Microtubules (MTs) are dynamic polymers composed of α- and β-tubulin heterodimers. Most organisms have multiple tubulin isotypes encoded by different genes. In C. elegans, nine α-tubulins and six β-tubulins are present. Among them, two α-tubulins (tba-1 and tba-2; hereafter called α1 and α2) and two β-tubulins (tbb-1 and tbb-2; hereafter called β1 and β2) are the principal isotypes during embryogenesis. While the depletion of α1, α2 or β1 does not affect viability, β2 depletion causes abnormally active spindle movement which leads to partial embryonic lethality, implying that the two β-tubulin isotypes distinctively contribute to the behaviors of MTs. However, how tubulin isotypes affect MT dynamics in vivo is still poorly understood.
To further understand the distinct contribution of tubulin isotypes in MT dynamics in C. elegans embryos, we have generated in-frame GFP insertions in the genomic locus of β1 or β2 using the CRISPR-Cas9 system. Live imaging of these strains revealed that β2 was incorporated into the spindle MTs twice as much as β1. We next measured the parameters of the MT dynamics in embryos lacking either β1 or β2, using a growing MT plus-end marker GFP::EBP-2. In embryos lacking β1, both the growth rate and the catastrophe frequency of MTs were slightly decreased, resulting in the equivalent length of MTs with the wild type despite the alteration of MT dynamics. On the other hand, in embryos lacking β2, the growth rate was severely decreased and the catastrophe frequency was increased, resulting in shorter MTs that lead to abnormally active spindle movement. The decreased growth rate (and resulting partial embryonic lethality) by β2 depletion was enhanced by α1 depletion, whereas it was partially suppressed by α2 depletion. These results indicate that different tubulin isotypes—and different combinations of αβ-heterodimers—distinctively affect MT dynamics.
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