Rab gap gyp8: determining function of a green fluorescent protein chimera and location of a gyp8 transmembrane domain
In the model eukaryote Saccharomyces cereviseae, there are 11 Rab GTPase proteins that regulate vesicular transport between membranebound compartments (Zerial et al. 2001). Signaling by Rab proteins is determined by whether a Rab is bound to guanosine triphosphate (GTP, active) or guanosine diphosphate (GDP, inactive). Rabs depend on accessory proteins called GAPs (GTPase accelerating proteins) to trigger GTP hydrolysis and return the Rab to its inactive, GDP-bound state. Gyp8 is a mostly uncharacterized Rab GAP in yeast, first reported as a regulator of Rab1/ Ypt1 (De Antoni et al. 2002), though the specific identity of the compartment to which Gyp8 localized was not determined. A later study (Sklan et al., 2007) identified a mammalian Gyp8 ortholog, TBC1D20, that acts on Rab1 to regulate traffic between the ER and Golgi. TBC1D20 contains a transmembrane domain that anchors the Rab GAP on membranes, but no study has experimentally investigated whether Gyp8 also contains a transmembrane domain. We sought to determine which cellular compartment(s) in yeast contain Gyp8 and whether Gyp8 is anchored by a transmembrane 52 5th Annual Student Research Symposium domain. We observed a chimera of green fluorescent protein (GFP) fused to Gyp8 co-localizes with markers of peroxisomes, but localization of GFP-Gyp8 shifts to the ER when peroxisomes were eliminated from cells. There may be a previously unknown role for Gyp8 in managing vesicular transport between the ER and peroxisomes. Truncation mutations of GFP-Gyp8 indicate that a C-terminal domain that contains a computationally predicted transmembrane domain is necessary and sufficient for localization of GFP-Gyp8 to peroxisomes.
"Rab gap gyp8: determining function of a green fluorescent protein chimera and location of a gyp8 transmembrane domain,"
OSR Journal of Student Research: Vol. 5
, Article 79.
Available at: https://scholarworks.lib.csusb.edu/osr/vol5/iss1/79