Localization of α-, γ-, and δ-tubulin in the hypotrich ciliate Stylonychia pustulata (Hyportrichida, Ciliophora)

Abstract

Ciliature microtubular organelles, which organize into ciliary units, are highly complex microtubular systems. To investigate the formation of ciliature microtubular organelles and the role that different tubulins play in this process, we examined the distribution and function of α-, γ-, and δ-tubulins in the hypotrich ciliate Stylonychia pustulata. Immunofluorescence analysis demonstrated that α-tubulin localized to all microtubular organelles, while γ- and δ-tubulins colocalized at basal bodies. Immunogold labeling revealed that γ- and δ-tubulins colocalize to the proximal part of basal bodies, suggesting that γ- and δ-tubulins probably interact and may function together. In addition, the permanent localization of γ-tubulin to the adoral zone of membranelles suggests that γ-tubulin play a role in the maintenance of oral ciliature base-associated microtubules, but does not play a role in the biogenesis of these ciliature. We observe that δ-tubulin is restricted to the basal body and the connective fibers of the basal body, suggesting this protein may be required for the extension and stability of the microtubules in these structures. Dramatically different concentrations of these three tubulins were observed in the cytoskeleton and the insoluble and soluble fractions of the cell, suggesting that these highly related proteins perform different functions in S. pustulata.