The glycoprotein cytoplasmic tail of Uukuniemi virus (Bunyaviridae) interacts with ribonucleoproteins and is critical for genome packaging.
We have analyzed the importance of specific amino acids in the cytoplasmic tail of the glycoprotein G(N) for packaging of ribonucleoproteins (RNPs) into virus-like particles (VLPs) of Uukuniemi virus (UUK virus), a member of the Bunyaviridae family. In order to study packaging, we added the G(N)/G(C) glycoprotein precursor (p110) to a polymerase I-driven minigenome rescue system to generate VLPs that are released into the supernatant. These particles can infect new cells, and reporter gene expression can be detected. To determine the role of UUK virus glycoproteins in RNP packaging, we performed an alanine scan of the glycoprotein G(N) cytoplasmic tail (amino acids 1 to 81). First, we discovered three regions in the tail (amino acids 21 to 25, 46 to 50, and 71 to 81) which are important for minigenome transfer by VLPs. Further mutational analysis identified four amino acids that were important for RNP packaging. These amino acids are essential for the binding of nucleoproteins and RNPs to the glycoprotein without affecting the morphology of the particles. No segment-specific interactions between the RNA and the cytoplasmic tail could be observed. We propose that VLP systems are useful tools for analyzing protein-protein interactions important for packaging of viral genome segments, assembly, and budding of other members of the Bunyaviridae family.
Overby
Anna K
AK
Ludwig Institute for Cancer Research, Stockholm Branch, Karolinska Institute, Box 240, SE-17177 Stockholm, Sweden.
Pettersson
Ralf F
RF
Neve
Etienne P A
EP
D000409:Alanine / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism*; D017930:Genes, Reporter / Q000235:genetics; D016679:Genome, Viral / Q000235:genetics*; D006023:Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009698:Nucleoproteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D012261:Ribonucleoproteins / Q000378:metabolism*; D018066:Uukuniemi virus / Q000235:genetics* / Q000378:metabolism* / Q000648:ultrastructure; D014771:Virion / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D019065:Virus Assembly
D006023:Glycoproteins; D009698:Nucleoproteins; D012261:Ribonucleoproteins; D000409:Alanine