title: Alix serves as an adaptor that allows human parainfluenza virus type 1 to interact with the host cell ESCRT system.
abstract: The cellular ESCRT (endosomal sorting complex required for transport) system functions in cargo-sorting, in the formation of intraluminal vesicles that comprise multivesicular bodies (MVB), and in cytokinesis, and this system can be hijacked by a number of enveloped viruses to promote budding. The respiratory pathogen human parainfluenza virus type I (HPIV1) encodes a nested set of accessory C proteins that play important roles in down-regulating viral transcription and replication, in suppressing the type I interferon (IFN) response, and in suppressing apoptosis. Deletion or mutation of the C proteins attenuates HPIV1 in vivo, and such mutants are being evaluated preclinically and clinically as vaccines. We show here that the C proteins interact and co-localize with the cellular protein Alix, which is a member of the class E vacuolar protein sorting (Vps) proteins that assemble at endosomal membranes into ESCRT complexes. The HPIV1 C proteins interact with the Bro1 domain of Alix at a site that is also required for the interaction between Alix and Chmp4b, a subunit of ESCRT-III. The C proteins are ubiquitinated and subjected to proteasome-mediated degradation, but the interaction with AlixBro1 protects the C proteins from degradation. Neither over-expression nor knock-down of Alix expression had an effect on HPIV1 replication, although this might be due to the large redundancy of Alix-like proteins. In contrast, knocking down the expression of Chmp4 led to an approximately 100-fold reduction in viral titer during infection with wild-type (WT) HPIV1. This level of reduction was similar to that observed for the viral mutant, P(C-) HPIV1, in which expression of the C proteins were knocked out. Chmp4 is capable of out-competing the HPIV1 C proteins for binding Alix. Together, this suggests a possible model in which Chmp4, through Alix, recruits the C proteins to a common site on intracellular membranes and facilitates budding.
authors.affiliation: Laboratory of Infectious Diseases, RNA Viruses Section, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Jim
authors.initials: J
authors.lastname: Boonyaratanakornkit
authors.forename: Henrick
authors.initials: H
authors.lastname: Schomacker
authors.forename: Peter
authors.initials: P
authors.lastname: Collins
authors.forename: Alexander
authors.initials: A
authors.lastname: Schmidt
mesh_terms: D001667:Binding, Competitive / Q000235:genetics; D015153:Blotting, Western; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D056827:Endosomal Sorting Complexes Required for Transport / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D047468:Immunoprecipitation; D013058:Mass Spectrometry; D010222:Parainfluenza Virus 1, Human / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: C481529:CHMP4A protein, human; D002135:Calcium-Binding Proteins; D018797:Cell Cycle Proteins; D017931:DNA Primers; D056827:Endosomal Sorting Complexes Required for Transport; C117020:PDCD6IP protein, human; D034741:RNA, Small Interfering; D014764:Viral Proteins; C068856:C protein, human parainfluenza virus type 1