TTLL12 is required for primary ciliary axoneme formation in polarized epithelial cells

Abstract

The primary cilium is a critical sensory organelle that is built of axonemal microtubules ensheathed by a ciliary membrane. In polarized epithelial cells, primary cilia reside on the apical surface and must extend these microtubules directly into the extracellular space and remain a stable structure. However, the factors regulating cross-talk between ciliation and cell polarization, as well as, axonemal microtubule growth and stabilization in polarized epithelia are not fully understood. In this study, we find TTLL12, a previously uncharacterized member of the Tubulin Tyrosine Ligase-Like (TTLL) family, localizes to the base of primary cilia and is required for cilia formation in polarized renal epithelial cells. We also show that TTLL12 directly binds to the tubulin heterodimer in vitro and regulates microtubule dynamics, stability, and post-translational modifications (PTMs). While all other TTLLs catalyze the addition of glutamate or glycine to microtubule C-terminal tails, TTLL12 uniquely affects tubulin PTMs by promoting both microtubule lysine acetylation and arginine methylation. Together, this work identifies a novel microtubule regulator and provides insight into the requirements for apical extracellular axoneme formation.

Novel inhibitors of TTLL12’s oncogenic potential that overcome suppression of ligation of nitrotyrosine to the C-terminus of detyrosinated α-tubulin

“…The mechanism by which TTLL12 inhibits nitrotyrosination remain to be discovered, and may involve protein-protein interactions rather than a direct enzymatic activity. Despite diverse attempts, TTLL12 has not been found to catalyse the various activities predicted from its resemblance to other proteins, (8) (9, 10) (12) suggesting that it is a pseudo-enzyme (10). TTLL12 has been reported to bind α-tubulin (10) (12), indicating that it may physically hinder tyrosine/nitrotyrosine ligation or promote removal.…”

“…Despite diverse attempts, TTLL12 has not been found to catalyse the various activities predicted from its resemblance to other proteins, (8) (9, 10) (12) suggesting that it is a pseudo-enzyme (10). TTLL12 has been reported to bind α-tubulin (10) (12), indicating that it may physically hinder tyrosine/nitrotyrosine ligation or promote removal.…”

“…TTLL12 has SET-like and TTL-like domains but no apparent consequential activity, as either a methylase, tyrosine ligase, glutamylase or glycylase (8-10). However, TTLL12 has indirect effects on tubulin detyrosination, acetylation and methylation, as well as histone methylation (10, 11) [preprint: (12)]. This poorly understood TTLL merits further study, and the development of tools to probes its functions, especially given its wide implication in pathologies and cellular functions.…”

“…A novel transcript isoform has been identified in human cancers (20). TTLL12 may contribute to tumour progression through effects on chromosomal ploidy (11), mitotic duration (10) and microtubule dynamics [preprint: (12)]. Interestingly, TTLL12 is very highly conserved in evolution (10, 21) and the only related protein in rice affects the dynamics and orientation of microtubules (22).…”

“…TTLL12 is: a target for serum autoantibodies after acute angle closure glaucoma attack (28), a candidate gene for triglycerides in mice (29), and a gene that is differently expressed during lactogenic differentiation of buffalo mammary epithelial cells (30). Interestingly, TTLL12 is required for the formation of primary ciliary axonemes, which are critical sensory organelles in polarized epithelial cells [preprint: (12)].…”