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

Nat Cell Biol. 2012 Nov;14(11):1159-68. doi: 10.1038/ncb2597. Epub 2012 Oct 21.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blotting, Western
  • Cell Cycle Proteins
  • Cell Line
  • Centrioles / metabolism*
  • Centrosome / metabolism*
  • HeLa Cells
  • Humans
  • Intracellular Signaling Peptides and Proteins / metabolism
  • Microscopy
  • Microscopy, Fluorescence / methods*
  • Microtubules / metabolism
  • Nerve Tissue Proteins / metabolism
  • RNA Interference

Substances

  • CDK5RAP2 protein, human
  • Cell Cycle Proteins
  • Intracellular Signaling Peptides and Proteins
  • Nerve Tissue Proteins