Subdiffraction-resolution fluorescence microscopy reveals a domain of the centrosome critical for pericentriolar material organization

Abstract

As the main microtubule-organizing centre in animal cells, the centrosome has a fundamental role in cell function. Surrounding the centrioles, the pericentriolar material (PCM) provides a dynamic platform for nucleating microtubules. Although the importance of the PCM is established, its amorphous electron-dense nature has made it refractory to structural investigation. By using SIM and STORM subdiffraction-resolution microscopies to visualize proteins critical for centrosome maturation, we demonstrate that the PCM is organized into two main structural domains: a layer juxtaposed to the centriole wall, and proteins extending farther away from the centriole organized in a matrix. Analysis of Pericentrin-like protein (PLP) reveals that its carboxy terminus is positioned at the centriole wall, it radiates outwards into the matrix and is organized in clusters having quasi-nine-fold symmetry. By RNA-mediated interference (RNAi), we show that PLP fibrils are required for interphase recruitment and proper mitotic assembly of the PCM matrix. Centrosomes, the microtubule nucleation centre of most cells, consist of two centrioles surrounded by pericentriolar material (PCM). The PCM has been considered as amorphous but, using subdiffraction fluorescence microscopy approaches, Agard and colleagues now reveal the organized structure of the PCM of Drosophila centrosomes.