replication neg RNA virus Flashcards
which are the only RNA viruses replicating in the nucleus? What is the advantage?
Borna Virus and Influenza virus
advantage: using the splicing apparatus
what are the fundamental characteristics of RNA minus strand replication?
1.- strand is not infectious
2.viral enzymes for formation of + strand needed
3.active RdRp in virus particle
4.replication via complete + strand intermediate
what are the characteristics of the genome of Orthomyxo-,, Paramyxo, and Rhabdoviridae?
-serves as template for mRNA synthesis
-always in a complex with proteins
-without proteins not infectious
-viral RdRp packed into virion
what are the characteristics of the Rhabdovirus (Vesicular Stomatits Virus VSV)?
Genome: not segmented and replicated in cytoplasm
5 Proteins: G (Glycoprotein), L and P (RNA polymerase), M (Matrix), N (Nucleocapsid protein –> transcription according to the attenuation model
3’end: Leader-N-P-M-G-L-5’end
viral RdRP (L): transcribes 5 monocistronic mRNAs
capping and polyadenylation
describe the cell cycle of VSV
1.binding and fusion: release of helical nucleocapsid
2.synthesis of 5 mRNA and leader RNA
3.Translation of viral mRNA
4.+ Strand synthesis
5.- strand synthesis
6.processing and localization of G-Protein
7.Packaging and release with M-Protein as driving force
what is the Attenuation model? In which virus does it play a role?
in VSV virus:
start of the mRNA synthesis at the 3’ end of the N gene, not the 3’ end of the genome (inefficient reinitiation after termination of transcription) –> attenuation: decreasing amounts of transcript from 3’ to 5’ (N>P>M>G>L) –> severely reducced amounts of L (RdRP)
Poly A via stuttering of viral polymerase at intergenic region
VSV transcription: what is the mechanism behind the attenuation of transcription? What model?
1.Initiation of mRNA synthesis by binding of L (RdRP) in complex with three P at the 3’ end of N gene
2-Termination of mRNA synthesis at intergenic region (ig)
3.inefficient reinitiation after termination of transcription at the 3’ end of the P gene
–> start stop model
How is the Poly A tail synthesized in VSV?
-Synthesis of 7A residues in IG region by VSV polymerase complementary to 7Us
-stuttering of polymerase: those 7 Us are transcribed multiple times
-Termination of mRNA synthesis at ig region after 200 A
-Inefficient reinitiation –> capping
VSV: how does the switch between mRNA and genome synthesis work?
switching from mRNA synthesis (early) to replication (late) via changes in the composition of the replicase complex:
L+P = transcription
L+N+P = replication
intracellualr accumulation of amount of N decisive for switch to replication:
N=low= mRNA synthesis
N=high= replication
what does the crystal structure of a nucleocapsid-like Nucleoprotein-RNA complex of respiratory syncytial virus look like?
10 N-proteins in complex with RNA Ring –> serves as template for viral RNA synthesis
How does the rabies virus switch from transcription to replication?
switch controlled by intracellular concentration of M-protein
low amount of M: Transcription
high amount of M: replication
Why are animal viruses relevant research objects?
-often very expensive outbreaks
-importamt experimental models
-transmission from animals to humans: zoonosis
why does rabies virus package - strand RNA into its virion?
no selective packaging but strand imbalance: more - than +
very strong RNA promotor at 3’ end of + strand –> overproportional - strand synthesis
how is the genome organized and structured in influenza virus?
-segmented genome: 8 segments –> 1-2 proteins (each associated with polymerase complex and NP)
-splicing –> nucleus
-reassortment
-viral RdRp (3 subunits: PB1, PB2, PA)
-cap snatching
what is cap snatching? How does it work?
5’ ends of cellular pre-,RNA in nucleus is cleaved off and serves as primer for viral mRNA synthesis
mechanism:
hydrolysis of a random capped nuclear mRA via nuclease activity of viral polymerase complex.
why is the influenza genome so variabel? why is it so problematic?
antigenic shift/reassortment:
exchange of surface proteins via reassortment (segments switched) leads to massiv change in antigenicity of the virus –> also possible between fowl, swine, humans: cross infections between swine and human possible but only rarely between human and birds
PROBLEM: no acquired protective immunity via previous infections or vaccinations
describe the replication cycle of influenza a virus
1.binding via HA to sialic acid residues
2.endocytosis and pH induced fusion via HA
3.nuclear transport of nucleocapsid via NLS in NP and in PB2
4.synthesis of 8 mRNAs in the nucleus including cap snatching
5.mRNA splicing of NS2/M2
6.synthesis of PA/PB1/PB2 (nuclear import)
7.export and translation of viral mRNAs
8.processing and localization of HA, NA
what role does the ribonucleotiprotein complex (RNP) play in influenza virus?
genomic RNA never naked –> always associated to RNP
polymerase complex mediates circularisation of genome segments
how does the mRNA synthesis of influenza A virus work?
permanent synthesis of capped and polyadenylated mRNA in nucleus:
3 RNA species: vRNA (-strand RNA), viral mRNA (cap, Poly A) and cRNA (+ strand RNA)
viral pol snatches cap –> cap-primer dependent initiation of transcribtion and elongation by viral polymerase
replication is mediated by NP
cRNA synthesis in cis
vRNA synthesis in trans
mRNA splicing
what controls/changes the activity of the replication complex in influenza virus? what is the consequence?
binding of specific sequences in the viral RNAs to pol subunits
–>functionally different replication complexes catalyse transcription or replication
what are the functions of the 5’ end cap in influenza virus?
-RNA stabilisation
-essential for translation initiation
-essential for binding mRNA to eIF4A which mediates ribosome binding to cap
how is the replication of influenza virus regulated? Why is there an inbalance between vRNA and cRNA?
RNA promotor: corksrew structure of communicating ends –> differently efficient RNA promotors in vRNA (weak promotor for sythesis of cRNA) and cRNA (strong promotor for synthesis of vRNA) –> vRNA:cRNA=10:1
–> because cRNA is only a replication intermediate, vRNA is packed into the virion
how are RNA segemnts of Influenza virus packed into new virions?
segment specific packaging signals at the 5’ and 3’ ends (NTR)
8 helical rods (genome) with different lenghts in the virion
why is there genetic diversity of negative strand RNA viruses?
missing accuracy of RNA polymerase (RdRp has no proof readign activity)
segmentation allows reassortment
