Sequential Roles for Phosphatidylinositol 3-Phosphate and Rab5 in Tethering and Fusion of Early Endosomes via Their Interaction with EEA1
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Early endosome antigen 1 (EEA1) is a 170-kDa polypeptide required for endosome fusion in mammalian cells. The COOH terminus of EEA1 contains a FYVE domain that interacts specifically with phosphatidylinositol 3-phosphate (PtdIns-3-P) and a Rab5 GTPase binding region adjacent to the FYVE domain. The dual interaction of EEA1 with both PtdIns-3-P and Rab5 has been hypothesized to provide the specificity required to target EEA1 to early endosomes. To test this hypothesis, we generated truncated (amino acids 1277--1411) and full-length EEA1 constructs containing point mutations in the COOH terminus that impair Rab5 but not PtdIns-3-P binding. These constructs localized to endosomes in intact cells as efficiently as their wild-type counterparts. Furthermore, overexpression of the truncated constructs, both wild-type and mutated, impaired the function of endogenous EEA1 resulting in the accumulation of small, untethered endosomes. These results suggest that association with Rab5 is not necessary for the initial binding and tethering functions of EEA1. A role for Rab5 binding was revealed, however, upon comparison of endosomes in cells expressing full-length wild-type or mutated EEA1. The mutant full-length EEA1 caused the accumulation of endosome clusters and suppressed the enlargement of endosomes caused by a persistently active form of Rab5 (Rab5Q79L). In contrast, expression of wild-type EEA1 with Rab5Q79L enhanced this enlargement. Thus, endosome tethering depends on the interaction of EEA1 with PtdIns-3-P, and its interaction with Rab5 appears to regulate subsequent fusion.
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Rab5A is a marker of early phagosomes and facilitates downstream maturation by recruiting EEA1, which then binds NSF, Rabex5, and Rabaptin-5 and subsequently syntaxin13 to form a fusogenic complex with endosomes.
“The small GTPase Rab5A is a marker of early phagosomes and facilitates downstream maturation by recruiting early endosomal antigen 1 (EEA1), which then binds NSF, Rabex5, and Rabaptin-5 and subsequently syntaxin13 to form a fusogenic complex with endosomes (22–24).”
From Macrophage defense mechanisms against intracellular bacteria (2015) · cites this paper as [24]
Cells expressing the active GTP-bound form of RAB5, RAB5 Q79L, show an enlargement of endosomes that is commonly used to facilitate discrimination between the limiting membrane and the lumen of these organelles.
“Cells expressing the active (GTP-bound) form of RAB5, RAB5 Q79L , show an enlargement of endosomes, which is commonly used to facilitate the discrimination between the limiting membrane and the lumen of these organelles. 44,45”
From NRBF2 is involved in the autophagic degradation process of APP-CTFs in Alzheimer disease models (2017) · cites this paper as [45]
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- Authors
- Lawe, D. C., Chawla, A., Merithew, E., Dumas, J., Carrington, W., Fogarty, K., Lifshitz, L., Tuft, R., Lambright, D., & Corvera, S.
- Affiliations
- University of Massachusetts Chan Medical School (US)
- Corresponding author
- Corvera, Silvia
- Venue
- Journal of Biological Chemistry
- Publisher
- Elsevier BV
- Published
- 2002-03
- Volume
- 277
- Issue
- 10
- Pages
- 8611-8617
- ISSN
- 0021-9258 (ISSN-L)
- OpenAlex
- W2084119414
- Cited by
- 156
- References
- 28
- Citations by year
- 2026: 2 · 2025: 1 · 2024: 11 · 2023: 5 · 2022: 2 · 2021: 12 · 2020: 9 · 2019: 6 · 2018: 5 · 2017: 8 · 2016: 4 · 2015: 5 · 2014: 6 · 2012: 8
- FWCI
- 4.61
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Journal of Biological Chemistry (publishedVersion, cc-by)
- PubMed (publishedVersion)
- eScholarship@UMassChan (submittedVersion)
- Funders
- National Institute of Diabetes and Digestive and Kidney Diseases (DK54479, R01DK054479)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular transport and secretion
- Health Sciences › Medicine › Epidemiology › Autophagy in Disease and Therapy
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Physiology › Calcium signaling and nucleotide metabolism
- Keywords
- EEA1, phosphatidylinositol 3-phosphate, Rab5, early endosomes, endosome fusion, FYVE domain, endosomal trafficking
- MeSH terms
- Amino Acid Sequence, Amino Acids, Animals, Cytosol, Luminescent Proteins, Membrane Fusion, Membrane Proteins, Microscopy, Fluorescence, Molecular Sequence Data, Mutation, Plasmids, Protein Binding, Endosomes, Recombinant Fusion Proteins, Recombinant Proteins, Transfection, Transferrin, Point Mutation, Protein Structure, Tertiary, Phosphatidylinositol Phosphates, COS Cells, rab5 GTP-Binding Proteins, Vesicular Transport Proteins, Green Fluorescent Proteins
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