Replication of Replication of Medium DNA Medium DNA

Virus;Virus;ssDNA VirusssDNA Virus

Replication of Replication of Medium DNA Medium DNA

Virus;Virus;ssDNA VirusssDNA Virus

Family Adenoviridae• “gland” lymphoid tissue in nasopharynx• Icosahedral capsid, 70-90 nm• Fiber projections (ligand for attachment)• Some adenovirus produce tumors in

experimental animals (hamsters)

Genus: Mastadenovirus

• “breast” • Mammalian species• Human adenovirus 1-50• Monkeys, horses, cows, pigs, dogs

Genus: Aviadenovirus• “bird” • Avian species

Adenovirus• Isolated in 1953 when searching for

common cold virus in tonsil & adenoid tissue culture

• Latent virus activated, replicates, CPE (cytopathic effect) of cells

Adenovirus Infections• Upper & lower respiratory infection

(URTI, LRTI)• Epidemic conjunctivitis• Gastroenteritis• Acute hemorrhagic cystitis• Lytic, chronic, latent, transform

cells

URTI: “Cold”• Rhinovirus• Coronavirus• Adenovirus• Parainfluenza virus

URTI: Pharyngitis• Adenovirus• Herpes simplex virus (HSV)• Enterovirus• Epstein-Barr virus (EBV)• Less likely: Influenza virus,

Parainfluenza virus, Rhinovirus, Coronavirus

URTI/LRTI: Croup (Laryngotracheobronchiti

s)

• Parainfluenza virus• Respiratory syncytial virus

LRTI: Bronchiolitis• Parainfluenza virus• Respiratory syncytial virus (RSV)

LRTI: Pneumonia (Lung)

• Influenza virus• Respiratory syncytial virus (RSV)• Parainfluenza virus• Adenovirus virus• Coronavirus

“Cold” Virus Symptoms• Fever• Cough• Headache• Sore throat• Muscle ache• Fatigue• ± Vomiting, diarrhea

Feels Like a “Cold” But It’s Not! See Clinician

if:• Fever >103°F• Fever 101°F for more than 3 days• Yellow/green discharge from lungs• Extreme difficulty swallowing• Shortness of breath• Chest pains• Earache, discharge from ears, hearing

loss

Adenovirus Genome: dsDNA

• Linear, 36 kbp• Ends have inverted terminal repeats (ssDNA

forms circular structure)• 5’ end - viral terminal protein (TP, 50 Kd)• Genes found on both DNA strands• Express proteins for: cell activation, DNA

synthesis, suppress host immune & inflammatory response, structural capsid

Adenovirus“Early” mRNA

• From both DNA strands using cell RNA pol II– “right” strand transcription (--->)– “left” strand transcription (<---)

• Four early transcription “units”: E1 – E4– each with separate promoter– E1 enhanced promoter activated by host cell DNA binding proteins

• mRNAs: 5’cap, 3’polyA tail, post-transcriptionally spliced

Adenovirus: “Early” Proteins

• E1 – transformation function, binds cell repressor proteins p105RB & p53

• E2 – DNA pol, TP, DNA-binding protein (shuts off early promoter)

• E3 – decreases host MHC proteins & inflammation

• E4 – decrease host T cell recognition

Host Cell Activation • Adenovirus E1 protein binds cell p105

RB protein• p105 RB no longer functions to down

regulate cell transcription• Cell remains active

Adenovirus DNA Replication

• “Continuous” DNA strand synthesis

• Replication complex:– viral DNA pol– TP (primer)– DNA-binding proteins

(viral & cellular)• Type 1 Replication:

– #1 strand copied into dsDNA

– #2 strand displaced into ssDNA intermediate

• Type 2 Replication: displaced ssDNA copied into dsDNA

Adenovirus: “Late” mRNA

• From one DNA strand – “right” strand transcription (--->)

• Single promoter: leader + five mRNAs (L1 – L5) by alternative splicing

• Translate mainly for structural proteins

Adenovirus: Splicing of mRNA

• First observation for splicing of eukaryotic mRNA

• Hybridize late “hexon” mRNA with single strand viral DNA

• EM observation that ds RNA:DNA not co-linear, with ssDNA “loops”

• Deduce that mRNA must be shorter (“exons” – 1,2,3 leader, hexon) and lack certain DNA regions (“introns”- A,B,C)

Adenovirus: Virus-Associated (VA) RNA

• By cell RNA pol III• Short RNA molecule blocks

antiviral action of interferon (stops activation of PKR)

• Also, E1A protein blocks interferon signal transduction

• Role in persistent viral infection

Adenoviral Vector for Gene Replacement

Therapy• Eliminate & cut out viral pathogenic gene• Insert therapeutic gene, i.e. insulin gene• Infect patient with virus vector to

produce insulin in pancreas• Complications due to adenovirus

infection of various other organs, results in toxicity

• Vector DNA eventually degraded by cell

Family Parvoviridae

• “small”• ssDNA, 5.5 kb• icosahedral capsid,

18-26 nm

• Genus: Parvovirus – mammals and birds• Genus: Dependovirus (adeno-associated

viruses) requires “helper” adenovirus for replication

• Genus: Densovirus - insects

Human Parvovirus B19• Infects mitotically active

erythroid precursor cells in bone marrow

• Erythemia infectiosum (fifth disease) in children – mild flu-like, “slapped cheek” rash

• Aplastic crisis in hemolytic anemia and in sickle-cell patients

• Associated with rheumatoid arthritis

Reading & Questions• Chapter 16: Replication Strategies

of Small and Medium-Sized DNA Viruses

Class Discussion – Lecture 10

• 1. Is there any similarity in function of adenovirus and SV40 virus “early” proteins?

• 2. Although adenovirus has a dsDNA genome, why can’t you assign polarity (+/-) to the two strands?

• 3. As human parvovirus, unlike SV40, does not have the ability to activate its host cell, how does it insure it will infect a cell that is actively dividing?