Rab5 is necessary for the biogenesis of the endolysosomal system in vivo

Zeigerer, Anja; Gilleron, Jérôme; Bogorad, Roman L.; Marsico, Giovanni; Nonaka, Hiroshi; Seifert, Stefan; Epstein-Barash, Hila; Kuchimanchi, Satya; Peng, Chang G.; Ruda, Vera M.; Conte-Zerial, Perla Del; Hengstler, Jan G.; Kalaidzidis, Yannis; Koteliansky, Victor; Zerial, Marino

doi:10.1038/nature11133

Cited by 328 publications

(375 citation statements)

References 49 publications

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“…We also found eHAV to be enriched for RAB5C and RAB7A (Fig. 2), and isoforms of both of these RAB GTPases are known to play prominent roles in endosomal trafficking and exosome biogenesis (12,26).…”

Section: Discussionmentioning

confidence: 87%

Protein composition of the hepatitis A virus quasi-envelope

McKnight

Xie

González‐López

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

124

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The Picornaviridae are a diverse family of RNA viruses including many pathogens of medical and veterinary importance. Classically considered "nonenveloped," recent studies show that some picornaviruses, notably hepatitis A virus (HAV; genus Hepatovirus) and some members of the Enterovirus genus, are released from cells nonlytically in membranous vesicles. To better understand the biogenesis of quasienveloped HAV (eHAV) virions, we conducted a quantitative proteomics analysis of eHAV purified from cell-culture supernatant fluids by isopycnic ultracentrifugation. Amino acid-coded mass tagging (AACT) with stable isotopes followed by tandem mass spectrometry sequencing and AACT quantitation of peptides provided unambiguous identification of proteins associated with eHAV versus unrelated extracellular vesicles with similar buoyant density. Multiple peptides were identified from HAV capsid proteins (53.7% coverage), but none from nonstructural proteins, indicating capsids are packaged as cargo into eHAV vesicles via a highly specific sorting process. Other eHAVassociated proteins (n = 105) were significantly enriched for components of the endolysosomal system (>60%, P < 0.001) and included many common exosome-associated proteins such as the tetraspanin CD9 and dipeptidyl peptidase 4 (DPP4) along with multiple endosomal sorting complex required for transport III (ESCRT-III)-associated proteins. Immunoprecipitation confirmed that DPP4 is displayed on the surface of eHAV produced in cell culture or present in sera from humans with acute hepatitis A. No LC3-related peptides were identified by mass spectrometry. RNAi depletion studies confirmed that ESCRT-III proteins, particularly CHMP2A, function in eHAV biogenesis. In addition to identifying surface markers of eHAV vesicles, the results support an exosome-like mechanism of eHAV egress involving endosomal budding of HAV capsids into multivesicular bodies.exosome | multivesicular body | ESCRT | extracellular vesicle | picornavirus T he Picornaviridae are a large and diverse family of positivestrand RNA viruses that include numerous pathogens (1). Classically considered "nonenveloped," these viruses package their single-stranded genomes within stable icosahedral protein capsids that are released from cells following cell lysis. However, recent work has revealed that several picornaviruses gain egress from cells in a nonlytic fashion within the lumen of extracellular vesicles shed from the cell. Most notably, hepatitis A virus (HAV; genus Hepatovirus), an important cause of enterically transmitted hepatitis in humans, replicates without cytopathic effect and is released from cells as membrane-wrapped, "quasienveloped" (eHAV) virions similar in size and density to exosomes (2). eHAV virions have been identified in sera collected from persons with acute hepatitis A, as well as in supernatant fluids of infected cell cultures. These virions are completely cloaked in host-derived membranes that protect the capsid from neutralizing antibody until the quasi-envelope is degraded wi...

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Section: Discussionmentioning

confidence: 87%

Protein composition of the hepatitis A virus quasi-envelope

McKnight

Xie

González‐López

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

124

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“…Briefly, activated Rab5 drives NSF-primed endosomes to tether and dock with each other via oligomerized EEA1, syntaxin12/13 [32] and possibly the Mon1/CCZ1 complex [36] for subsequent homotypic fusion to generate a fused large endosome. Proteins in the fused large endosomes are either recycled back to the plasma membrane or transported to the trans-Golgi network or lysosome for destruction [37]. We used RNA interference technology [38] to knock down Numb and Numblike to characterize their roles in substrate trafficking.…”

Section: Introductionmentioning

confidence: 99%

Numb regulates vesicular docking for homotypic fusion of early endosomes via membrane recruitment of Mon1b

Shao

Liu

et al. 2016

Cell Res

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Numb is an endocytic protein that plays crucial roles in diverse cellular processes such as asymmetric cell division, cell migration and differentiation. However, the molecular mechanism by which Numb regulates endocytic trafficking is poorly understood. Here, we demonstrate that Numb is a docking regulator for homotypic fusion of early endosomes (EEs). Numb depletion causes clustered but unfused EEs, which can be rescued by overexpressing cytosolic Numb 65 and Numb 71 but not plasma membrane-attached Numb 66 or Numb 72. Time-lapse analysis reveals that paired vesicles tend to tether but not fuse with each other in the absence of Numb. We further show that Numb binds to another docking regulator, Mon1b, and is required for the recruitment of cytosolic Mon1b to the EE membrane. Consistent with this, deletion of Mon1b causes similar defects in EE fusion. Our study thus identifies a novel mechanism by which Numb regulates endocytic sorting by mediating EE fusion.

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“…Titration of Rab5 showed that the endocytic system was resilient against depletion of Rab5 and collapsed only when depletion exceeded approximately 80 % of control levels. Comparison of the experimental data with the model simulations supported the scenario whereby the endosomes are depleted, but those remaining maintain a constant level of molecular machinery (Zeigerer et al 2012). The mathematical model also explains why the relatively wide variations in Rab5 expression have only a minor influence on the numbers of endosomes, but the system collapses as soon as critical thresholds are exceeded.…”

mentioning

confidence: 60%

“…The importance of Rab5 in endosome biogenesis was tested using a combination of mathematical model and experimental validation in the mouse livers as model system. To study the dependency of the endosome number on the levels of Rab5, a mathematical model was formulated to predict the consequences of the loss of Rab5 for the number of early endosomes (Zeigerer et al 2012). Different scenarios were considered.…”

mentioning

confidence: 99%

The virtual liver: state of the art and future perspectives

et al. 2014

Self Cite

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clarifying the underlying principles. The mathematical models formalize the relationship between individual components, test their interactions in a virtual setting and may even simulate influences that are (still) difficult to analyse experimentally. In recent years, model simulations have been instrumental to elucidate mechanisms and principles that were not accessible by traditional approaches. To promote systems biology research in the field of the liver with the aim to gain a better understanding of the basic mechanisms of liver function as well as key principles of liver Developments over the past two decades have improved our ability to obtain comprehensive and quantitative data, for example, by genome-wide analysis of gene expression, proteomics, lipidomics and metabolomics. Moreover, both imaging and image analysis have been improved which offers new possibilities to quantify the three-dimensional organization of cells and tissues. However, research in disease pathogenesis is often hampered by the difficulty to understand the complex, time-resolved interplay among numerous components. Here, mathematical modelling helps

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Rab5 is necessary for the biogenesis of the endolysosomal system in vivo

Cited by 328 publications

References 49 publications

Protein composition of the hepatitis A virus quasi-envelope

Protein composition of the hepatitis A virus quasi-envelope

Numb regulates vesicular docking for homotypic fusion of early endosomes via membrane recruitment of Mon1b

The virtual liver: state of the art and future perspectives

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