Characterization of a Temperature-Sensitive Vertebrate Clathrin Heavy Chain Mutant as a Tool to Study Clathrin-Dependent Events In Vivo

2010 | journal article. A publication with affiliation to the University of Göttingen.

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​Characterization of a Temperature-Sensitive Vertebrate Clathrin Heavy Chain Mutant as a Tool to Study Clathrin-Dependent Events In Vivo​
Neumann-Staubitz, P.; Hall, S. L.; Kuo, J. & Jackson, A. P.​ (2010) 
PLoS ONE5(8) art. e12017​.​ DOI: https://doi.org/10.1371/journal.pone.0012017 

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Authors
Neumann-Staubitz, Petra; Hall, Stephanie L.; Kuo, Joseph; Jackson, Antony P.
Abstract
Clathrin and clathrin-dependent events are evolutionary conserved although it is believed that there are differences in the requirement for clathrin in yeast and higher vertebrates. Clathrin is a long-lived protein and thus, with clathrin knockdowns only long-term consequences of clathrin depletion can be studied. Here, we characterize the first vertebrate temperature-sensitive clathrin heavy chain mutant as a tool to investigate responses to rapid clathrin inactivation in higher eukaryotes. Although we created this mutant using a clathrin cryo-electron microscopy model and a yeast temperature-sensitive mutant as a guide, the resulting temperature-sensitive clathrin showed an altered phenotype compared to the corresponding yeast temperature-sensitive clathrin. First, it seemed to form stable triskelions at the non-permissive temperature although endocytosis was impaired under these conditions. Secondly, as a likely consequence of the stable triskelions at the non-permissive temperature, clathrin also localized correctly to its target membranes. Thirdly, we did not observe missorting of the lysosomal enzyme beta-glucuronidase which could indicate that the temperature-sensitive clathrin is still operating at the non-permissive temperature at the Golgi or, that, like in yeast, more than one TGN trafficking pathway exists. Fourthly, in contrast to yeast, actin does not appear to actively compensate in general endocytosis. Thus, there seem to be differences between vertebrates and yeast which can be studied in further detail with this newly created tool.
Issue Date
2010
Status
published
Publisher
Public Library Science
Journal
PLoS ONE 
ISSN
1932-6203
Sponsor
Deutsche Forschungsgemeinschaft DFG (German Research Foundation)

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