Lec 9 Paramyxovirus

Question 1

Order: Mononegavirales

Answer 1

Family: Bornaviridae
Family: Filoviridae
Family: Paramyxoviridae
Family: Rhabdoviridae (bullet shaped virions) Share some similarities.

Question 2

Paramyxoviridae

Answer 2

Pleomorphic (differing membranes on the outside, stolen from host), enveloped, helical, -ve sense RNA virus. Not segmented genome though. Core is encapsidated w/ NP (nucleocapsid) protein, which aggregate to form arrays of proteins.

Associated with broad range of disease
Respiratory (paraflu, RSV)
Glandular (mumps)
Epithelial (measles)
Chronic? (MS) Debatable 

Resembles Rhabdovirus in transcription and regulation,

Orthomyxovirus at the membrane level

Question 3

Paramyxoviridae Traits

Answer 3

Paramyxovirus
Sendai virus, type species, pathogenic in Mice, “King of Fusion” Parainfluenza type 1
Human Parainfluenza Types 1&3, 
respiratory infections
flu-like virus of humans

HPIV3 alone is responsible for ~11% of pediatric respiratory Hospitalizations and is the predominant cause of Croup in young infants

Rubulavirus
Mumps virus, glandular infection
Newcastle Disease Virus, avian virus

Human Parainfluenza types 2, 4a, 4b, similar symptoms, virus differs in properties. Very severe in children and very powerful in humans in general.

Viruses belonging to the paramyxovirus family, particularly respiratory syncytial virus (RSV), the recently identified human metapneumovirus (1), and the human parainfluenza viruses (HPIVs) types 1, 2, and 3, cause the majority of respiratory childhood cases of croup, bronchiolitis, and pneumonia worldwide (2).

HPIV3 alone is responsible for approximately 11% of pediatric respiratory hospitalizations in the US (3, 4) and is the predominant cause of croup in young infants, while HPIV1 and -2 tend to infect older children and adolescents

Question 4

Syncytia in Paramyxoviridae

Answer 4

Syncytia - Multinuclear Cell, Respond to the expression of measles virus genes. Giant cells formed when many cells fuse together because they are infected by Sendai virus. Have a fusion protein that fuses infected cells w/ others and uninfected neighbours!

Question 5

2 Paramyxoviridae

Morbillivirus and Pneumovirus

Answer 5

Morbillivirus
Measles Virus, typically mild childhood disease, more virulent with age, may be linked to chronic infections (MS)
Canine Distemper Virus, pathogen of dogs

Sub family Pneumovirinae
Pneumovirus
Human Respiratory Syncytial Virus (RSV), important respiratory infection in infants

Question 6

Paramyxoviruses

Henipavirus (Emerging Viruses)

Niphavirus and Hendravirus

Localized infections (community wise)

Answer 6

Henipavirus (Emerging Viruses) Zoonosis! and person to person

NIPAH VIRUS first arose in Malaysia and Singapore in1998-99, subsequently caused a number of outbreaks primarily in Bangladesh
Highly pathogenic causing a severe febrile encephalitis in humans with high mortality rate
Pigs and bats may be normal reservoir (Zoonosis)
Can spread person to person

HENDRA VIRUS also causes severe encaphalitis and death in adults and children
Natural reservoir is flying foxes (bats)

R0 is basic reproduction Number in Bangladesh outbreak was low 0.48 which is below 1 suggesting that the outbreak will die out.

Question 7

Paramyxovirus Structure

Answer 7

Pleomorphic virus range from 100-800 nm average at 150-350 nm, spherical in shape

Membrane of host plasma membrane lipid and 2 VIRAL SPIKE PROTEINS (PEPLOMERS) and matrix protein underneath membrane, gives shape.

Question 8

Paramyxovirus Structural Proteins

M Protein

**

Answer 8

Matrix Protein (M). Most common protein in vision. 
basic protein

Matrix Protein also
Forms paracrystaline layer under membrane
Organizes & stabilizes virus by interaction of cytoplasmic tail with HN, F, and NP (nucleocapsid protein)
Also orients nucleocapsid to HN and F proteins.
HN protein in paramyxovirus has abilities of HA and GA from orthomyxoviridae!
Most abundant protein

Question 9

Paramyxovirus Fusion Protein (F)

Answer 9

MUST BE CLEAVED to FORM DISULFIDE-LINKED DIMER (F1 & F2) to be active, stick outside from virus.

Peptide on base of integral membrane protein triggers fusion. Involved in membrane fusion and hemolysis (kill RBC)

Question 10

Paramyxovirus Hemagglutinin-Neuraminadase (HN)

Answer 10

HN Attachment glycoprotein to host cell receptors (Sialic Acid)

Paraflu and Rubulavirus HN have both sialic acid binding and cleavage activity

morbillivirus HN has only binding activity, needs something else to perform cleavage*

Question 11

Structure of Paramyxovirus helical nucleocapsid

Answer 11

Left handed helix with the nuclelotides in white responsible for initiation of replication and transcription

Nucleocapsid proteins encapsidate RNA to form this helix. Attach on specific sequence (leader genome and promoter)

MP protein forms long arrays. There are also individual subunits which form the ends of the arrays.

Question 12

Paramyxovirus proteins

Nucleocapsid Core Structure

NP

P Protein

L Protein

Answer 12

Core structure made up of gRNA (genomic RNA), NP, P, and L proteins, remains intact during infection, is itself infectious and carries out transcription in vitro. Can isolate nucleocapsid w/o membrane.

Nucleocapsid Protein (NP)
Serves to coat gRNA
-Associates with L and P in transcription-replication and with M in docking (P and L form polymerase)

NP is not typical of RNA binding proteins,
-ve charged,
no typical RNA binding motifs

Phosphoprotein (P)
highly phosphorylated
-With L is involved in all RNA synthesis; and with NP is involved in gRNA encapsidation and regulation of mRNA to replication switch
-P gene can code for a number of other proteins in overlapping reading frames depending on virus.
-P Proteins believed to regulate switch from transcription to replication

Large Protein (L)
Serves as polymerase RNAdRNAp, with help of P and NP proteins
Adds 5’ caps and 3’ poly A tail to mRNA
L is the Least abundant but biggest protein

Question 13

Paramyxoviridae Viral Genome Specifities

Answer 13

Viral genome is ss-RNA, -ve sense, non-segmented, 5-7 X 106, 50S in size
50 nucleotide 3’ leader sequence and a 50 nucleotide 5’ tail (trailer) sequence enclose coding region

3’-N-P/C/V/-M-F-HN-L-5’

Genes are separated by short INTER CISTRONIC NUCLEOTIDE SEQUENCES (ICS) allow mRNA to form.

Green nucleocapsid protein envelops whole thing. purple P proteins attach to some places.

Question 14

Paramyxoviridae Life cycle

Answer 14

Absorption to receptors, pulled to host cell membrane, fusion protein fuses the 2 membranes and releases nucleocapsid core into the cytoplasm of the host, uncoats. There are still some HF and N inserted into the membrane and if they are up against the neighbouring cell to form syncytia! Allows bigger cytoplasm and nutrients for virus to form virions!

Different b/c the virus is not taken up into an endosome.

When -ssRNA from nucleocapsid core is release the genome is transcribed into + mRNA w/ 5’ cap and poly A tail. Translated into the different proteins which allow switch to replication mode where genome is replicated into anti genome, then replicated into -ssRNA viral genome. They are both encapsidated w/ the NP protein, mRNA isn’t!

H N and F proteins are inserted into rough ER and move to PM, and glycolylated.

Question 15

Paramyxovirus Attachment, Penetration, Uncoating

Answer 15

Attachment via HN protein binding sialic acid residues on cell surface. If you add Sialidase the infection is blocked.

Receptor binding triggers fusion
Sendai: gangliosides serve as receptor
Measles: CD46 serves as receptor
Sialidase treatment blocks infection

Fusion by F protein at plasma membrane pH independent. Endosomal membrane fusion in orthomyxoviridae is acidic pH dependent!

Binding of HN induces activation of F by conformational change

Uncoating limited to core release from membrane into cytoplasm
*Is the cell infected before the core even enters?

Question 16

Paramyxovirus Transcription

Answer 16

Like Rhabdovirus transcription starts from 3’ leader sequence producing mRNA for each gene by termination-reinitiation, with each mRNA capped and polyadenylated. When RNA pol ives down genome, when it hots an INTERCISTRONIC REGION it stops and reinitiates on the next gene.

B/c of the larger genome, in some viruses there is more equal transcription of some genomes where on other viruses there is a larger drop off on other proteins.

mRNA copy numbers dependant on reinitiation,
for Measles same chance at each site
for Sendai big drop off between M-F and HN-L

L and P associated with NP serve polymerase function,
C,V, and D genes coded in P gene region regulate transcription process

Question 17

Paramyxovirus Transcription Specifics

Answer 17

The individual genes produced are translated by RNAP. On lower RNAP complex the +ssRNA is read.

When cell is first infected N levels will be low and transcription of the genes will occur leading to translation.

When N builds up then get switch to production of antigenome and hence genome all which bind up N Protein

Virus must first make NP proteins. If it goes right into transcription the genome would be degraded! Viral proteins must protect anti genome and genome!

Question 18

Paramyxovirus Genome Replication

Answer 18

gRNA replicated from complete cRNA template, both in core structure. Higher levels of cRNA found in Paramyxovirus

Again the switch to replication from mRNA synthesis believed to be driven by high protein concentration

where high P levels allow P to act as chaperone protein filling in ICS regions with NP allowing a read-through to give cRNA using RNAP instead of transcribing mRNA. This is where auxiliary proteins work!

C/V/D genes products also believed to play role here

Question 19

Paramyxovirus Assembly & Release

Answer 19

Core formed in 2 steps

First NP protein binds at specific sites in leader sequences leading to encapsidation of gRNA (cRNA) during synthesis
P and L proteins then associate to serve as RDRP

As proteins build up, HN and F are inserted into golgi apparatus’ membrane, and r transported to plasma membrane and are glycosylated. As they are inserted into host membrane,
synthesis cycle and inserted into plasma membrane

M protein serves to dock core structure with viral membrane protein rich regions and virus buds through plasma membrane with HN protein serving in release of virus from cell

Core inserts near where HN and F are.

Question 20

Paramyxovirus Virus Budding

Answer 20

Fuzziness around the cell membrane is where HN proteins are formed. M proteins formed underneath, pulls F and HN around where a ribonucleic capsid core is associated with.

Question 21

Paramyxovirus Distinct Characteristics

Answer 21

RdRp in virion transcribes genome into mRNAs

Single transcriptional promoter at 3’of gRNA

mRNAs made by start-stop mechanism

Full length genome or anti-genome RNA is always present in assembled nucleocapsids

Some Paramyxoviruses edit P proteins using mRNA generating, distinct proteins

Some Paramyxoviruses induce cell-cell Fusions

Generate multinucleated cells syncytia

Question 22

Signs and symptoms of Nipah Virus infections

Answer 22

Human infections range from asymptomatic infection to fatal encephalitis.

Infected people initially develop influenza-like symptoms of fever, headaches, myalgia (muscle pain), vomiting and sore throat.

This can be followed by dizziness, drowsiness, altered consciousness, and neurological signs that indicate acute encephalitis.

Some people can also experience atypical pneumonia and severe respiratory problems, including acute respiratory distress.

Encephalitis and seizures occur in severe cases, progressing to coma within 24 to 48 hours.

The incubation period (interval from infection to onset of symptoms) varies from four to 45 days.

The case fatality rate is estimated at 40% to 75%; however, this rate can vary by outbreak depending on local capabilities for surveillance investigations.