The Expression and Function of Tubulin Isotypes in Caenorhabditis elegans

Abstract

Microtubules, made from the polymerization of the highly conserved α/β-tubulin heterodimers, serve as important components of the cytoskeleton in all eukaryotic cells. The existence of multiple tubulin isotypes in metazoan genomes and a dazzling variety of tubulin posttranslational modifications (PTMs) prompted the “tubulin code” hypothesis, which proposed that microtubule structure and functions are determined by the tubulin composition and PTMs. Evidence for the tubulin code has emerged from studies in several organisms with the characterization of specific tubulins for their expression and functions. The studies of tubulin PTMs are accelerated by the discovery of the enzymes that add or remove the PTMs. In tubulin research, the use of simple organisms, such as Caenorhabditis elegans, has been instrumental for understanding the expression and functional specialization of tubulin isotypes and the effects of their PTMs. In this review, we summarize the current understanding of the expression patterns and cellular functions of the nine α-tubulin and six β-tubulin isotypes. Expression studies are greatly facilitated by the CRISPR/Cas9-mediated endogenous GFP knock-in reporters and the organism-wide single cell transcriptomic studies. Meanwhile, functional studies benefit from the ease of genetic manipulation and precise gene replacement in C. elegans. These studies identified both ubiquitously expressed tubulin isotypes and tissue-specific isotypes. The isotypes showed functional redundancy, as well as functional specificity, which is likely caused by the subtle differences in their amino acid sequences. Many of these differences concentrate at the C-terminal tails that are subjected to several PTMs. Indeed, tubulin PTM, such as polyglutamylation, is shown to modulate microtubule organization and properties in both ciliated and non-ciliated neurons. Overall, studies from C. elegans support the distinct expression and function patterns of tubulin isotypes and the importance of their PTMs and offer the promise of cracking the tubulin code at the whole-genome and the whole-organism level.

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“…S3 A) (Shapovalov and Dunbrack, 2011). Remarkably, deletion of TBA-5 (or loss-of-function TBA-5) has negligible effects on cilia due to the functional redundancy of tubulin isotypes (Hao et al, 2011; Lu and Zheng, 2022), while A19V mutation transformed the configuration of TBA-5 to ‘gain-of-function’, thereby destructing ciliary MTs upon its assembly into MTs, resulting in the loss of the ciliary distal segments (Fig. 3 A) (Hao et al, 2011).…”

“…Leveraging the C. elegans system -- renowned for its suitability for genetic manipulation and live-cell imaging -- we carried out initial investigations. The C. elegans genome harbors nine α-tubulin and six β-tubulin isotypes (Lu and Zheng, 2022). Supposing that regions of low sequence conservation might be more permissive for tagging, we targeted the H1-S2 loop of the tubulin chain, a notably variable region (Fig.…”

“…Concurrently, glh-1::T2A::gfp1-10 , a knock-in (KI) allele of the germline-specific helicase GLH-1, provided endogenous GFP1-10 fragments as the T2A self-split sequence ensured a definitive partition between GLH-1 and GFP1-10 (Goudeau et al, 2021). During early-stage embryonic development, TBB-1 and TBB-2 are the exclusively expressed β-tubulin isotypes (Honda et al, 2017; Lu and Zheng, 2022). Previous studies revealed that RNA interference (RNAi)-mediated depletion of TBB-1 did not induce apparent phenotypic abnormalities including embryonic lethality, suggesting the predominant role of TBB-2 in mitotic MT formation (Honda et al, 2017).…”

“…Concurrently, glh-1::T2A::gfp1-10, a knock-in (KI) allele of the germlinespecific helicase GLH-1, provided endogenous GFP1-10 fragments as the T2A selfsplit sequence ensured a definitive partition between GLH-1 and GFP1-10 (Goudeau et al, 2021). During early-stage embryonic development, TBB-1 and TBB-2 are the exclusively expressed β-tubulin isotypes (Honda et al, 2017;Lu and Zheng, 2022).…”

“…Leveraging the C. elegans system --renowned for its suitability for genetic manipulation and live-cell imaging --we carried out initial investigations. The C. elegans genome harbors nine α-tubulin and six β-tubulin isotypes (Lu and Zheng, 2022).…”