scholarly journals The ciliary targeting of membrane proteins by a ternary complex comprising transportin1, Rab8 and the ciliary targeting signal

2016 ◽  
Author(s):  
Viswanadh Madugula ◽  
Lei Lu
Keyword(s):  
Ternary Complex ◽  
Molecular Level ◽  
Nucleotide Exchange ◽  
Lateral Transport ◽  
Retinol Dehydrogenase ◽  
Transport Pathway ◽  
Binding Motifs ◽  
Ciliary Membrane ◽  
Guanine Nucleotide Exchange ◽  
Targeting Signal

AbstractThe sensory functions of cilia are dependent on the enrichment of ciliary resident proteins. While it is known that ciliary targeting signals (CTSs) specifically target ciliary proteins to cilia, it is still unclear how CTSs facilitate the entry and retention of ciliary residents at the molecular level. We found that non-ciliary membrane reporters can passively diffuse to cilia via the lateral transport pathway and the translocation of membrane reporters through the ciliary diffusion barrier is facilitated by importin binding motifs/domains. Screening known CTSs of ciliary membrane residents uncovered that fibrocystin, photoreceptor retinol dehydrogenase, rhodopsin and retinitis pigmentosa 2 interact with transportin1 (TNPO1) via previously identified CTSs. We further discovered that a novel ternary complex, comprising TNPO1, Rab8 and CTS, can assemble or disassemble under the guanine nucleotide exchange of Rab8. Our study suggests a novel mechanism in which TNPO1/Rab8/CTS complex mediates selective entry and retention of cargos within cilia.

scholarly journals Ubiquitin binding by the CUE domain promotes endosomal localization of the Rab5 GEF Vps9

2015 ◽  
Vol 26 (7) ◽  
pp. 1345-1356 ◽  
Author(s):  
Tess Shideler ◽  
Daniel P. Nickerson ◽  
Alexey J. Merz ◽  
Greg Odorizzi
Keyword(s):  
Membrane Trafficking ◽  
Guanine Nucleotide Exchange Factors ◽  
Multivesicular Bodies ◽  
Nucleotide Exchange ◽  
Binding Motifs ◽  
Endosomal Sorting ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factors ◽  
Ubiquitin Binding ◽  
Cue Domain

Vps9 and Muk1 are guanine nucleotide exchange factors (GEFs) in Saccharomyces cerevisiae that regulate membrane trafficking in the endolysosomal pathway by activating Rab5 GTPases. We show that Vps9 is the primary Rab5 GEF required for biogenesis of late endosomal multivesicular bodies (MVBs). However, only Vps9 (but not Muk1) is required for the formation of aberrant class E compartments that arise upon dysfunction of endosomal sorting complexes required for transport (ESCRTs). ESCRT dysfunction causes ubiquitinated transmembrane proteins to accumulate at endosomes, and we demonstrate that endosomal recruitment of Vps9 is promoted by its ubiquitin-binding CUE domain. Muk1 lacks ubiquitin-binding motifs, but its fusion to the Vps9 CUE domain allows Muk1 to rescue endosome morphology, cargo trafficking, and cellular stress-tolerance phenotypes that result from loss of Vps9 function. These results indicate that ubiquitin binding by the CUE domain promotes Vps9 function in endolysosomal membrane trafficking via promotion of localization.

scholarly journals RAPGEF5 regulates nuclear translocation of β-catenin

2017 ◽  
Author(s):  
John N. Griffin ◽  
Florencia del Viso ◽  
Anna R. Duncan ◽  
Andrew Robson ◽  
Saurabh Kulkarni ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Nuclear Translocation ◽  
Nuclear Transport ◽  
Nucleotide Exchange ◽  
Transport Pathway ◽  
Disease States ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Pathway Activation ◽  
Intensive Investigation

SUMMARYCanonical Wnt signaling coordinates many critical aspects of embryonic development, while dysregulated Wnt signaling contributes to common diseases, including congenital malformations and cancer. The nuclear localization of β-catenin is the defining step in pathway activation. However, despite intensive investigation, the mechanisms regulating β-catenin nuclear transport remain undefined. In a patient with congenital heart disease and heterotaxy, a disorder of left-right patterning, we previously identified the guanine nucleotide exchange factor, RAPGEF5. Here, we demonstrate that RAPGEF5 regulates left-right patterning via Wnt signaling. In particular, RAPGEF5, regulates the nuclear translocation of β-catenin independently of both β-catenin cytoplasmic stabilization and the importin β1/Ran mediated transport system. We propose a model whereby RAPGEF5 activates the nuclear GTPases, Rap1/2, to facilitate the nuclear transport of β-catenin, defining a parallel nuclear transport pathway to Ran. Our results suggest new targets for modulating Wnt signaling in disease states.

scholarly journals Lateral transport of Smoothened from the plasma membrane to the membrane of the cilium

2009 ◽  
Vol 187 (3) ◽  
pp. 365-374 ◽  
Author(s):  
Ljiljana Milenkovic ◽  
Matthew P. Scott ◽  
Rajat Rohatgi
Keyword(s):  
Plasma Membrane ◽  
Protein Trafficking ◽  
Primary Cilia ◽  
Protein Transport ◽  
Transmembrane Protein ◽  
Signaling Proteins ◽  
Lateral Transport ◽  
Transport Pathway ◽  
Ciliary Membrane ◽  
Specific Proteins

The function of primary cilia depends critically on the localization of specific proteins in the ciliary membrane. A major challenge in the field is to understand protein trafficking to cilia. The Hedgehog (Hh) pathway protein Smoothened (Smo), a 7-pass transmembrane protein, moves to cilia when a ligand is received. Using microscopy-based pulse-chase analysis, we find that Smo moves through a lateral transport pathway from the plasma membrane to the ciliary membrane. Lateral movement, either via diffusion or active transport, is quite distinct from currently studied pathways of ciliary protein transport in mammals, which emphasize directed trafficking of Golgi-derived vesicles to the base of the cilium. We anticipate that this alternative route will be used by other signaling proteins that function at cilia. The path taken by Smo may allow novel strategies for modulation of Hh signaling in cancer and regeneration.

scholarly journals ARL3 and ARL13B GTPases participate in distinct steps of INPP5E targeting to the ciliary membrane

Biology Open ◽  
2021 ◽  
Vol 10 (9) ◽  
Author(s):  
Sayaka Fujisawa ◽  
Hantian Qiu ◽  
Shohei Nozaki ◽  
Shuhei Chiba ◽  
Yohei Katoh ◽  
...  
Keyword(s):  
Guanine Nucleotide Exchange Factor ◽  
Membrane Localization ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Cellular Functions ◽  
Ciliary Membrane ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Direct Binding ◽  
Ciliary Localization

ABSTRACT INPP5E, a phosphoinositide 5-phosphatase, localizes on the ciliary membrane via its C-terminal prenyl moiety, and maintains the distinct ciliary phosphoinositide composition. The ARL3 GTPase contributes to the ciliary membrane localization of INPP5E by stimulating the release of PDE6D bound to prenylated INPP5E. Another GTPase, ARL13B, which is localized on the ciliary membrane, contributes to the ciliary membrane retention of INPP5E by directly binding to its ciliary targeting sequence. However, as ARL13B was shown to act as a guanine nucleotide exchange factor (GEF) for ARL3, it is also possible that ARL13B indirectly mediates the ciliary INPP5E localization via activating ARL3. We here show that INPP5E is delocalized from cilia in both ARL3-knockout (KO) and ARL13B-KO cells. However, some of the abnormal phenotypes were different between these KO cells, while others were found to be common, indicating the parallel roles of ARL3 and ARL13B, at least concerning some cellular functions. For several variants of ARL13B, their ability to interact with INPP5E, rather than their ability as an ARL3-GEF, was associated with whether they could rescue the ciliary localization of INPP5E in ARL13B-KO cells. These observations together indicate that ARL13B determines the ciliary localization of INPP5E, mainly by its direct binding to INPP5E.

scholarly journals Fusel Alcohols Regulate Translation Initiation by Inhibiting eIF2B to Reduce Ternary Complex in a Mechanism That May Involve Altering the Integrity and Dynamics of the eIF2B Body

2010 ◽  
Vol 21 (13) ◽  
pp. 2202-2216 ◽  
Author(s):  
Eleanor J. Taylor ◽  
Susan G. Campbell ◽  
Christian D. Griffiths ◽  
Peter J. Reid ◽  
John W. Slaven ◽  
...  
Keyword(s):  
Ternary Complex ◽  
Tiling Array ◽  
Dependent Manner ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Alcohol Sensitivity ◽  
Critical Determinant ◽  
Cytoplasmic Body ◽  
Guanine Nucleotide Exchange ◽  
Fusel Alcohols

Recycling of eIF2-GDP to the GTP-bound form constitutes a core essential, regulated step in eukaryotic translation. This reaction is mediated by eIF2B, a heteropentameric factor with important links to human disease. eIF2 in the GTP-bound form binds to methionyl initiator tRNA to form a ternary complex, and the levels of this ternary complex can be a critical determinant of the rate of protein synthesis. Here we show that eIF2B serves as the target for translation inhibition by various fusel alcohols in yeast. Fusel alcohols are endpoint metabolites from amino acid catabolism, which signal nitrogen scarcity. We show that the inhibition of eIF2B leads to reduced ternary complex levels and that different eIF2B subunit mutants alter fusel alcohol sensitivity. A DNA tiling array strategy was developed that overcame difficulties in the identification of these mutants where the phenotypic distinctions were too subtle for classical complementation cloning. Fusel alcohols also lead to eIF2α dephosphorylation in a Sit4p-dependent manner. In yeast, eIF2B occupies a large cytoplasmic body where guanine nucleotide exchange on eIF2 can occur and be regulated. Fusel alcohols impact on both the movement and dynamics of this 2B body. Overall, these results confirm that the guanine nucleotide exchange factor, eIF2B, is targeted by fusel alcohols. Moreover, they highlight a potential connection between the movement or integrity of the 2B body and eIF2B regulation.

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