Over-expression of Rififylin, a new RING finger and FYVE-like domain-containing protein, inhibits recycling from the endocytic recycling compartment.
Endocytosed membrane components are recycled to the cell surface either directly from early/sorting endosomes or after going through the endocytic recycling compartment (ERC). Studying recycling mechanisms is difficult, in part due to the fact that specific tools to inhibit this process are scarce. In this study, we have characterized a novel widely expressed protein, named Rififylin (Rffl) for RING Finger and FYVE-like domain-containing protein, that, when overexpressed in HeLa cells, induced the condensation of transferrin receptor-, Rab5-, and Rab11-positive recycling tubulovesicular membranes in the perinuclear region. Internalized transferrin was able to access these condensed endosomes but its exit from this compartment was delayed. Using deletion mutants, we show that the carboxy-terminal RING finger of Rffl is dispensable for its action. In contrast, the amino-terminal domain of Rffl, which shows similarities with the phosphatidylinositol-3-phosphate-binding FYVE finger, is critical for the recruitment of Rffl to recycling endocytic membranes and for the inhibition of recycling, albeit in a manner that is independent of PtdIns(3)-kinase activity. Rffl overexpression represents a novel means to inhibit recycling that will help to understand the mechanisms involved in recycling from the ERC to the plasma membrane.
Coumailleau
Franck
F
Unité Biologie du Développement, CNRS URA 2578, Institut Pasteur, 75724 Paris Cedex 15, France.
Das
Vincent
V
Alcover
Andres
A
Raposo
Graça
G
Vandormael-Pournin
Sandrine
S
Le Bras
Stéphanie
S
Baldacci
Patricia
P
Dautry-Varsat
Alice
A
Babinet
Charles
C
Cohen-Tannoudji
Michel
M
D000595:Amino Acid Sequence; D000818:Animals; D051017:Apoptosis Regulatory Proteins; D001483:Base Sequence; D004705:Endocytosis / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism* / Q000648:ultrastructure; D006367:HeLa Cells; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D014168:Transferrin / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases; D016335:Zinc Fingers
D051017:Apoptosis Regulatory Proteins; D011506:Proteins; D011993:Recombinant Fusion Proteins; D014168:Transferrin; C502761:Rffl protein, mouse; D044767:Ubiquitin-Protein Ligases