Journal of Cell Science, Vol 108, Issue 9 3013-3028, Copyright © 1995 by Company of Biologists

JOURNAL ARTICLES

Monoclonal and polyclonal antibodies detect a new type of post-translational modification of axonemal tubulin

N Levilliers, A Fleury and AM Hill
Laboratoire de Biologie Cellulaire 4, URA 1134 CNRS, Universite de Paris-Sud, Orsay, France.

Polyclonal (PAT) and monoclonal (AXO 49) antibodies against Paramecium axonemal tubulin were used as probes to reveal tubulin heterogeneity. The location, the nature and the subcellular distribution of the epitopes recognized by these antibodies were, respectively, determined by means of: (i) immunoblotting on peptide maps of Paramecium, sea urchin and quail axonemal tubulins; (ii) immunoblotting on ciliate tubulin fusion peptides generated in E. coli to discriminate antibodies directed against sequential epitopes (reactive) from post-translational ones (non reactive); and (iii) immunofluorescence on Paramecium cells, using throughout an array of antibodies directed against tubulin sequences and post-translational modifications as references. AXO 49 monoclonal antibody and PAT serum were both shown to recognize epitopes located near the carboxyl-terminal end of both subunits of Paramecium axonemal tubulin, whereas the latter recognized additional epitopes in alpha-tubulin; AXO 49 and a fraction of the PAT serum proved to be unreactive over fusion proteins; both PAT and AXO 49 labelled a restricted population of very stable microtubules in Paramecium, consisting of axonemal and cortical ones, and their reactivity was sequentially detected following microtubule assembly; finally, both antibodies stained two upward spread bands in Paramecium axonemal tubulin separated by SDS-PAGE, indicating the recognition of various alpha- and beta-tubulin isoforms displaying different apparent molecular masses. These data, taken as a whole, definitely establish that PAT and AXO 49 recognize a post-translational modification occurring in axonemal microtubules of protozoa as of metazoa. This modification appears to be distinct from the previously known ones, and all the presently available evidence indicates that it corresponds to the very recently discovered polyglycylation of Paramecium axonemal alpha- and beta-tubulin.
CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    Twitter    What's this?

This article has been cited by other articles:

				N. Sharma, J. Bryant, D. Wloga, R. Donaldson, R. C. Davis, M. Jerka-Dziadosz, and J. Gaertig Katanin regulates dynamics of microtubules and biogenesis of motile cilia J. Cell Biol., September 7, 2007; 178(6): 1065 - 1079. [Abstract] [Full Text] [PDF]

				L. Xia, B. Hai, Y. Gao, D. Burnette, R. Thazhath, J. Duan, M.-H. Bre, N. Levilliers, M. A. Gorovsky, and J. Gaertig Polyglycylation of Tubulin Is Essential and Affects Cell Motility and Division in Tetrahymena thermophila J. Cell Biol., May 29, 2000; 149(5): 1097 - 1106. [Abstract] [Full Text] [PDF]

				K Million, J Larcher, J Laoukili, D Bourguignon, F Marano, and F Tournier Polyglutamylation and polyglycylation of alpha- and beta-tubulins during in vitro ciliated cell differentiation of human respiratory epithelial cells J. Cell Sci., January 12, 1999; 112(23): 4357 - 4366. [Abstract] [PDF]

				M.-H. Bré, V. Redeker, J. Vinh, J. Rossier, and N. Levilliers Tubulin Polyglycylation: Differential Posttranslational Modification of Dynamic Cytoplasmic and Stable Axonemal Microtubules in Paramecium Mol. Biol. Cell, September 1, 1998; 9(9): 2655 - 2665. [Abstract] [Full Text]

				K. Johnson The axonemal microtubules of the Chlamydomonas flagellum differ in tubulin isoform content J. Cell Sci., January 2, 1998; 111(3): 313 - 320. [Abstract] [PDF]

				M. Bre, V Redeker, M Quibell, J Darmanaden-Delorme, C Bressac, J Cosson, P Huitorel, J. Schmitter, J Rossler, T Johnson, et al. Axonemal tubulin polyglycylation probed with two monoclonal antibodies: widespread evolutionary distribution, appearance during spermatozoan maturation and possible function in motility J. Cell Sci., January 4, 1996; 109(4): 727 - 738. [Abstract] [PDF]