Super-resolution Microscopy Reveals Compartmentalization of Peroxisomal Membrane Proteins

J Biol Chem. 2016 Aug 12;291(33):16948-62. doi: 10.1074/jbc.M116.734038. Epub 2016 Jun 16.


Membrane-associated events during peroxisomal protein import processes play an essential role in peroxisome functionality. Many details of these processes are not known due to missing spatial resolution of technologies capable of investigating peroxisomes directly in the cell. Here, we present the use of super-resolution optical stimulated emission depletion microscopy to investigate with sub-60-nm resolution the heterogeneous spatial organization of the peroxisomal proteins PEX5, PEX14, and PEX11 around actively importing peroxisomes, showing distinct differences between these peroxins. Moreover, imported protein sterol carrier protein 2 (SCP2) occupies only a subregion of larger peroxisomes, highlighting the heterogeneous distribution of proteins even within the peroxisome. Finally, our data reveal subpopulations of peroxisomes showing only weak colocalization between PEX14 and PEX5 or PEX11 but at the same time a clear compartmentalized organization. This compartmentalization, which was less evident in cases of strong colocalization, indicates dynamic protein reorganization linked to changes occurring in the peroxisomes. Through the use of multicolor stimulated emission depletion microscopy, we have been able to characterize peroxisomes and their constituents to a yet unseen level of detail while maintaining a highly statistical approach, paving the way for equally complex biological studies in the future.

Keywords: STED microscopy; membrane protein; membrane trafficking; microscopy; peroxisome; protein import; super-resolution optical microscopy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Carrier Proteins / metabolism*
  • Cell Line
  • Humans
  • Intracellular Membranes / metabolism*
  • Membrane Proteins / metabolism*
  • Microscopy
  • Peroxisome-Targeting Signal 1 Receptor
  • Peroxisomes / metabolism*
  • Receptors, Cytoplasmic and Nuclear / metabolism*
  • Repressor Proteins / metabolism*


  • Carrier Proteins
  • Membrane Proteins
  • PEX14 protein, human
  • PEX5 protein, human
  • Peroxisome-Targeting Signal 1 Receptor
  • Receptors, Cytoplasmic and Nuclear
  • Repressor Proteins
  • sterol carrier proteins