Ebola virus matrix protein VP40 interaction with human cellular factors Tsg101 and Nedd4.
The Ebola virus matrix protein VP40 is a major viral structural protein and plays a central role in virus assembly and budding at the plasma membrane of infected cells. For efficient budding, a full amino terminus of VP40 is required, which includes a PPXY and a PT/SAP motif, both of which have been proposed to interact with cellular proteins. Here, we report that Ebola VP40 can interact with cellular factors human Nedd4 and Tsg101 in vitro. We show that WW domain 3 of human Nedd4 is necessary and sufficient for binding to the PPXY motif of VP40, which requires an oligomeric conformation of VP40. Single particle electron microscopy reconstructions indicate that WW3 of Nedd4 is in close contact with the N-terminal domain of hexameric VP40. In contrast, the ubiquitin enzyme variant domain of Tsg101 was sufficient for binding to the PT/SAP motif of VP40, regardless of the oligomeric state of the matrix protein. These results suggest that hNedd4 and Tsg101 may play complimentary roles at a late stage of the assembly process, by recruiting cellular factors of two independent pathways to the site of budding at the plasma membrane.
Timmins
Joanna
J
European Molecular Biology Laboratory, 6 rue Jules Horowitz, 38042 Grenoble, France.
Schoehn
Guy
G
Ricard-Blum
Sylvie
S
Scianimanico
Sandra
S
Vernet
Thierry
T
Ruigrok
Rob W H
RW
Weissenhorn
Winfried
W
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D056827:Endosomal Sorting Complexes Required for Transport; D007091:Image Processing, Computer-Assisted; D008025:Ligases / Q000235:genetics / Q000378:metabolism*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D000075702:Nedd4 Ubiquitin Protein Ligases; D009698:Nucleoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D020349:Surface Plasmon Resonance; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D014758:Viral Core Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication; D000099177:Tsg101 Protein
D002135:Calcium-Binding Proteins; D004268:DNA-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D008025:Ligases; D000075702:Nedd4 Ubiquitin Protein Ligases; D009698:Nucleoproteins; D011994:Recombinant Proteins; D014157:Transcription Factors; D044767:Ubiquitin-Protein Ligases; D014758:Viral Core Proteins; D000099177:Tsg101 Protein; C084819:nucleoprotein VP40, Ebola virus; C000618483:Nedd4 protein, human