Immunology &Virology II – Virology

Dr Mohammed Alsharifi
Vaccine Research Group, SBS
Virus Structure
What are viruses?
Structure of virus particles (virions)

Classification of viruses into families

Examples: Poliovirus, HCV, HBV, HIV, Influenza A virus, Herpes viruses

Pathogenesis of Virus infection

1. Transmission
2. Replication inside the host
Single cell infection
Virus Replication
3. Virus-host interaction [and outcomes]
Immune response
Courtesy of WHO
Transient and Persistent infection

Epidemiology of virus infections

Control of virus infections
Diagnosis of virus infections
http://www.healthoxygen.com
 Replication of DNA viruses

Priming of DNA synthesis

1. RNA - Okazaki fragments

2. DNA - hairpin structure

3. Protein - covalently attached to 5’ end

http://en.wikipedia.org/wiki/Okazaki_fragments

Parvoviridae

http://www.virology.ws/2010/12/10/unexpected-
endogenous-viruses/
 Two basic strategies for DNA replication

Strand displacement strategy

Adenoviruses (protein) 5’ 3’
Parvoviruses (DNA hairpin) 3’ 5’
Poxviruses (DNA hairpin)
5’

Primer 3’
3’ 5’
 Two basic strategies for DNA replication
Leading 3’

Replication fork strategy

5’
3’
Herpesviruses
Papillomaviruses
Lagging 5’

www.shmoop.com/dna/dna-replication.html
 Baltimore Classification
Strategy used by viruses to make mRNA

(+) ss RNA (-) ss DNA Parvoviridae - Parvovirus

Retroviridae - HIV
Adenoviridae - Adenovirus
(+/-) ds DNA Hepadnaviridae – HBV
Herpesviridae - VZV

Reoviridae - Rotavirus
Flaviviridae – HCV (+) RNA
Picornaviridae - Poliovirus = mRNA
(+/-) ds RNA

Orthomyxoviridae
(-) ss RNA Influenza A
 ADENOVIRUSES

http://gryllosblog.wordpress.com
Non-enveloped (Naked),
Icosahedral viruses with fibers at vertices
70nm diameter
Linear ds DNA genome (36-38 kbp)

DNA is associated with virally coded basic

proteins in virion (do not use cell histones to
package virion DNA)

47 known serotypes

Clinical infections:
Respiratory and gastroenteritis

http://www.hindawi.com/journals/ah/2013/176418/fig1/

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136627/pdf/pnas.201408462.pdf
 CAR: Coxsackievirus and Adenovirus Receptor.

http://arthritis-research.com/content/4/Suppl%203/S215/figure/F1?highres=y
Complex program of transcription from 7 early, intermediate
and late promoters
Transcription, replication and assembly
occur in the nucleus

pre-Terminal Protein (pTP)-Pol complex

http://viralzone.expasy.org/all_by_species/1940.html
http://www.microbiologytext.com/5th_ed/book/displayarticle/aid/479
 Hepadnaviridae: Hepatitis B Virus

Structure:
Enveloped virus, 42 nm
• Surface antigens - L, M & S
• Icosahedral NC - core protein (C)
• Virions contain DNA dep DNA pol
• (+/-) ds RC DNA genome, 3200 nt

http://microbiology2009.wikispaces.com/hepatitis+B
 Hepadnaviridae: Hepatitis B Virus

http://www.emedicinehealth.com
http://web.uct.ac.za/depts/mmi/stannard/emim
ages.html

HBsAg spheres &

filaments, 22 nm

Enveloped virus, 42 nm

DNA genome-containing virions & “empty” surface antigen

particles & HBeAg circulate in the bloodstream
 Hepatitis B Virus - Replication
Target cells: Hepatocytes (liver)
Receptor(s): NTCP
(Sodium/taurocholate co-transporting polypeptide)

Rc-DNA Entry: RM endocytosis

Viral genome:
cccDNA Relaxed circular (RC) DNA
genome
Packaging
Nucleus
RC DNA converted to a covalently closed
circular DNA (cccDNA)
RT
cccDNA binds histones stable
“mini chromosome”

Template for host RNA pol II

Overlapping open reading frames
Progeny virus
(ORF) present in (-) DNA strand

Pre-genomic RNA & mRNAs produced

 Hepatitis B virus genome - 3200 nt

Overlapping open reading frames (ORF)

present in (-) DNA strand “Increase
coding capacity”

Pre-genomic RNA 1.3 x genome size

• Pre-genome is reverse transcribed to
RC DNA
• Also acts as mRNA for C, Pol, &
HBeAg

Shorter mRNAs for S (HBsAg) & X

(HBxAg)
uni-kiel.de.infmed.research.hildt.fig.1
 Hepatitis B Virus - Replication

Rc-DNA

Cytoplasm
cccDNA Protein production
Virus polymerase (RdDp) produced
RdDp + pre-genomic RNA + core antigen = NC
Packaging RNA is Reverse Transcribed to RC (+/-) ds DNA
NC enveloped & released via ER golgi
or enter nucleus form pool of cccDNA
RT
RC DNA can enter nucleus
10-30 copies of cccDNA per
infected hepatocyte

Progeny virus
 Hepatitis B Virus - Replication

(+/-) ds RC DNA

(+/-) ds cccDNA

Host RNA pol II

(+) RNA pre-genome

& mRNA
RT
RNA dep DNA pol
RNaseH

(-) ss DNA

DNA dep DNA pol

http://www.intechopen.com/books/viral-replication/hepatitis-b-virus-genetic- (+/-) ds RC DNA

diversity-disease-pathogenesis
HBV infection of single cell

1. Attachment: Receptor
2. Entry: (RM Endocytosis)
3. Release of genome to nucleus
(RC DNA – cccDNA)
4. mRNA & proteins
5. Assembly of NC
6. Packaging RNA pregenome & RT
7. Reverse transcription inside NC
(RT)
8. Release by budding (ER/Golgi)
(no matrix)
Hepadnaviruses: e.g. HBV Retroviruses: e.g. HIV

DNA genome Dimeric RNA genome

Assembly

Reverse
Reverse Genomic RNA transcription
transcription
ccc DNA

Pre-genomic ds DNA
Integration
RNA
Assembly

Hepadnaviruses & Retroviruses replicate via reverse transcription

 Herpesviridae - Human Herpes Viruses
Herpes simplex Virus (HSV / HHV1, 2 / alpha)
• Cold sores, genital herpes
Varicella zoster virus (VZV/ HHV3 / alpha)
• Chickenpox, shingles
Epstein Barr Virus (EBV/ HHV4 / gamma)
• Infectious Mononucleosis (Glandular fever)
• Burkitt’s Lymphoma & Nasopharyngeal CA
Cytomegalovirus (CMV / HHV5 / beta)
• Congenital CMV
Human Herpes Virus 6 & 7 (HHV6 & HHV7 / beta)
• Infantile Febrile Illness
Karposi’s Sarcoma Herpes Virus (KSHV / HHV8 / gamma)
• Multiple Myeloma
 Herpesviridae
Herpes simplex (HSV / HHV1, 2)
• Cold sores, genital herpes
Varicella zoster virus (VZV/ HHV3)
• Chickenpox, shingles

Structure:
ds DNA genome,
Icosahedral NC (6 proteins)
Tegument proteins (10-20 proteins)
Enveloped (10 proteins)
Size 150-200 nm,

Genome:
(+/-) ds linear DNA, 125 kb

Budding:
Buds initially from nucleus then into
the ER and released by exocytosis

http://www.bio.davidson.edu
 Herpes viruses enter cells by surface fusion

Host cell infection:

HSV envelope binds heparan sulphate proteoglycans (HSPG)
VZV glycoprotein E (gE) binds to insulin-degrading enzyme (IDE)
Herpes viruses enter cells by surface fusion
Nucleocapsid & tegument proteins enter cytoplasm
Move by dynein-mediated transport to the nucleus
(+/-) ds DNA genome enters the nucleus via nuclear pore
 Herpes virus infection
Genome delivered to nucleus
Replication of DNA “rolling circle”

3 Classes of mRNA produced

• Immediate Early (IE) mRNA
• Early (E) mRNA
• Late (L) mRNA

3 Classes of proteins produced

• Regulatory (IE)
• Non-structural (E)
• Structural (L)

nature reviews microbiology 1794-fig 2 Lytic

Herpes mRNA Production & Replication
2
1 3
Early (E) mRNA
Immediate Early (IE) Late (L) mRNA
mRNA Non-Structural
proteins (E) Structural
Regulatory proteins (IE)
proteins (L)

RNA polymerase II
produces mRNA
(+/-) ds DNA

Rolling Circle Replication

DNA dep DNA pol
Structural Proteins
(+/-) ds DNA for new virions
Herpes replicates in nucleus via “rolling circle”

pathmicro.med.sc.edu mhunt dna18

Replication:
The linear genome is circularized by DNA ligase IV/XRCC4
Rolling circle replication generating concatemers of viral genomes
New genomes cleaved & packaged in NC
NC exist nucleus: 1st budding at the nuclear membrane
Release of herpesvirus - Lytic infection cycle
Herpes viruses replicate in the cell nucleus

NC budds at the nuclear membrane

“primary envelopment”
Virions lose envelope and teguments
proteins assemble to protect the NC
Tegument proteins coordinate 2nd
envelopment at the ER
Release of infectious virions by exocytosis

darwin.bio.uci.edu wagner
 Herpes virus infection of a single cell
e.g. epithelial cell in the skin

1. Attachment (Heparin Sulphate)

2. Entry (Surface fusion)
3. Release of genome (nucleus)
4. mRNA & proteins (IE, E, L)
5. Replication (Rolling circle)
6. Assembly of NC (icosahedral)
7. Packaging of genome & DdDp *
8. Release by budding - tegument
(enveloped 2x; nucleus & ER)

* Packaging of DdDp controversial

http://www.sciencedirect.com/science/article/pii/S01681
70204003247
Virology - Reading material
•Fenner and White’s Medical Virology, 5th Ed, 2017.
•Fields Virology, 6th Ed, 2014; 5th Ed, 2007.
•Lippincott Illustrated Reviews – Microbiology
•Prescott’s Microbiology; 8th Ed, 2011: Chapters 4, 5, 25, & 37
•Prescott, Harley, Klein’s Microbiology; 7th Ed, 2008: Chapters 16, 18, 36
& 37
•Principles of Virology: Molecular Biology, Pathogenesis & Control; S.J.
Flint et al, 2nd Ed, 2004; 1st Ed 2000.
•Muylaert I, Tang KW, Elias P. Replication and recombination of herpes
simplex virus DNA. J Biol Chem. 2011 May 6;286(18):15619-24. Review.