Article

Rabenosyn-5, a Novel Rab5 Effector, Is Complexed with Hvps45 and Recruited to Endosomes through a Fyve Finger Domain

Max-Planck-Institute for Molecular Cell Biology and Genetics, 01307 Dresden, Germany.
The Journal of Cell Biology (Impact Factor: 9.83). 11/2000; 151(3):601-12.
Source: PubMed

ABSTRACT

Rab5 regulates endocytic membrane traffic by specifically recruiting cytosolic effector proteins to their site of action on early endosomal membranes. We have characterized a new Rab5 effector complex involved in endosomal fusion events. This complex includes a novel protein, Rabenosyn-5, which, like the previously characterized Rab5 effector early endosome antigen 1 (EEA1), contains an FYVE finger domain and is recruited in a phosphatidylinositol-3-kinase-dependent fashion to early endosomes. Rabenosyn-5 is complexed to the Sec1-like protein hVPS45. hVPS45 does not interact directly with Rab5, therefore Rabenosyn-5 serves as a molecular link between hVPS45 and the Rab5 GTPase. This property suggests that Rabenosyn-5 is a closer mammalian functional homologue of yeast Vac1p than EEA1. Furthermore, although both EEA1 and Rabenosyn-5 are required for early endosomal fusion, only overexpression of Rabenosyn-5 inhibits cathepsin D processing, suggesting that the two proteins play distinct roles in endosomal trafficking. We propose that Rab5-dependent formation of membrane domains enriched in phosphatidylinositol-3-phosphate has evolved as a mechanism for the recruitment of multiple effector proteins to mammalian early endosomes, and that these domains are multifunctional, depending on the differing activities of the effector proteins recruited.

Download full-text

Full-text

Available from: Bernard Hoflack
  • Source
    • "e found that DRG2 interacted with Rab5 , indicating that interaction between DRG2 and Rab5 mediates the localization of DRG2 to Rab5 endosomes . Rab5 recruits PI - 3 - kinases in a GTP - dependent manner , resulting in localized synthesis of PI - 3P ( Christoforidis et al . , 1999b ) . In turn , EEA1 ( Simonsen et al . , 1998 ) and Rabenosyn - 5 ( Nielsen et al . , 2000 ) , Rab5 effectors , bind PI3P through the interaction of a FYVE domain in a PI3K - dependent manner . In this study , we also found that , while DRG2 did not colocalize with Rabenosyn - 5 on endosomes , it colocalized and interacted with EEA1 on endosomes . In addition , colocalization and interaction of DRG2 with EEA1 on endosomes wer"
    [Show abstract] [Hide abstract]
    ABSTRACT: The small GTPase Rab5 regulates the early endocytic pathway of transferrin (Tfn) and Rab5 deactivation is required for Tfn recycling. Rab5 deactivation is achieved by RabGAP5, a GTPase-activating protein, on the endosomes. Here, we report that recruitment of RabGAP5 is insufficient to deactivate Rab5 and that developmentally regulated GTP-binding protein 2 (DRG2) is required for Rab5 deactivation and Tfn recycling. DRG2 was associated with phosphatidylinositol 3-phosphate (PI(3)P)-containing endosomes. It colocalized and interacted with EEA1 and Rab5 on endosomes in a phosphatidylinositol-3-kinase-dependent manner. DRG2 depletion did not affect Tfn uptake and recruitment of RabGAP5 and Rac1 to Rab5 endosomes. However, it resulted in impairment of interaction between Rab5 and RabGAP5, Rab5 deactivation on endosomes, and Tfn recycling. Ectopic expression of shRNA-resistant DRG2 rescued Tfn recycling in DRG2 depleted cells. Our results demonstrate DRG2 as an endosomal protein and a key regulator for Rab5 deactivation and Tfn recycling.
    Preview · Article · Nov 2015 · Molecular biology of the cell
  • Source
    • "motor protein complexes). Rab5 has been shown to interact with at least 20 identified proteins through the use of Rab5-GST fusion proteins in affinity chromatography preloaded with either GTP or GDP and after binding to effector molecules, eluted with GDP or GTP, respectively[14]: Rabaptin-5[15], RIN3[16], Rabex-5 GEF[17], Rabenosyn-5[18],[19], Rabankyrin-5[20], EEA1[21], PI-3 kinases hVPS34 and p85α-p110β[22],[23], PI-4, PI-5 phosphatases[24], kinesin type motor proteins for microtubule dependent movement[25], APPL1 and APPL2[26], syntaxin-13[27], GGAs[28], Huntingtin and HAP40[29]. The nature of the function of Rab proteins (cycling between the “on” and “off” state) may make for only transient interactions with effectors, which from an experimental perspective, are difficult to trap. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Abnormal intracellular accumulation or transport of lipids contributes greatly to the pathogenesis of human diseases. In the liver, excess accumulation of triacylglycerol (TG) leads to fatty liver disease encompassing steatosis, steatohepatitis and fibrosis. This places individuals at risk of developing cirrhosis, hepatocellular carcinoma or hepatic decompensation and also contributes to the emergence of insulin resistance and dyslipidemias affecting many other organs. Excessive accumulation of TG in adipose tissue contributes to insulin resistance as well as to the release of cytokines attracting leucocytes leading to a pro-inflammatory state. Pathological accumulation of cholesteryl ester (CE) in macrophages in the arterial wall is the progenitor of atherosclerotic plaques and heart disease. Overconsumption of dietary fat, cholesterol and carbohydrates explains why these diseases are on the increase yet offers few clues for how to prevent or treat individuals. Dietary regimes have proven futile and barring surgery, no realistic alternatives are at hand as effective drugs are few and not without side effects. Overweight and obesity-related diseases are no longer restricted to the developed world and as such, constitute a global problem. Development of new drugs and treatment strategies are a priority yet requires as a first step, elucidation of the molecular pathophysiology underlying each associated disease state. The lipid droplet (LD), an up to now overlooked intracellular organelle, appears at the heart of each pathophysiology linking key regulatory and metabolic processes as well as constituting the site of storage of both TGs and CEs. As the molecular machinery and mechanisms of LDs of each cell type are being elucidated, regulatory proteins used to control various cellular processes are emerging. Of these and the subject of this review, small GTPases belonging to the Rab protein family appear as important molecular switches used in the regulation of the intracellular trafficking and storage of lipids.
    Preview · Article · May 2014
  • Source
    • "Interestingly, only the combination of both tethers led to significant fusion , suggesting that one cannot substitute for the other. These two tethers also coordinate Rab function with SNARE assembly on early endosomes, because rabenosyn-5 binds the Sec1/Munc18-like Vps45 protein (Nielsen et al. 2000; Morrison et al. 2008), and EEA1 binds "
    [Show abstract] [Hide abstract]
    ABSTRACT: Organelles within the endomembrane system are connected via vesicle flux. Along the endocytic pathway, endosomes are among the most versatile organelles. They sort cargo through tubular protrusions for recycling or through intraluminal vesicles for degradation. Sorting involves numerous machineries, which mediate fission of endosomal transport intermediates and fusion with other endosomes or eventually with lysosomes. Here we review the recent advances in our understanding of these processes with a particular focus on the Rab GTPases, tethering factors, and retromer. The cytoskeleton has also been recently recognized as a central player in membrane dynamics of endosomes, and this review covers the regulation of the machineries that govern the formation of branched actin networks through the WASH and Arp2/3 complexes in relation with cargo recycling and endosomal fission.
    Full-text · Article · Mar 2014 · Cold Spring Harbor perspectives in biology
Show more