Quasispecies Theory and the Behavior of RNA

Viruses

Sumeeta Singh, Steve Bowers, Greg Rice, Tom McCarty

BINF 70402/19/13

What is a virus?• Viruses are obligate intracellular parasites.

• Small infectious agents bearing nucleic acid instructions.

• Classified based on the form of nucleic acid, DNA or RNA.

Virus types• Plus strand RNA virus• Minus strand RNA virus• Double strand RNA virus

• Retrovirus

• Single strand DNA virus• Double strand DNA virus

RNA Virus types

DNA Virus types

“Quasispecies Theory”• Mathematical framework describing

evolution of macromolecules (Eigen, 1971)

• Extends the classic population genetics ideas of mutation-selection into quasispecies (Eigen, Schuster, 1977)

• Eventually borrowed to describe RNA virus evolution dynamics

Virus Replication Error• Plus strand RNA virus• Minus strand RNA virus • Double strand RNA virus

• Retrovirus

• Single strand DNA virus• Double strand DNA virus

RNA dependent- RNA polymeraseError rate =10-3 to 10-5

DNA polymeraseError rate =10-7 to 10-9

Reverse TranscriptaseError rate =10-4 to 10-5

Requirements and Consequences• Polymerase responsible for high error

rates.• Estimated that each single and some

double nucleotide sequence changes occur.

• Resulting in a collection of related sequences around a “master” sequence.

• Variation is related to ability to survive (population genetics) AND probability of occurring based on sequence neighbors (quasispecies).

Virus Replication Swarm

Virus Replication Swarm

“Quasispecies Theory”1971

1977

Quasispecies and RNA viruses

• Survival of the “flattest”

• Error Catastrophe

• Fidelity and Fitness

Survival of the Fittest or Survival of the FlattestA flat species is a species which exists in a genetically diverse group. Not dominated by one variant.

A quasispecies must be a flat species.

A fit species (in this part of the presentation) is a species which reproduces very fast.

Flat Species

Properties of a flat species

A flat species will have high mutation rates.

A flat species is able to mutate without a major effect on fitness.

Advantages of a flat speciesDifferent mutants in a flat quasispecies can help each other.

Flat species are better able to adapt.

Dengue-1 VirusLives as a quasispeciesOne variant of the virus which is

found in high concentrations cannot survive on its own.

It can only survive because other viruses (infecting the same cell) create the protein which it lacks.

Viroid ExperimentTheory: More genetically diverse

(flatter) species are better able to adapt to mutations.

Viroid 1 - CSVd – very fitViroid 2- CChVMd – more flat, but less

fitProcedure: Infect plants with each viroid Subject the plants to two environment either normal, or UVC light

Results UVC light will cause mutationsResults: Under normal conditions the fitter viroid did better, then the flatter viroid. Under the UVC light the two were about equal Under the UVC light the flatter became more diverse. The fitter did not become more diverse.

Fitter or Flatter, which is better able to survive?The fit species will grow faster in

an ideal environment.The flat species will be able to

adapt more quickly.

Error ThresholdRNA Viruses have a high mutation rate

The point at which accumulated mutations reduce fitness: - Too much mutation can lead to loss of vital information - Too little mutation can lead to host defenses overcoming the virus

Error Threshold: position in informational space where a phase transition occurs such that the genomic sequence information can no longer be perpetuated.

The greatest fitness is when mutation rates approach the error threshold

Model of Error Catastrophe

http://www.pnas.org/content/98/12/6895.full

Error CatastropheExtinction of a virus as a result of excessive

RNA mutations – lethal mutagenesis

Decrease viral fitness by increasing the rate at which new mutations appear.

There is an intrinsic limit to the maximum variability of viral genetic information before it loses meaning. If an RNA virus quasispecies

goes beyond that mutation limit, the population will no longer be viable.

Increasing Mutation RateIonizing radiation (eg X-rays) –

cause mutations by damaging DNA.

Base Analogs – chemicals that replace one of the usual nucleotides in the DNA. These mutagens cause copying errors.

APOBEC3GHumans have the ability to

induce lethal mutagenesis Protein found inside cells that has

a very specific antiviral roleCytidine deaminase enzymeUnfortunately, HIV has the ability

to bind to APOBEC3G proteins and cause their degradation

APOBEC3G

It de-aminates the cytosine base, thus mutating it to a uracil base

APOBEC3G

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Fidelity and Fitness: Mutation Rate

• Evolutionary Theory: viral error rates

• RNA Virus: low fidelity generates diverse population of variants

• Homogenous population vs dynamic environment

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Fidelity and Fitness: Mutation Rate

• Ribavirin and Lethal Mutagenesis

• Hypothesis: mutant with low mutation rate less sensitive to LM and resistant to ribavirin

• Poliovirus experimental groups: G64S polymerase mutation

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Fidelity and Fitness: Mutation Rate/Pathogenicity

3D-G64S Virus Is Less Pathogenic than Wild-Type Virus in Mice

Competition between 3D-G64S and Wild-Type Virus in Mice

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Fidelity and Fitness: Virulence

• G64S species attenuated in transgenic mouse model for poliovirus infection

• Virulence determined by diversity of coinfecting population

• Quasispecies diversity, rather than the selection of individual variants, correlates with enhanced virulence

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Fidelity and Fitness: Attenuation

• Vaccine Design• Vignuzzi(2008): G64

engineered mutants stimulated high titers of neutralizing antibodies in mice

• Fidelity modulation as therapeutic strategy

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Future Perspectives

• How do models apply to infected hosts?• What is best measure of viral fitness in

dynamic population?• How does population diversity influence

pathogenesis (subpopulation cooperation)?

• Improved assays for characterizing viral populations

• Modern techniques: Deep sequencing, Molecular barcoding

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