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Transcript of The DNA -> RNA -> Protein Pathway year/Micro/powerpoint/Positive Strand... · The DNA -> RNA ->...
http://www.bioteach.ubc.ca/MolecularBiology/AMonksFlourishingGarden/
The DNA -> RNA -> Protein Pathway
RNA Polymerase =enzyme that makes
mRNA from theDNA gene template
•Plus-strand RNA viruses have genomes comprised of RNA. They haveno DNA in their replication cycles.
•In most cases, the replication cycle takes place in the cytoplasm of thecells.
•No involvement of the transcription machinery in the nucleus; so, theplus-strand RNA viruses produce their own enzymes for RNAtranscription and replication, which recognize RNA as the template.
•The plus-strand RNA viruses do use the host cell’s translation machineryto generate viral proteins.
•Many plus-strand RNA viruses produce numerous viral proteins from asingle “gene.”
Plus-strand RNA Viruses
Types of Plus-Strand RNA Viruses with single-stranded, non-segmentedplus-strand genomes
Picornaviruses(ex. Rhinovirus)
Flaviviruses(ex. West Nile Virus)
Longpolyprotein;subsequently
cleaved
OneYesNo+
(mRNA)I
CoronavirusesOne for eachmRNAMultipleYesNo
+(mRNA)
II
Example virus familyTypes of proteinproducts
Types ofmRNAs
RNA byitself
infectious?
Polymerasein Virions?
GenomePolarity
Class
Picornavirus Diseases
ENTEROVIRUSES (stable in GI tract)
•Poliomyelitis•Hepatitis A Virus•Coxsackieviruses•Enteroviruses Types 68-72•Echoviruses
RHINOVIRUSES (not stable in GI tract)
•Common Cold•Numerous subtypes
Poliovirus Pathogenesis
Only a fraction of patients develop paralytic disease
•Asymptomatic infection – 90%
•Abortive/minor illness – 5%
•Non-paralytic progression to the CNS – 1%-2%
•Paralytic poliovirus – 0.1%-2%
•3-4 days after minor illness
•Virus infects motor neurons in anterior horn of spinal cord and themotor cortex
Enterovirus Replication and DiseaseProgression
Primary InfectionMucosa and lymphoid tissues
Tonsils, pharynx, Peyer’s Patches
Viremia
Infection of secondary target tissue
Secondary replicationPhase
SYMPTOMS
Release of virus to environment
via fecal shedding
Protection by Antibodies
•Secretory antibodies can prevent primaryinfection
•Serum antibodies prevent viremic spread totarget tissues
Poliovirus Vaccination
•Salk Vaccine•Killed Virus (formalin inactivated)•IPV
•Sabin Vaccine•Live, attenuated viruses•OPV
•Attenuated strains can revert to virulent forms•Vaccinees shed the attenuated viruses in feces
•Plusses•Minuses
WHO Eradication of Poliovirus
Eradication campaign started in 1988Current target date for eradication: Beyond 2009
http://www.polioeradication.org/content/general/casemap.shtml
Total cases in 2006 Compared to same period in 2005
Globally 1403 1349
-in endemic countries: 1300 552
-in non-endemic countries: 103 797
WHO Eradication of Poliovirus:
2006 vs. 2005
http://www.polioeradication.org/casecount.asp
Is Eradication Possible?•Oral Poliovirus Vaccine
•Preferred for Third World Vaccination•Cheaper, No Sterile Needles Required•Campaign of usage has drastically reduced wild poliovirus transmission•Major issue is REVERSION of OPV strains to virulent forms
•Laboratory Stocks•Poliovirus present in many laboratories around the World•Poliovirus contaminants in stocks of other viruses, i.e. Coxsackievirus,Rhinovirus
•Bioterrorism•cDNA copies of poliovirus genome exist in many laboratories; transfectioninto tissue culture cells results in virus production•The poliovirus genome has been generated on a gene synthesizer (Plus-strand RNA viral genomes alone are infectious!)
Picornavirus Diseases
ENTEROVIRUSES (stable in GI tract)
•Poliomyelitis•Hepatitis A Virus•Coxsackieviruses•Enteroviruses Types 68-72•Echoviruses
RHINOVIRUSES (not stable in GI tract)
•Common Cold•Numerous subtypes
Human Rhinovirus Structure
AAAAAAAA
VPg
5’-NTROpen Reading Frame Encoding Polyprotein
~7200 bases
~630 nts
http://www.rcsb.org/pdb/molecules/pdb20_3.html
Human Rhinovirus Structure
Virus exterior = protein shell
Icosahedral shape
Cross-section
Viral RNA genome inside
Protein capsid layer outside
http://www.rcsb.org/pdb/molecules/pdb20_3.html
Rhinovirus Binding to Receptor and Antibodies
ICAM-1 RECEPTORBOUND TO RHINOVIRUS
ANTIBODIES BOUNDTO RHINOVIRUS
http://rhino.bocklabs.wisc.edu/virusworld/images/r14_rhino+receptor.jpg
ICAM-1 Receptor Binds to Canyon onRhinovirus Surface
http://www.ag.uidaho.edu/mmbb/p_gustin_k.htm
Rhinovirus Polyprotein Processing: Method forGeneration of Individual Viral Proteins
Rhinovirus Pathogenesis•Entry of virus into upper respiratory tract
•Hands and fomites•Inhalation of droplets that contain virus
•Unable to replicate in the GI tract (not stable to acid)
•Receptor is ICAM-1 (Intercellular Adhesion Molecule 1)
•Preferentially replicates at 33oC
•Over 100 serotypes•Reason for repeat infections throughout lifetime•Antigenic sites change, but receptor-binding site is protected
New Rhinovirus Therapies62 million cases of Rhinovirus infection in U.S. each year
Cause more than 50% of respiratory tract infections
•Traditional vaccine approaches as with poliovirus not possible (toomany serotypes), so antiviral therapies must exploit unique properties ofthe virus
•Soluble ICAM-1: Blocks virus binding to its receptor
•AG7088: 3C protease inhibitor (being formulated for intranasaldelivery)
•Pleconaril: Binds a pocket in the capsid; interferes with attachment anduncoating
•Promising early clinical studies – reduction in disease length•Not approved due to possible drug-drug interactions•Reformulations being developed
Figure 1. Genomic structure of flaviviruses. The flavivirus genome is 11,000 to 12,000 nucleotides long. Both the 5'- and 3'- ends containnoncoding (NC) regions. The genome encodes 10 proteins, 3 of which are structural proteins (C, M, and E), and 7 of which arenonstructural proteins (NS1, NS2a, NS2b, NS3, NS4a, NS4b, and NS5). The M protein is synthesized as a precursor (prM) protein. The prMprotein is processed to pr + M protein late in the virus maturation by a convertase enzyme (furin).
http://www.cdc.gov/ncidod/eid/vol7no4/petersenG1.htm
Flavivirus Gene Structure
(West Nile Virus = member ofFlavivirus family)
http://www.cdc.gov/ncidod/eid/vol7no4/petersenG2.htmhttp://news.uns.purdue.edu/UNS/html4ever/031009.Kuhn.westnile.html
West Nile Virus Structure
Figure 2 The WNV replication cycle. A. Attachment and entry of the virion. B. Uncoating and translation of the virion RNA. C.Proteolytic processing of the polyprotein. D. Synthesis of the minus-strand RNA from the virion RNA. E. Synthesis of nascentgenome RNA from the minus-strand RNA. F. Transport of structural proteins to cytoplasmic vesicle membranes. G.Encapsidation of nascent genome RNA and budding of nascent virions. H. Movement of nascent virions to the cell surface. I.Release of nascent virions. SHA, slowly sedimenting hemagglutinin, a subviral particle that is also sometimes released.
http://arjournals.annualreviews.org/doi/full/10.1146/annurev.micro.56.012302.160654;jsessionid=i8FGn3DIgEu8
West Nile Virus Replication Cycle
*The WNV receptorin vertebrate cells isαVβ3 integrin, whichis highly conserved invertebrates
Coronaviruses
•Another virus group with members responsible for the “common cold”
•Coronavirus member in the news = SARS Virus
http://www.lapublichealth.org/acd/images/SARS_CoV_Picture.gif
http://www.csic.es/hispano/charlas/2002/m1/m1abr-02/m1abr-02.htm
http://www.cmlab.csie.ntu.edu.tw/~jsyeh/SARS/images/coronavirus-Alankann.gif
Coronaviruses
Coronavirus Receptor = ACE-2
http://www.rndsystems.com/DAM_public/5531.gif
ACE-2 = angiotensin-converting enzyme 2
Expressed in heart, lung, kidney, GI tract
Coronavirus Spike Proteins:Virulence Factors
http://www.nyas.columbia.edu/sars/web/slide_presentations/s3_2/04.html