During mitosis, the centrosome expands its capacity to nucleate microtubules. Understanding the mechanisms of centrosomal microtubule nucleation is, however, constrained by a lack of knowledge of the amount of soluble and polymer tubulin at mitotic centrosomes. Here we combined light microscopy and serial-section electron tomography to measure the amount of dimer and polymer at mitotic centrosomes in early C. elegans embryos. We show that a C. elegans one-cell stage centrosome at metaphase contains more than ten thousand microtubules with a total polymer concentration of 230 μM. Centrosomes concentrate soluble α/β tubulin by about tenfold over the cytoplasm, reaching peak values of 470 μM, giving a combined total monomer and polymer tubulin concentration at centrosomes of up to 660 μM. These findings support in vitro data suggesting that microtubule nucleation in C. elegans centrosomes is driven in part by concentrating soluble tubulin.
How Amira-Avizo Software is used
For segmentation and automatic tracing of microtubules, we used the AMIRA Software with an extension to the filament editor. Using the Amira software, corrected three-dimensional models were then stitched in z to obtain complete volumes of the recorded centrosomes .