The ultrastructure of mono- and holocentric plant centromeres: an immunological investigation by structured illumination microscopy and scanning electron microscopy

摘要: The spatial distribution of the three centromere-associated proteins α-tubulin, CENH3, and phosphorylated histone H2A (at threonine 120, H2AThr120ph) was analysed by indirect immunodetection at monocentric cereal chromosomes holocentric Luzula elegans super-resolution light microscopy scanning electron (SEM). Using structured illumination (SIM) as technique on squashed specimens SEM uncoated isolated specimens, three-dimensional (3D) visualized centromeres. Technical aspects 3D are explained in detail. We show that CENH3 forms curved “pads” mainly around lateral centromeric region primary constriction metacentric chromosomes. H2AThr120ph is present both pericentromere. α-tubulin-labeled microtubule bundles attach to CENH3-containing chromatin structures, either single with a V-shaped attachment centromere or split bordering pericentromeric flanks. In L. chromosomes, located predominantly groove each chromatid proven subsequent FIB/FESEM ablation reconstruction. α-tubulin localizes edges groove. no additional intermediate structures between microtubules were observed. established models sites for plant