) A model for microtubule-associated protein 4 structure. Domains defined by comparisons of human, mouse and bovine sequences.J. Biol. Chem. 266, 21 886–96.

    Google Scholar 

  • wiche,g. (1989) High microtubule-associated proteins: properties and functions.Biochem. J. 259, 1–12.

    PubMed  CAS  Google Scholar 

  • wiche,g.,oberkanins,c. &himmler,a. (1991) Molecular structure and function of microtubule-associated proteins.Int. Rev. of Cytol. 124, 217–73.

    CAS  Google Scholar 

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  1. INSERM, Unité 381, Ontogénèse et pathologie du système digestif, 3 Av. Molière, 67200, Strasbourg, France

    P. Neuville, M. T. Vanier, L. Michalik & J. F. Launay

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  1. P. Neuville
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  2. M. T. Vanier
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  3. L. Michalik
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  4. J. F. Launay
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Neuville, P., Vanier, M.T., Michalik, L. et al. In situ localization with digoxigenin-labelled probes of tau-related mRNAs in the rat pancreas. Histochem J 27, 565–574 (1995). https://doi.org/10.1007/BF02388454

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  • DOI: https://doi.org/10.1007/BF02388454

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