Article

Organization of the Influenza Virus Replication Machinery

National Resource for Automated Molecular Microscopy, Department of Cell Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
Science (Impact Factor: 33.61). 11/2012; 338(6114). DOI: 10.1126/science.1227270
Source: PubMed

ABSTRACT

Influenza virus ribonucleoprotein complexes (RNPs) are central to the viral life cycle and in adaptation to new host species. RNPs are composed of the viral genome, viral polymerase, and many copies of the viral nucleoprotein. In vitro cell expression of all RNP protein components with four of the eight influenza virus gene segments enabled structural determination of native influenza virus RNPs by cryo-EM. The cryo-EM structure reveals the architecture and organization of the native RNP, thereby defining the attributes of its largely helical structure and how polymerase interacts with NP and the viral genome. Observations of branched-RNP structures in negative stain EM and their putative identification as replication intermediates suggest a mechanism for viral replication by a second polymerase on the RNP template.

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    • "NP is a major component of the ribonucleoprotein (RNP) complexes. The RNP consists of RNA, multiple copies of NP, and polymerase subunits (PB1, PB2 and PA), together forming a large loop that is twisted into a helical rod-like structure [19] [20]. NP plays regulatory roles in transcription, replication, and virus maturation [21], and it is one of the most conserved proteins among influenza A viruses with >90% protein sequence homology [22] [23]. "
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    • "NP is a major component of the ribonucleoprotein (RNP) complexes. The RNP consists of RNA, multiple copies of NP, and polymerase subunits (PB1, PB2 and PA), together forming a large loop that is twisted into a helical rod-like structure [19] [20]. NP plays regulatory roles in transcription, replication, and virus maturation [21], and it is one of the most conserved proteins among influenza A viruses with >90% protein sequence homology [22] [23]. "
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    ABSTRACT: There are two major limitations to vaccine preparedness in the event of devastating influenza pandemics: the time needed to generate a vaccine and rapid generation of sufficient amounts. DNA vaccination could represent a solution to these problems, but efficacy needs to be enhanced. In a separate line of research, it has been established that targeting of vaccine molecules to antigen-presenting cells enhances immune responses. We have combined the two principles by constructing DNA vaccines that encode bivalent fusion proteins; these target hemagglutinin to MHC class II molecules on antigen-presenting cells. Such DNA vaccines rapidly induce hemagglutinin-specific antibodies and T cell responses in immunized mice. Responses are long-lasting and protect mice against challenge with influenza virus. In a pandemic situation, targeted DNA vaccines could be produced and tested within a month. The novel DNA vaccines could represent a solution to pandemic preparedness in the advent of novel influenza pandemics.
    No preview · Article · Mar 2015 · Expert Review of Vaccines
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    • "Sample dilution was adjusted to achieve a homogeneous separation of particles. Samples were stained as previously described (Moeller et al., 2012) (Tao et al., 2013) using a 2% uranyl formate solution. EM micrographs were acquired using a Tecnai F20 Twin transmission electron microscope operating at 200 kV, using a dose of $45 e À /A ˚ 2 and nominal underfocus ranging from 0.7 mm to 1.7 mm. "
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    Full-text · Article · Jan 2015 · Structure
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