The YPLGVG sequence of the Nipah virus matrix protein is required for budding.
BACKGROUND
Nipah virus (NiV) is a recently emerged paramyxovirus capable of causing fatal disease in a broad range of mammalian hosts, including humans. Together with Hendra virus (HeV), they comprise the genus Henipavirus in the family Paramyxoviridae. Recombinant expression systems have played a crucial role in studying the cell biology of these Biosafety Level-4 restricted viruses. Henipavirus assembly and budding occurs at the plasma membrane, although the details of this process remain poorly understood. Multivesicular body (MVB) proteins have been found to play a role in the budding of several enveloped viruses, including some paramyxoviruses, and the recruitment of MVB proteins by viral proteins possessing late budding domains (L-domains) has become an important concept in the viral budding process. Previously we developed a system for producing NiV virus-like particles (VLPs) and demonstrated that the matrix (M) protein possessed an intrinsic budding ability and played a major role in assembly. Here, we have used this system to further explore the budding process by analyzing elements within the M protein that are critical for particle release.


RESULTS
Using rationally targeted site-directed mutagenesis we show that a NiV M sequence YPLGVG is required for M budding and that mutation or deletion of the sequence abrogates budding ability. Replacement of the native and overlapping Ebola VP40 L-domains with the NiV sequence failed to rescue VP40 budding; however, it did induce the cellular morphology of extensive filamentous projection consistent with wild-type VP40-expressing cells. Cells expressing wild-type NiV M also displayed this morphology, which was dependent on the YPLGVG sequence, and deletion of the sequence also resulted in nuclear localization of M. Dominant-negative VPS4 proteins had no effect on NiV M budding, suggesting that unlike other viruses such as Ebola, NiV M accomplishes budding independent of MVB cellular proteins.


CONCLUSIONS
These data indicate that the YPLGVG motif within the NiV M protein plays an important role in M budding; however, involvement of any specific components of the cellular MVB sorting pathway in henipavirus budding remains to be demonstrated. Further investigation of henipavirus assembly and budding may yet reveal a novel mechanism(s) of viral assembly and release that could be applicable to other enveloped viruses or have therapeutic implications.
Patch
Jared R
JR
Department of Microbiology and Immunology, Uniformed Services University, Bethesda, Maryland 20814, USA. Jared.Patch@ARS.USDA.GOV
Han
Ziying
Z
McCarthy
Sarah E
SE
Yan
Lianying
L
Wang
Lin-Fa
LF
Harty
Ronald N
RN
Broder
Christopher C
CC
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D002460:Cell Line; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D045405:Nipah Virus / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D016415:Sequence Alignment; D014763:Viral Matrix Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017201:Virus Shedding
D014763:Viral Matrix Proteins