The axonemes of eukaryotic cilia and flagella display high tubulin glutamylation heterogeneity, yet the functional significance of this variation remains elusive. We previously showed that long-chain polyglutamylation is crucial for ciliary motility in Chlamydomonas. However, the respective contributions of long-chain polyglutamylation versus short-chain species to motility remain unclear, as existing mutants did not allow for a clear functional dissection of these two modification states. Here, we generated mutants deficient in deglutamylases, cytosolic carboxypeptidases (CCPs) 1, 2, and 5. Importantly, CCP5 is known to remove the branch-point glutamate residue, the final step in deglutamylation. While axonemal polyglutamylation levels remained largely unaffected in these mutants, abundance of short-chain glutamylation was significantly increased in both the axonemal and cytoplasmic microtubules of ccp5-1, consistent with CCP5’s role as a branch-point deglutamylase. Although each single mutant exhibited slightly reduced swimming velocity, the loss of CCP5 in the tpg1 background lacking long polyglutamate side chains resulted in a significant restoration of motility. These findings indicate that the abundance of short-chain species, regulated by CCP5, plays a distinct role in modulating ciliary motility, particularly in the absence of long polyglutamate side chains. This suggests that even minimal glutamylation can functionally support dynein-driven microtubule sliding.
Rinka Sasaki, Toshiyuki Oda, Tomohiro Kubo (2026) Tubulin monoglutamylation is sufficient to rescue the ciliary motility defects in a Chlamydomonas polyglutamylation deficient mutant., Frontiers in cell and developmental biology, Volume 14, pp. 1857091
Published in June 5, 2026 (Electronic publication in June 5, 2026, midnight )
(Abstract) INTRODUCTION: The axonemes of eukaryotic cilia and flagella display high tubulin glutamylation heterogeneity, yet the functional significance of this variation remains elusive. We previously showed that long-chain polyglutamylation is crucial for ciliary motility in Chlamydomonas. However, the respective contributions of long-chain polyglutamylation versus short-chain species to motility remain unclear, as existing mutants did not allow for a clear functional dissection of these two modification states. METHODS: To investigate the roles of different glutamylation states, we generated Chlamydomonas mutants deficient in the deglutamylases cytosolic carboxypeptidases (CCPs) 1, 2, and 5. Tubulin modifications and ciliary motility were analyzed. RESULTS: While axonemal polyglutamylation levels remained largely unaffected in these mutants, abundance of short-chain glutamylation was significantly increased in both the axonemal and cytoplasmic microtubules of ccp5-1, consistent with CCP5's role as a branch-point deglutamylase. Although each single mutant exhibited slightly reduced swimming velocity, the loss of CCP5 in the tpg1 background lacking long polyglutamate side chains resulted in a significant restoration of motility. DISCUSSION: These findings indicate that the abundance of short- chain species, regulated by CCP5, plays a distinct role in modulating ciliary motility, particularly in the absence of long polyglutamate side chains. This suggests that even minimal glutamylation can functionally support dynein- driven microtubule sliding.
Sasaki, Rinka, Oda, Toshiyuki, Kubo, Tomohiro (2026/01/01), Tubulin Monoglutamylation is Sufficient to Rescue the Ciliary Motility Defects in a Chlamydomonas Polyglutamylation Deficient Mutant, bioRxiv, 2026.04.09.717589
Published in April 9, 2026
(Abstract) None