Specific antibodies for tyrosinated and detyrosinated tubulin recognize retina tubulin subpopulations that do not participate in the posttranslational tyrosination/detyrosination cycle

We have used the monoclonal YL 1/2 (Tyr antibody) and polyclonal (Glu antibody) antibodies, specific for tyrosinated and detyrosinated tubulin, respectively, to determine the levels and cellular distribution of these tubulin species in chick retina during development. At embryonic day 4, detyrosinated tubulin was restricted to the ganglion cells of the fundic region. As development progresses, immunofluorescence also appears, first, in the outermost zone of the retina and then in the plexiform layers. The Tyr antibody staining was found in the different layers and it was fairly homogeneous in distribution. Analysis by dot immunobinding showed that the ratios of tyrosinated to detyrosinated tubulin obtained at different ages do not agree with those obtained previously by an enzymatic method based on the incorporation of [14C]tyrosine found that the lack of coincidence is due to the fact that a fraction of the tubulin species determined by the Tyr and Glu antibodies does not participate in the posttranslational tyrosination/detyrosination cycle. This is a novel concept that should be considered in the interpretations of immunofluorescence studies concerning the cellular distribution of tyrosinated and detyrosinated tubulin.

[1]  H. Barra,et al.  Tyrosination State of Tubulin and the Activity of Tubulin:Tyrosine Ligase and Tubulin Carboxypeptidase in the Developing Retina of the Chick , 1989, Journal of neurochemistry.

[2]  R. Margolis,et al.  Complete separation of tyrosinated, detyrosinated, and nontyrosinatable brain tubulin subpopulations using affinity chromatography. , 1989, Biochemistry.

[3]  H. Barra,et al.  Relationship between the tyrosination state of tubulin and the activities of tubulin:tyrosine ligase and tubulin carboxypeptidase in rat muscle during development. , 1988, European journal of biochemistry.

[4]  K. Gull,et al.  Tubulin post-translational modifications and the construction of microtubular organelles in Trypanosoma brucei. , 1988, Journal of cell science.

[5]  D. Cleveland,et al.  A survey of the alpha‐tubulin gene family in chicken: unexpected sequence heterogeneity in the polypeptides encoded by five expressed genes. , 1988, EMBO Journal.

[6]  G. Gundersen,et al.  Enhanced stability of microtubules enriched in detyrosinated tubulin is not a direct function of detyrosination level , 1988, The Journal of cell biology.

[7]  M. Kirschner,et al.  Posttranslational modification and microtubule stability , 1987, The Journal of cell biology.

[8]  E. Karsenti,et al.  Control of microtubule nucleation and stability in Madin-Darby canine kidney cells: the occurrence of noncentrosomal, stable detyrosinated microtubules , 1987, The Journal of cell biology.

[9]  K. Weber,et al.  Turnover of the carboxy-terminal tyrosine of alpha-tubulin and means of reaching elevated levels of detyrosination in living cells. , 1987, Journal of cell science.

[10]  T. Kreis,et al.  Microtubules containing detyrosinated tubulin are less dynamic. , 1987, The EMBO journal.

[11]  G. Gundersen,et al.  Postpolymerization detyrosination of alpha-tubulin: a mechanism for subcellular differentiation of microtubules , 1987, The Journal of cell biology.

[12]  K. Weber,et al.  Tubulin-tyrosine ligase has a binding site on beta-tubulin: a two- domain structure of the enzyme , 1987, The Journal of cell biology.

[13]  K. Gull,et al.  Distinct localization and cell cycle dependence of COOH terminally tyrosinolated alpha-tubulin in the microtubules of Trypanosoma brucei brucei , 1987, The Journal of cell biology.

[14]  G. Gundersen,et al.  Microtubule arrays in differentiated cells contain elevated levels of a post-translationally modified form of tubulin. , 1986, European journal of cell biology.

[15]  M. Takeichi,et al.  Expression of N-cadherin adhesion molecules associated with early morphogenetic events in chick development , 1986, Nature.

[16]  J. Fletcher.  Dating of art-historical artefacts , 1986, Nature.

[17]  W. Halfter,et al.  The formation of the axonal pattern in the embryonic avian retina , 1985, The Journal of comparative neurology.

[18]  G. Gundersen,et al.  Distinct populations of microtubules: Tyrosinated and nontyrosinated alpha tubulin are distributed differently in vivo , 1984, Cell.

[19]  P. Greengard,et al.  A quantitative dot-immunobinding assay for proteins using nitrocellulose membrane filters. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[20]  M. Willingham,et al.  A rat monoclonal antibody reacting specifically with the tyrosylated form of alpha-tubulin. I. Biochemical characterization, effects on microtubule polymerization in vitro, and microtubule polymerization and organization in vivo , 1983, The Journal of cell biology.

[21]  J. Kilmartin,et al.  Rat monoclonal antitubulin antibodies derived by using a new nonsecreting rat cell line , 1982, The Journal of cell biology.

[22]  H. Barra,et al.  Total tubulin and its aminoacylated and non-aminoacylated forms during the development of rat brain. , 1980, European journal of biochemistry.

[23]  M. Flavin,et al.  Tubulin tyrosylation in vivo and changes accompanying differentiation of cultured neuroblastoma-glioma hybrid cells. , 1979, Journal of Biological Chemistry.

[24]  G. Borisy,et al.  Modification of the C-terminus of brain tubulin during development. , 1978, Biochemical and biophysical research communications.

[25]  D. Raybin,et al.  Modification of tubulin by tyrosylation in cells and extracts and its effect on assembly in vitro , 1977, The Journal of cell biology.

[26]  G. G. Deanin,et al.  The distribution of tyrosyltubulin ligase in brain and other tissues. , 1976, Biochemical and biophysical research communications.

[27]  A. Kahn.  An autoradiographic analysis of the time of appearance of neurons in the developing chick neural retina. , 1974, Developmental biology.

[28]  H. Barra,et al.  A SOLUBLE PREPARATION FROM RAT BRAIN THAT INCORPORATES INTO ITS OWN PROTEINS [14C]ARGININE BY A RIBONUCLEASE‐SENSITIVE SYSTEM AND [14C]TYROSINE BY A RIBONUCLEASE‐INSENSITIVE SYSTEM , 1973, Journal of neurochemistry.

[29]  S. Goldberg,et al.  Topographical development of the ganglion cell fiber layer in the chick retina. A whole mount study , 1972, The Journal of comparative neurology.

[30]  F. Greenwood,et al.  THE PREPARATION OF I-131-LABELLED HUMAN GROWTH HORMONE OF HIGH SPECIFIC RADIOACTIVITY. , 1963, The Biochemical journal.

[31]  Oliver H. Lowry,et al.  Protein measurement with the Folin phenol reagent. , 1951, The Journal of biological chemistry.

[32]  K. Sullivan.  Structure and utilization of tubulin isotypes. , 1988, Annual review of cell biology.

[33]  H. Barra,et al.  Tyrosination-detyrosination of the COOH-terminus of alpha-tubulin in oocytes and embryos of Bufo arenarum. , 1987, Comparative biochemistry and physiology. B, Comparative biochemistry.

[34]  G. Matsumoto,et al.  Cytoplasmic tubulin from squid nerve fully retains C-terminal tyrosine. , 1982, Journal of Biochemistry (Tokyo).

[35]  A. Coulombre,et al.  Correlations of structural and biochemical changes in the developing retina of the chick. , 1955, The American journal of anatomy.