Three-dimensional reconstruction of the mammalian centriole from cryoelectron micrographs: the use of common lines...

The microtubule organizing center of the animal cell (S. D. Fuller et al., 1992, Curr. Opin. Struct. Biol. 2, 264-274; D. M. Glover et al., 1993, Sci. Am. 268, 62-68; E. B. Wilson, 1925), (The Cell in Development and Heredity) comprises two centrioles and the pericentriolar material. We have completed several three-dimensional reconstructions of individual centrioles from tilt series of cryoelectron micrographs. The reconstruction procedure uses minimization of the common lines residual to define the orientation of the centriolar minefold symmetry axis and then uses this symmetry to generate a structure by weighted backprojection to 28-nm resolution. Many of the features of these reconstructions agree with previous, conventional transmission electron microscopy studies (M. Paintrand et al., 1992, J. Struct. Biol. 108, 107-128). The microtubule barrel of the centriole is roughly 500 nm long and 300 nm in diameter and the microtubule bundles appear to taper toward the distal end. In addition, we see a handedness to the pericentriolar material at the base (distal end) of the centriole which is opposite to the skew of the microtubule triplets. The region at which the microtubule barrel joins this base is intriguingly complex and includes an internal cylindrical feature which is a site of gamma tubulin localization.