TTLL10 can perform tubulin glycylation when co-expressed with TTLL8

Edited by Noboru Mizushima

Received 16 March 2009; received in revised form 27 April 2009; accepted 2 May 2009. published online 08 May 2009.

Abstract

Tubulin can undergo unusual post-translational modifications, glycylation and glutamylation. We previously failed to find glycylase (glycine ligase) for tubulin while identifying TTLL10 as a polyglycylase for nucleosome assembly protein 1. We here examine whether TTLL10 performs tubulin glycylation. We used a polyclonal antibody (R-polygly) raised against a poly(glycine) chain, which does not recognize monoglycylated protein. R-polygly strongly reacted with mouse tracheal cilia and axonemal tubulins. R-polygly detected many proteins in cell lysates co-expressing TTLL10 with TTLL8. Two-dimensional electrophoresis revealed that the R-polygly-reactive proteins included α- and β-tubulin. R-polygly labeling signals overlapped with microtubules. These results indicate that TTLL10 can strongly glycylate tubulin in a TTLL8-dependent manner. Furthermore, these two TTLL proteins can glycylate unidentified 170-, 110-, 75-, 40-, 35-, and 30-kDa acidic proteins.

Abbreviations: NAP1, nucleosome assembly protein 1, PTM, post-translational modification, TTL, tubulin tyrosine ligase, TTLL, TTL-like, R-polygly, anti-poly-glycine polyclonal antibody, mAb, monoclonal antibody, cDNA, complementary DNA, PCR, polymerase chain reaction, PBS, phosphate-buffered saline

Keywords: Glycylase, Monoglycylase, Polyglycylase, Glutamylation, Glutamylase, Tubulin tyrosine ligase

a Department of Molecular Anatomy, Molecular Imaging Advanced Research Center, Hamamatsu University School of Medicine, 1-20-1 Handayama, Hamamatsu, Shizuoka 431-3192, Japan

b Mitsubishi Kagaku Institute of Life Sciences (MITILS), Machida, Tokyo 194-8511, Japan

Corresponding author. Address: Department of Molecular Anatomy, Molecular Imaging Advanced Research Center, Hamamatsu University School of Medicine, 1-20-1 Handayama, Hamamatsu, Shizuoka 431-3192, Japan. Fax: +81 53 435 2292.

PII: S0014-5793(09)00360-3

doi:10.1016/j.febslet.2009.05.003

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