Small GTPase regulation of GPCR anterograde trafficking.
ABSTRACT The physiological functions of heterotrimeric G protein-coupled receptors (GPCRs) are dictated by their intracellular trafficking and precise targeting to the functional destinations. Over the past decades, most studies on the trafficking of GPCRs have focused on the events involved in endocytosis and recycling. By contrast, the molecular mechanisms underlying anterograde transport of newly synthesized GPCRs from the endoplasmic reticulum (ER) to the cell surface have only now begun to be revealed. In this review we discuss current advances in understanding the role of Ras-like GTPases, specifically the Rab and Sar1/ARF subfamilies, in regulating cell-surface transport of GPCRs en route from the ER and the Golgi.
Article: Non-conventional trafficking of the cystic fibrosis transmembrane conductance regulator through the early secretory pathway.[show abstract] [hide abstract]
ABSTRACT: The mechanism(s) of cystic fibrosis transmembrane conductance regulator (CFTR) trafficking from the endoplasmic reticulum (ER) through the Golgi apparatus, the step impaired in individuals afflicted with the prevalent CFTR-DeltaF508 mutation leading to cystic fibrosis, is largely unknown. Recent morphological observations suggested that CFTR is largely absent from the Golgi in situ (Bannykh, S. I., Bannykh, G. I., Fish, K. N., Moyer, B. D., Riordan, J. R., and Balch, W. E. (2000) Traffic 1, 852-870), raising the possibility of a novel trafficking pathway through the early secretory pathway. We now report that export of CFTR from the ER is regulated by the conventional coat protein complex II (COPII) in all cell types tested. Remarkably, in a cell type-specific manner, processing of CFTR from the core-glycosylated (band B) ER form to the complex-glycosylated (band C) isoform followed a non-conventional pathway that was insensitive to dominant negative Arf1, Rab1a/Rab2 GTPases, or the SNAp REceptor (SNARE) component syntaxin 5, all of which block the conventional trafficking pathway from the ER to the Golgi. Moreover, CFTR transport through the non-conventional pathway was potently blocked by overexpression of the late endosomal target-SNARE syntaxin 13, suggesting that recycling through a late Golgi/endosomal system was a prerequisite for CFTR maturation. We conclude that CFTR transport in the early secretory pathway can involve a novel pathway between the ER and late Golgi/endosomal compartments that may influence developmental expression of CFTR on the cell surface in polarized epithelial cells.Journal of Biological Chemistry 04/2002; 277(13):11401-9. · 4.77 Impact Factor
Article: Seven transmembrane receptor core signaling complexes are assembled prior to plasma membrane trafficking.[show abstract] [hide abstract]
ABSTRACT: Much is known about beta2-adrenergic receptor trafficking and internalization following prolonged agonist stimulation. However, less is known about outward trafficking of the beta2-adrenergic receptor to the plasma membrane or the role that trafficking might play in the assembly of receptor signaling complexes, important for targeting, specificity, and rapidity of subsequent signaling events. Here, by using a combination of bioluminescence resonance energy transfer, bimolecular fluorescence complementation, and confocal microscopy, we evaluated the steps in the formation of the core receptor-G protein heterotrimer complex. By using dominant negative Rab and Sar GTPase constructs, we demonstrate that receptor dimers and receptor-G betagamma complexes initially associate in the endoplasmic reticulum, whereas G alpha subunits are added to the complex during endoplasmic reticulum-Golgi transit. We also observed that G protein heterotrimers adopt different trafficking itineraries when expressed alone or with stoichiometric co-expression with receptor. Furthermore, deliberate mistargeting of specific components of these complexes leads to diversion of other members from their normal subcellular localization, confirming the role of these early interactions in targeting and formation of specific signaling complexes.Journal of Biological Chemistry 12/2006; 281(45):34561-73. · 4.77 Impact Factor
Article: Mutant rab8 Impairs docking and fusion of rhodopsin-bearing post-Golgi membranes and causes cell death of transgenic Xenopus rods.[show abstract] [hide abstract]
ABSTRACT: Rab8 is a GTPase involved in membrane trafficking. In photoreceptor cells, rab8 is proposed to participate in the late stages of delivery of rhodopsin-containing post-Golgi membranes to the plasma membrane near the base of the connecting cilium. To test the function of rab8 in vivo, we generated transgenic Xenopus laevis expressing wild-type, constitutively active (Q67L), and dominant negative (T22N) forms of canine rab8 in their rod photoreceptors as green fluorescent protein (GFP) fusion proteins. Wild-type and constitutively active GFP-rab8 proteins were primarily associated with Golgi and post-Golgi membranes, whereas the dominant negative protein was primarily cytoplasmic. Expression of wild-type GFP-rab8 had minimal effects on cell survival and intracellular structures. In contrast, GFP-rab8T22N caused rapid retinal degeneration. In surviving peripheral rods, tubulo-vesicular structures accumulated at the base of the connecting cilium. Expression of GFP-rab8Q67L induced a slower retinal degeneration in some tadpoles. Transgene effects were transmitted to F1 offspring. Expression of the GFP-rab8 fusion proteins appears to decrease the levels of endogenous rab8 protein. Our results demonstrate a role for rab8 in docking of post-Golgi membranes in rods, and constitute the first report of a transgenic X. laevis model of retinal degenerative disease.Molecular Biology of the Cell 09/2001; 12(8):2341-51. · 4.94 Impact Factor