scholarly journals TIP47 is a key effector for Rab9 localization

2006 ◽  
Vol 173 (6) ◽  
pp. 917-926 ◽  
Author(s):  
Dikran Aivazian ◽  
Ramon L. Serrano ◽  
Suzanne Pfeffer
Keyword(s):  
Steady State ◽  
Human Genome ◽  
Effector Proteins ◽  
Rab Gtpases ◽  
Interacting Protein ◽  
Cellular Concentration ◽  
Membrane Compartments ◽  
Effector Binding ◽  
Distinct Membrane

The human genome encodes ∼70 Rab GTPases that localize to the surfaces of distinct membrane compartments. To investigate the mechanism of Rab localization, chimeras containing heterologous Rab hypervariable domains were generated, and their ability to bind seven Rab effectors was quantified. Two chimeras could bind effectors for two distinctly localized Rabs; a Rab5/9 hybrid bound both Rab5 and Rab9 effectors, and a Rab1/9 hybrid bound to certain Rab1 and Rab9 effectors. These unusual chimeras permitted a test of the importance of effector binding for Rab localization. In both cases, changing the cellular concentration of a key Rab9 effector, which is called tail-interacting protein of 47 kD, moved a fraction of the proteins from their parental Rab localization to that of Rab9. Thus, relative concentrations of certain competing effectors could determine a chimera's localization. These data confirm the importance of effector interactions for Rab9 localization, and support a model in which effector proteins rely on Rabs as much as Rabs rely on effectors to achieve their correct steady state localizations.

A model for Rab GTPase localization

2005 ◽  
Vol 33 (4) ◽  
pp. 627-630 ◽  
Author(s):  
S. Pfeffer
Keyword(s):  
Steady State ◽  
Cellular Localization ◽  
Rab Gtpase ◽  
Rab Proteins ◽  
Rab Gtpases ◽  
Macromolecular Assemblies ◽  
Complex Interactions ◽  
Membrane Compartment ◽  
Membrane Bound ◽  
Distinct Membrane

The human genome encodes almost 70 Rab GTPases. These proteins are C-terminally geranylgeranylated and are localized to the surfaces of distinct membrane-bound compartments in eukaryotic cells. This mini review presents a working model for how Rabs achieve and maintain their steady-state localizations. Data from a number of laboratories suggest that Rabs participate in the generation of macromolecular assemblies that generate functional microdomains within a given membrane compartment. Our data suggest that these complex interactions are important for the cellular localization of Rab proteins at steady state.

Targeting Rab GTPases to distinct membrane compartments

2004 ◽  
Vol 5 (11) ◽  
pp. 886-896 ◽  
Author(s):  
Suzanne Pfeffer ◽  
Dikran Aivazian
Keyword(s):  
Rab Gtpases ◽  
Membrane Compartments ◽  
Distinct Membrane

scholarly journals Rab GTPase localization and Rab cascades in Golgi transport

2012 ◽  
Vol 40 (6) ◽  
pp. 1373-1377 ◽  
Author(s):  
Suzanne R. Pfeffer
Keyword(s):  
Effector Proteins ◽  
Membrane Microdomains ◽  
Rab Gtpase ◽  
Rab Gtpases ◽  
Nucleotide Exchange ◽  
Golgi Transport ◽  
Guanine Nucleotide Dissociation Inhibitor ◽  
Membrane Bound ◽  
Effector Binding ◽  
Endocytic Pathways

Rab GTPases are master regulators of membrane traffic. By binding to distinct sets of effector proteins, Rabs catalyse the formation of function-specifying membrane microdomains. They are delivered to membranes by a protein named GDI (guanine-nucleotide-dissociation inhibitor) and are stabilized there after nucleotide exchange by effector binding. In the present mini-review, I discuss what we know about how Rab GTPases are delivered to the correct membrane-bound compartments and how Rab GTPase cascades order Rabs within the secretory and endocytic pathways. Finally, I describe how Rab cascades may establish the distinct compartments of the Golgi complex to permit ordered processing, sorting and secretion of secretory cargoes.

scholarly journals Membrane association but not identity is required for LRRK2 activation and phosphorylation of Rab GTPases

2019 ◽  
Vol 218 (12) ◽  
pp. 4157-4170 ◽  
Author(s):  
Rachel C. Gomez ◽  
Paulina Wawro ◽  
Pawel Lis ◽  
Dario R. Alessi ◽  
Suzanne R. Pfeffer
Keyword(s):  
Plasma Membrane ◽  
Cell Biology ◽  
Family Members ◽  
Membrane Association ◽  
Rab Gtpases ◽  
Membrane Compartments ◽  
Membrane Bound ◽  
Lrrk2 Kinase ◽  
Familial Parkinson’S Disease ◽  
Distinct Membrane

LRRK2 kinase mutations cause familial Parkinson’s disease and increased phosphorylation of a subset of Rab GTPases. Rab29 recruits LRRK2 to the trans-Golgi and activates it there, yet some of LRRK2’s major Rab substrates are not on the Golgi. We sought to characterize the cell biology of LRRK2 activation. Unlike other Rab family members, we show that Rab29 binds nucleotide weakly, is poorly prenylated, and is not bound to GDI in the cytosol; nevertheless, Rab29 only activates LRRK2 when it is membrane bound and GTP bound. Mitochondrially anchored, GTP-bound Rab29 is both a LRRK2 substrate and activator, and it drives accumulation of active LRRK2 and phosphorylated Rab10 on mitochondria. Importantly, mitochondrially anchored LRRK2 is much less capable of phosphorylating plasma membrane–anchored Rab10 than soluble LRRK2. These data support a model in which LRRK2 associates with and dissociates from distinct membrane compartments to phosphorylate Rab substrates; if anchored, LRRK2 can modify misdelivered Rab substrates that then become trapped there because GDI cannot retrieve them.

scholarly journals Structures ofDrosophila melanogasterRab2 and Rab3 bound to GMPPNP

2015 ◽  
Vol 71 (1) ◽  
pp. 34-40 ◽  
Author(s):  
Jennifer A. Lardong ◽  
Jan H. Driller ◽  
Harald Depner ◽  
Christoph Weise ◽  
Astrid Petzoldt ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Intracellular Trafficking ◽  
Large Family ◽  
Cysteine Residue ◽  
Effector Proteins ◽  
Rab Proteins ◽  
Rab Gtpases ◽  
Ras Proteins ◽  
Transport Vesicles ◽  
Switch Regions

Rab GTPases belong to the large family of Ras proteins. They act as key regulators of membrane organization and intracellular trafficking. Functionally, they act as switches. In the active GTP-bound form they can bind to effector proteins to facilitate the delivery of transport vesicles. Upon stimulation, the GTP is hydrolyzed and the Rab proteins undergo conformational changes in their switch regions. This study focuses on Rab2 and Rab3 fromDrosophila melanogaster. Whereas Rab2 is involved in vesicle transport between the Golgi and the endoplasmatic reticulum, Rab3 is a key player in exocytosis, and in the synapse it is involved in the assembly of the presynaptic active zone. Here, high-resolution crystal structures of Rab2 and Rab3 in complex with GMPPNP and Mg2+are presented. In the structure of Rab3 a modified cysteine residue is observed with an enigmatic electron density attached to its thiol function.

scholarly journals Phytophthora effector targets a novel component of small RNA pathway in plants to promote infection

2015 ◽  
Vol 112 (18) ◽  
pp. 5850-5855 ◽  
Author(s):  
Yongli Qiao ◽  
Jinxia Shi ◽  
Yi Zhai ◽  
Yingnan Hou ◽  
Wenbo Ma
Keyword(s):  
Small Rna ◽  
Rna Silencing ◽  
Effector Proteins ◽  
Helicase Domain ◽  
Small Interfering Rnas ◽  
Developmental Defects ◽  
Interacting Protein ◽  
Homologous Genes ◽  
Unidentified Component ◽  
Virulence Target

A broad range of parasites rely on the functions of effector proteins to subvert host immune response and facilitate disease development. The notorious Phytophthora pathogens evolved effectors with RNA silencing suppression activity to promote infection in plant hosts. Here we report that the Phytophthora Suppressor of RNA Silencing 1 (PSR1) can bind to an evolutionarily conserved nuclear protein containing the aspartate–glutamate–alanine–histidine-box RNA helicase domain in plants. This protein, designated PSR1-Interacting Protein 1 (PINP1), regulates the accumulation of both microRNAs and endogenous small interfering RNAs in Arabidopsis. A null mutation of PINP1 causes embryonic lethality, and silencing of PINP1 leads to developmental defects and hypersusceptibility to Phytophthora infection. These phenotypes are reminiscent of transgenic plants expressing PSR1, supporting PINP1 as a direct virulence target of PSR1. We further demonstrate that the localization of the Dicer-like 1 protein complex is impaired in the nucleus of PINP1-silenced or PSR1-expressing cells, indicating that PINP1 may facilitate small RNA processing by affecting the assembly of dicing complexes. A similar function of PINP1 homologous genes in development and immunity was also observed in Nicotiana benthamiana. These findings highlight PINP1 as a previously unidentified component of RNA silencing that regulates distinct classes of small RNAs in plants. Importantly, Phytophthora has evolved effectors to target PINP1 in order to promote infection.

scholarly journals Functional characterization of the human tRNA methyltransferases TRMT10A and TRMT10B

2020 ◽  
Vol 48 (11) ◽  
pp. 6157-6169 ◽  
Author(s):  
Elisa Vilardo ◽  
Fabian Amman ◽  
Ursula Toth ◽  
Annika Kotter ◽  
Mark Helm ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Steady State ◽  
Human Genome ◽  
Functional Significance ◽  
Functional Characterization ◽  
Trna Methyltransferase ◽  
Steady State Levels

Abstract The TRM10 family of methyltransferases is responsible for the N1-methylation of purines at position 9 of tRNAs in Archaea and Eukarya. The human genome encodes three TRM10-type enzymes, of which only the mitochondrial TRMT10C was previously characterized in detail, whereas the functional significance of the two presumably nuclear enzymes TRMT10A and TRMT10B remained unexplained. Here we show that TRMT10A is m1G9-specific and methylates a subset of nuclear-encoded tRNAs, whilst TRMT10B is the first m1A9-specific tRNA methyltransferase found in eukaryotes and is responsible for the modification of a single nuclear-encoded tRNA. Furthermore, we show that the lack of G9 methylation causes a decrease in the steady-state levels of the initiator tRNAiMet-CAT and an alteration in its further post-transcriptional modification. Our work finally clarifies the function of TRMT10A and TRMT10B in vivo and provides evidence that the loss of TRMT10A affects the pool of cytosolic tRNAs required for protein synthesis.

scholarly journals Spatiotemporal organization and protein dynamics involved in regulated exocytosis of MMP-9 in breast cancer cells

2019 ◽  
Vol 151 (12) ◽  
pp. 1386-1403 ◽  
Author(s):  
Dominique C. Stephens ◽  
Nicole Osunsanmi ◽  
Kem A. Sochacki ◽  
Tyrel W. Powell ◽  
Justin W. Taraska ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Secretory Vesicle ◽  
Effector Proteins ◽  
Cellular Transport ◽  
Secretory Vesicles ◽  
Rab Gtpases ◽  
Regulated Exocytosis ◽  
Endocytic Proteins

Altered regulation of exocytosis is an important mechanism controlling many diseases, including cancer. Defects in exocytosis have been implicated in many cancer cell types and are generally attributed to mutations in cellular transport, trafficking, and assembly of machinery necessary for exocytosis of secretory vesicle cargo. In these cancers, up-regulation of trafficking and secretion of matrix metalloproteinase-9 (MMP-9), a proteolytic enzyme, is responsible for degrading the extracellular matrix, a necessary step in tumor progression. Using TIRF microscopy, we identified proteins associated with secretory vesicles containing MMP-9 and imaged the local dynamics of these proteins at fusion sites during regulated exocytosis of MMP-9 from MCF-7 breast cancer cells. We found that many regulators of exocytosis, including several Rab GTPases, Rab effector proteins, and SNARE/SNARE modulator proteins, are stably assembled on docked secretory vesicles before exocytosis. At the moment of fusion, many of these components are quickly lost from the vesicle, while several endocytic proteins and lipids are simultaneously recruited to exocytic sites at precisely that moment. Our findings provide insight into the dynamic behavior of key core exocytic proteins, accessory proteins, lipids, and some endocytic proteins at single sites of secretory vesicle fusion in breast cancer cells.

DPP10 is an inactivation modulatory protein of Kv4.3 and Kv1.4

2006 ◽  
Vol 291 (5) ◽  
pp. C966-C976 ◽  
Author(s):  
Hong-Ling Li ◽  
Yu-Jie Qu ◽  
Yi Chun Lu ◽  
Vladimir E. Bondarenko ◽  
Shimin Wang ◽  
...  
Keyword(s):  
Steady State ◽  
Interacting Protein ◽  
Voltage Gated ◽  
Time To Peak ◽  
Kv Channel ◽  
Closed State ◽  
Channel Inactivation ◽  
Recovery From Inactivation

Voltage-gated K+ channels exist in vivo as multiprotein complexes made up of pore-forming and ancillary subunits. To further our understanding of the role of a dipeptidyl peptidase-related ancillary subunit, DPP10, we expressed it with Kv4.3 and Kv1.4, two channels responsible for fast-inactivating K+ currents. Previously, DPP10 has been shown to effect Kv4 channels. However, Kv1.4, when expressed with DPP10, showed many of the same effects as Kv4.3, such as faster time to peak current and negative shifts in the half-inactivation potential of steady-state activation and inactivation. The exception was recovery from inactivation, which is slowed by DPP10. DPP10 expressed with Kv4.3 caused negative shifts in both steady-state activation and inactivation of Kv4.3, but no significant shifts were detected when DPP10 was expressed with Kv4.3 + KChIP2b (Kv channel interacting protein). DPP10 and KChIP2b had different effects on closed-state inactivation. At −60 mV, KChIP2b nearly abolishes closed-state inactivation in Kv4.3, whereas it developed to a much greater extent in the presence of DPP10. Finally, expression of a DPP10 mutant consisting of its transmembrane and cytoplasmic 58 amino acids resulted in effects on Kv4.3 gating that were nearly identical to those of wild-type DPP10. These data show that DPP10 and KChIP2b both modulate Kv4.3 inactivation but that their primary effects are on different inactivation states. Thus DPP10 may be a general modulator of voltage-gated K+ channel inactivation; understanding its mechanism of action may lead to deeper understanding of the inactivation of a broad range of K+ channels.

scholarly journals A role for Yip1p in COPII vesicle biogenesis

2003 ◽  
Vol 163 (1) ◽  
pp. 57-69 ◽  
Author(s):  
Matthew Heidtman ◽  
Catherine Z. Chen ◽  
Ruth N. Collins ◽  
Charles Barlowe
Keyword(s):  
Secretory Pathway ◽  
Protein Transport ◽  
Genetic Interaction ◽  
Temperature Sensitive ◽  
Rab Gtpases ◽  
Direct Role ◽  
Interacting Protein ◽  
Copii Vesicle ◽  
Vesicle Biogenesis

Yeast Ypt1p-interacting protein (Yip1p) belongs to a conserved family of transmembrane proteins that interact with Rab GTPases. We encountered Yip1p as a constituent of ER-derived transport vesicles, leading us to hypothesize a direct role for this protein in transport through the early secretory pathway. Using a cell-free assay that recapitulates protein transport from the ER to the Golgi complex, we find that affinity-purified antibodies directed against the hydrophilic amino terminus of Yip1p potently inhibit transport. Surprisingly, inhibition is specific to the COPII-dependent budding stage. In support of this in vitro observation, strains bearing the temperature-sensitive yip1-4 allele accumulate ER membranes at a nonpermissive temperature, with no apparent accumulation of vesicle intermediates. Genetic interaction analyses of the yip1-4 mutation corroborate a function in ER budding. Finally, ordering experiments show that preincubation of ER membranes with COPII proteins decreases sensitivity to anti-Yip1p antibodies, indicating an early requirement for Yip1p in vesicle formation. We propose that Yip1p has a previously unappreciated role in COPII vesicle biogenesis.

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