Prenylated Rab Acceptor Protein Is a Receptor for Prenylated Small GTPases

Abstract

Localization of Ras and Ras-like proteins to the correct subcellular compartment is essential for these proteins to mediate their biological effects. Many members of the Ras superfamily (Ha-Ras, N-Ras, TC21, and RhoA) are prenylated in the cytoplasm and then transit through the endomembrane system on their way to the plasma membrane. The proteins that aid in the trafficking of the small GTPases have not been well characterized. We report here that prenylated Rab acceptor protein (PRA1), which others previously identified as a prenylation-dependent receptor for Rab proteins, also interacts with Ha-Ras, RhoA, TC21, and Rap1a. The interaction of these small GTPases with PRA1 requires their post-translational modification by prenylation. The prenylation-dependent association of PRA1 with multiple GTPases is conserved in evolution; the yeast PRA1 protein associates with both Ha-Ras and RhoA. Earlier studies reported the presence of PRA1 in the Golgi, and we show here that PRA1 co-localizes with Ha-Ras and RhoA in the Golgi compartment. We suggest that PRA1 acts as an escort protein for small GTPases by binding to the hydrophobic isoprenoid moieties of the small GTPases and facilitates their trafficking through the endomembrane system.Ras proteins (Ha-Ras, K-Ras, N-Ras, TC21, and Ras1/2) regulate cell growth in eukaryotic cells, and perturbation of signaling pathways by mutation and constitutive activation of Ras proteins is a common occurrence in a wide spectrum of human tumors (1). In addition to regulating cell proliferation, Ras proteins also regulate differentiation, cell death, and cell survival (1). The Ras proteins are members of a large superfamily of low molecular weight GTP-binding proteins, which include members that regulate the actin cytoskeleton (Rho and Rac), vesicle trafficking (Rabs), and nuclear transport (Ran).The biological activity of Ras proteins is controlled by a regulated GTP/GDP cycle (2). The GTP-bound, or active, Ras relays signals to downstream effector proteins. In the case of Ha-Ras, numerous effector proteins have been described, including serine/threonine kinases (c-Raf, A-Raf, and B-Raf), lipid kinases (type I phosphatidylinositol 3-kinase), and guanine nucleotide dissociation stimulators for Ral (Ral GDS 1 and RGL) (1). Activation of Ras effector pathways leads to proliferation, differentiation, cell death, and cell survival. Which biological outcome predominates is somewhat of a mystery but seems to depend on cell type and the coordinate integration of signaling pathways activated and/or inhibited within a cell. Members of the Ras superfamily are subject to post-translational modifications. The spectrum of modifications depends on the composition of the carboxyl terminus. In the case of HaRas, the protein is subject to prenylation, proteolysis, carboxylmethylation, and S-acylation/palmitoylation (3). Prenylation is the covalent attachment of farnesyl or geranylgeranyl isoprenoids at or near the carboxyl terminus of the GTPase. For Ras family members, prenylation occurs at a conserv...

Bnip3 Mediates the Hypoxia-induced Inhibition on Mammalian Target of Rapamycin by Interacting with Rheb

“…This observation suggests that PRA1 interacts with Rheb requires the C-terminal isoprenylation. PRA1 has been suggested to act as an escort protein for small GTPases by binding to the hydrophobic isoprenoid moieties of the small GTPases and facilitates their trafficking through the endomembrane system (54). Therefore, PRA1 may have a similar function for Rheb intracellular trafficking.…”

“…About 5 ϫ 10 6 clones were screened for HIS3 and lacZ gene expression using protocols as described (54). Positive clones were verified by reassessing for growth on Trp Ϫ /Leu Ϫ /His Ϫ plates and galactosidase activity by co-transforming bait plasmid and pGAD-GH vectors derived from positive yeast clones.…”

“…For example, Bnip3 clones were isolated six times from the HeLa library and two times from mouse embryo library. PRA1 has been shown to interact with several small GTPases (including Ras, Rap, Rho, TC21, and Rab) in a prenylation-dependent manner (54). Rheb protein is isoprenylated at the C-terminal CAAX motif.…”

ThePRA1Gene Family in Arabidopsis

“…In chemical synapses, PRA1 associates with Piccolo, a zinc finger protein of the presynaptic cytoskeletal matrix, suggesting a function in synaptic vesicle trafficking (Fenster et al, 2000). PRA1 also interacts with other prenylated small GTPases, such as the mouse Ha-Ras, N-Ras, TC21, and RhoA, as well as with the v-SNARE protein VAMP2 (Martincic et al, 1997;Figueroa et al, 2001), and it plays a role as modulator of the neural Glu transporter excitatory amino acid carrier 1 (Akiduki et al, 2007). Because of its capacity to interact with GTPases and SNARE proteins, PRA1 might connect these two large groups of proteins to efficiently control vesicle docking and fusion events.…”

An Arabidopsis Prenylated Rab Acceptor 1 Isoform, AtPRA1.B6, Displays Differential Inhibitory Effects on Anterograde Trafficking of Proteins at the Endoplasmic Reticulum

“…The Ypt-interacting protein (YIP) family displays an ability for binding Rabs/Ypts (Shisheva et al, 1994;Wada et al, 1997;Yang et al, 1998;Liang and Li, 2000;Figueroa et al, 2001;Pfeffer and Aivazian, 2004;Spang, 2004). The YIP family members, containing multiple transmembrane domains ranging from two to five, are divided into seven subfamilies (Yip1-Yip6 and Yif1) based on sequence homology (Pfeffer and Aivazian, 2004).…”