title: Endofin, an endosomal FYVE domain protein.
abstract: KIAA0305 is an uncharacterized member of the FYVE domain protein family. It is closely related to SARA, with about 50% identity in the carboxyl-terminal 800-amino acid region. Indirect immunofluorescence microscopy using polyclonal antibodies raised against KIAA0305 revealed that it is enriched in early endosomes. The Myc-tagged version is also faithfully targeted to the early endosome. We have tentatively called KIAA0305 endofin (for endosome-associated FYVE-domain protein). The association of endofin with endosomes is mediated by its FYVE domain because deletion mutants lacking the central FYVE finger motif are distributed in the cytoplasm. In addition, a single point mutation in the FYVE finger motif at cysteine residue 753 (C753S) is sufficient to abolish its endosomal association. Its endosomal localization is also sensitive to the phosphatidylinositol 3-kinase inhibitor, wortmannin. Using in vitro liposome binding assays, we demonstrate that Myc-tagged endofin associates preferentially with phosphatidylinositol 3-phosphate, whereas the C753S point mutant was unable to do so. We also show that endofin co-localizes with SARA but that they are not associated in a common complex because they failed to co-immunoprecipitate in co-expressing cells. Endofin also does not associate with Smad2 nor behave like SARA in affecting transforming growth factor-beta signaling. At high levels of expression, both endofin and SARA can cause an endosome aggregation/fusion effect. In COS7 cells, which can support high levels of exogenous protein expression, both proteins can also cause other structural anomalies in the endocytic pathway, as represented by enlarged vesicular structures. These endosomal aggregates/fusions accumulated endocytosed epidermal growth factor. Taken together, this report provides evidence to suggest that endofin and the highly related SARA are endosomal proteins with potential roles in regulating membrane traffic.
authors.affiliation: Membrane Biology Laboratory, Institute of Molecular and Cell Biology, Singapore 117609, Singapore.
authors.forename: L F
authors.initials: LF
authors.lastname: Seet
authors.forename: W
authors.initials: W
authors.lastname: Hong
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D002352:Carrier Proteins / Q000032:analysis* / Q000737:chemistry; D002449:Cell Aggregation; D002460:Cell Line; D011992:Endosomes / Q000737:chemistry*; D004815:Epidermal Growth Factor / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D012697:Serine Endopeptidases; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D016335:Zinc Fingers
chemical_list: D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D016212:Transforming Growth Factor beta; D004815:Epidermal Growth Factor; D019869:Phosphatidylinositol 3-Kinases; C116420:ZFYVE16 protein, human; D012697:Serine Endopeptidases