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Reoviridae
Molecular Virology
Reoviridae
Respiratory enteric orphan (呼腸孤病毒)
Depends on the presence of a segmented, dsRNA genome
the structure characters
replication strategy
Reoviridae consists of 12 genera
Mammalian orthoreovirus prototypes
Type 1 (Lang, T1L):from a healty child
Type 2 (Jones, T2L):from a child with diarrhea
Type 3 (Dearing, T3D):
from a child with diarrhea or (Abney, T3A) an upper respiratory illness
Avian reoviruses
Viral arthritis (VA)
Pale bird syndrome (PBS)
dsRNA Viruses
Family
Genera
Genome Segments
Hosts
Reoviridae
12
10, 11, or 12
(See individual genera)
Orthoreovirus
10
Mammals, Birds, Reptiles
Aquareovirus
11
Fish, Molluscs
Cypovirus
10
Insects
Idnoreovirus
10
Insects
Fijivirus
10
Plants, Insectsb
Oryzavirus
10
Plants, Insectsb
Mycoreovirus
11 or 12
Fungi
Rotavirus
11
Mammals, Birds
Orbivirus
10
Mammals, Birds, Arthropodsb
Coltivirus
12
Mammals, Arthropodsb
Phytoreovirus
12
Plants, Insectsb
Seadornavirus
12
Mammals, Insectsb
Turreteda
Nonturreteda
dsRNA Viruses
Family
Genera
Genome Segments
Hosts
Birnaviridae
3
2
Birds, Fish, Insects
Totiviridae
3
1
Fungi, Protozoa
Partitiviridae
3
2
Fungi, Plants
Chrysoviridae
1
4
Fungi, Plants
Hypoviridae
1
1
Fungi
Cystoviridae
1
3
Bacteria
aProposed
division based on presence or absence of turret-like protein projecting around fivefold
axes from innermost capsid layer.
bServe as vectors for transmission to other hosts.
Mammalian v.s. Avian reovirus
Cell fusion
－
＋
HA
＋
－
Fusogenic reovirus
Nonfusogenic reovirus
Avian reovirus can grow in mammalian cell line
Properties of nonfusogenic mammalian reovirus
Genome
Perfect double-stranded RNA
10 gene segments in three size classes (L, M, S)
Total size ~23,500 base pairs
Gene segments encode either one or two proteins each
Gene segments are transcribed into full-length mRNAs
Plus strands of gene segments have 5’caps
Nontranslated regions at segment termini are short
Gene segments can undergo reassortment between virus
strains
Short subgenomic RNA
Properties of nonfusogenic mammalian reovirus
Particles
Spherical, with icosahedral (5:3:2) symmetry
Nonenveloped
Total diameter ~85 nm (excluding σ 1 fibers)
Two concentric protein capsids: outer capsid subunits in T = 13
lattice, arrangement of inner capsid subunits in T = 1
8 structural proteins: 4 proteins in outer capsid [λ 2, μ1 (mostly
as cleavage fragments μ 1N and μ 1C), σ 1, and σ 3] and 4
proteins in inner capsid (λ 1, λ 3, μ 2, and σ 2)
Subviral particles (ISVPs and cores) can be generated from fully
intact particles (virions) by controlled proteolysis
Cell-attachment protein σ 1 can extend from the virion and ISVP
surface as a long fiber
Protein λ 2 forms pentamers that protrude from the core surface
Properties of nonfusogenic mammalian reovirus
Replication
Fully cytoplasmic
Sialic acid can serve as a cell surface receptor for recognition by
cell-attachment protein σ 1.
Proteolytic processing of outer capsid proteins σ 3 and μ 1/μ 1C
is essential to infection and can occur either extracellularly or in
endo/lysosomes.
Uncoating of parent particles is incomplete: genomic dsRNA
does not exit particles to enter the cytoplasm.
Transcription and capping of viral mRNAs occur within particles
and are mediated by particle-associated enzymes.
Segment assortment and packaging involves mRNAs.
Minus-strand synthesis occurs within assembling particles.
Mature virions are inefficiently released from infected cells by
lysis.
Ten Segmented dsRNA
Gene organization
Viral protein
MonoDi-
cistronic S1
Tri-
RNA
FIGURE 52.2 Coding strategies of the ten genome segments of reovirus type 3 Dearing. Segments are drawn approximately to scale and are
oriented so that left-to-right corresponds to 5??to 3??for the protein-coding plus strands. The segment names and lengths in nucleotides (nuc #)
are listed at left. The portion of each segment encompassed by protein-coding sequences is hatched. Short nontranslated regions at the ends of
each segment remain unshaded, and the lengths of these regions in nucleotides (nuc #) are designated above. Names of encoded proteins and their
lengths in amino acids (aa #) are listed at right. The S1 segment encodes two proteins as shown: The ?1s protein (offset, lighter hatching) initiates
at a second initiator codon in a different reading frame from ?1 (darker hatching). A second protein, 繕NSC (lighter hatching) also arises from the
M3 segment but in its case is thought to initiate at a second initiator codon in the same reading frame as 繕NS (darker hatching).
Structure of virus particles
Structural composition of orthoreovirus
Structural proteins of reovirus
Core proteins consisting of λ1, λ3, μ2 and σ2
proteins form internal transcriptase complexes
Outer capsid proteins consisting of λ2, μ1C, σ1
and σ3 proteins form external turrets and
nodules
Core structural proteins
Encoding
Segment
Protein
Mass (kd)
Presence in
Particle Formsa
L1
λ3
142
V, I, C
RNA-dependent RNA
polymerase
L3
λ1
143
V, I, C
Binds RNA, Zn metalloprotein,
NTPase, RNA helicase, RNA
triphosphatase
M1
μ2
83
V, I, C
Binds RNA, NTPase, a MAP
S2
σ2
47
V, I, C
Binds dsRNA
aV.I.C.:
virus particle ISVP, core.
Function or Property
Core structural proteins
Unclear in the complete core organization
σ2/λ1 is 2/1 and forms a complex
Iodination is less easily carried out on σ2 than λ1
Contains transcriptase and replication activity
Outer capsid structural proteins
Encoding
Segment
Protein
Mass (kd)
Presence in
Particle Formsa
L2
λ2
145
V, I, C
Guanylyltransferase,
methyltransferases, capping
M2
μ1/μ1C
76
V, I
N-myristoylated, cleaved into
fragments, role in penetration,
role in transcriptase activation
S1
σ1
49
V, I
Cell-attachment protein,
hemagglutinin, primary
serotype determinant,
apoptosis
S4
σ3
41
V
Sensitive to protease
degradation, binds dsRNA,
zinc metalloprotein, effects on
cellular translation
aV,
virion; I, ISVP; C, core.
Function or Property
Outercapsid structural proteins
• μ1C is proteolytically cleaved from μ1.
Glycoprotein.
• In cytoplasm of virus-infected cells, 95% of protein
complexed with σ3 is μ 1C (not μ1). Cleavage of μ1
may be linked to the formation of a complex with σ3.
• μ1C/σ3 is 1/1 in vivion.
• Sequences are highly conserved between viruses and
serve an structural constraints on the function.
• Cleavage site located at 42 and 43 AA of μ1 from Nterminus.
μ1C protein cleavage
Digestion of μ1C protein
Site for μ1c protein digestion
σ1 protein
• Proximately close to the λ2
• Is tetramer
• N-terminus is helix (Fiber), located in a
λ2 channel.
• Has an α-helix coiled-coil structure
Primary structure of σ1 protein
λ2 protein
Viral core: 60 nm
Icosahedron: 12 verticles, 20 faces, and 30 edges
Care spike: 12
Each spike consists of 5 λ2 protein, projects out
5-6 nm above the surface of the core.
Identified by mAb to λ2, has a hole or channel
formation under EM
Non-structural of reovirus
• Non-fusogenic mammalian reovirus
μNS σNS σ1S
• Fusogenic reovirus
μNS σNS
P10: induces cell fusion
P17: contains nuclear leader signal (NLS)
Viral replication cycle
Absorption
Receptor: sialic acid (some intergrins may involve)
Penetration
Uncoating
incomplete uncoating
Virion—ISVP—Core
Transcription
Newly synthesized +strand mRNA within
Core and using conservative transcription model
Early transcripts with 5’ capped mRNA
Late transcripts without 5’ capped mRNA
Early transcripts: L1, M3, S3 and S4 gene segments
All ten genes are transcribed at the same time in vitro
May have a cellular repressor prevent late transcription and a
protein encoded by early gene results in derepression
Replication of viral RNA genome
Using conservative transcription model
mRNA used as templates for “-” strand RNA
synthesis (no 5’ capping)
Single round for “-” RNA synthesis
σNS assorts 10 segment ssRNA for packaging
Late transcription of virus
Transcribed to be a non 5’ capped mRNA for late
protein synthesis
Appears at 4-6 h after infection, reaches to be
maximum at 12 h and then decrease after 12 h
Assembly and release
σ3 is the last one to the complete particle
Once σ3 is assembly, transcriptional activities are
terminated
Important Terminology
•
•
•
•
Reassortment
Cell fusion from within
Cell fusion from without
Persistent infection
Protein interactions in viral factories