Microscopy tracks transport of proteins in living cells

March 11, 2014

A 3D illustration of polarized epithelial cells with the nuclei colored blue. Some of the cells express a fluorescent protein (green) at the apical cell membrane.

Using spinning-disk confocal and total internal reflection fluorescence (TIRF) microscopy methods, scientists at the University of Freiburg in Germany have discovered a new mechanism by which proteins are transported to the outer membrane in epithelial cells (polarized cells that line the organs in the human body). Currently, this is a process that is not well understood, and further understanding of it will help to treat diseases in which this transport is impaired, such as cystic fibrosis and autosomal dominant polycystic kidney disease (ADPKD).

With the help of the two microscopy methods and the coordinated transport of fluorescent proteins in living cells, biophysicist Dr. Roland Thünauer and junior professor Dr. Winfried Römer, Institute of Biology II and member of the Cluster of Excellence BIOSS Centre for Biological Signalling Studies of the University of Freiburg, tracked the path of a protein from its synthesis to its arrival at the apical cell membrane. In collaboration with researchers from Weill Cornell Medical College (New York, NY), and CBL GmbH (Linz, Austria), they uncovered new mechanisms and signaling molecules responsible for sorting proteins and transporting them to the apical membrane.

Cell membrane proteins are synthesized in the endoplasmic reticulum. They are then sent to the so-called Golgi apparatus and shipped from there to cell compartments or the cell membrane via small vesicles. The researchers discovered that proteins destined for the apical membrane are, after leaving the Golgi apparatus, also sorted at an additional compartment—the apical recycling endosomes (AREs). The protein Rab11 plays a key role in this process: After leaving the AREs, the apical proteins are again packaged into vesicles and sent to the target membrane. Rab11 is also involved in the final stage of their journey, as the vesicles fuse with the outer membrane, allowing the proteins to reach the apical cell surface.

Full findings appear in the Proceedings of the National Academy of Sciences (PNAS); for more information, please visit http://dx.doi.org/10.1073/pnas.1304168111.