Lecture 10: Horizontal & Viral Genetics

Where did the placenta originate from?

Where did the placenta originate from?

Syncytin gene (repurposed ortholog of the retro viral env gene)

What is horizontal gene transfer?

The transfer of a genetic element from one organism to another within the same generation

Horizontal transmission has directionality meaning

There is a donor and recipient

Prokaryotes carry their genome as a _____, and some bacteria harbor extra chromosomal genetic elements known as _____

Nucleoid, Plasmids

What three common mechanisms of horizontal transfer are in bacteria?

Transformation, Conjugation, and Transduction

The simplest type of horizontal inheritance is transformation. Define transformation

Bacterial cells acquire and incorporate DNA from their environment

What is the name of bacteria which are able to acquire and incorporate DNA from the environment?

Competent cells

Are competent bacterial species rare?

Yes, ~100 species are actually competent

Transformation Process

1. Free DNA is digested while the other strand is taken up

2. the DNA is replicated

3. The dsDNA is incorporated

Transformation in the lab is an example of what type of selection?

Artificial selection which is a type of Strong, positive selection

What is conjugation?

Direct transfer of DNA from donor to recipient through physical contact

In conjugation what is donated?

The donor transfers a F+ plasmid through a pilus so both cells become F+ cells.

Self transmissible vs mobilizable

Self transmissible can create a pilus and move through pilus itself. A mobilizable plasmid depends on another F+ facto to form a pilus

What is transduction?

Foreign DNA is introduced to a cell by a viral vector

Who is transduction important to?

Bacteria, plants, and animals

Transduction cycle consists of 6 steps what are they?

1. Attachment-a virus attaches to the surface of a host cell

2. Entry- the virus or viral genome enters the host cell

3. Integration- some but not all viruses integrate their genome into host genome

4. Synthesis of viral components- viral proteins,DNA, or RNA are made by host

5. Viral Assembly-viral components assemble into virus particles

6. Release-viruses are released from the host cell

What is the lytic cycle?

Lysis and death of the host cell releases phages

What is the Lysogenic cycle?

Generations of bacteria divide with a prophage hidden inside the nucleoid

What are emerging viruses?

Viruses that have arisen recently and are more adapted to infect and damage cells than previous strains

Viral size, virus structure, genome size, and genome characters of HIV (Retroviridae)

120 nm, enveloped, 9200 bp, diploid + sense ssRNA, replicated via intermediate

Viral size, virus structure, genome size, and genome characters of Influenza (Orthomyxoviridae)

100 nm, helical capsid, 13500 bp, - sense ssRNA

Viral size, virus structure, genome size, and genome characters of SARS-CoV-2 (Coronaviridae)

120 nm, helical capsid, 30,000 bp, +sense ssRNA, no DNA intermediate

Positive sense vs Negative sense

Positive sense is 5’-3’, viral RNA can serve as mRNA and be directly translated into viral proteins.

Negative sense is 3’-5’, must first be converted to a positive RNA before being infectious

HIV (human immunodeficiency virus) causes AIDS, it has a diploid +ssRNA genome and uses what type of intermediate?

Double stranded DNA intermediate

HIV structure

HIV genome

What is the gag gene?

Stands for group-specific antigen and encodes a polyprotein used for viral assembly and capsid formation

What proteins are within the gag gene

Matrix protein, nucleocapsid protein, capsid protein, and p6

What is the pol gene?

Stands for polymerase and encodes a polyprotein which is cleaved into enzymes needed for viral replication and viral assembly

What enzymes does the pol gene encode

Integrase, reverse transcriptase, HIV protease

What is the env gene

Short for envelop, encodes a polyprotein which when cleaved has proteins that are part of the viral envelop

What proteins does the env gene encode

Gp 120 and gp 41

HIV reverse transcriptase contains.. which degrades RNA

RNase H

Reverse transcriptase is terrible at making DNA leading to

High mutation rates

During HIV integration, reverse transcriptase uses which two structures/sequences as primers?

PBS (with tRNA) and PPT (polypurine tract) as both are resistant to RNase H but ultimately are degraded by Rhase H

Reverse transcriptase makes _____ _____

DsDNA provirus

When is HIV DNA called a provirus?

After integration

the activation of the provirus occurs via

NF-kB, a transcription factor

Function of the Tat protein

Reenters the nucleus and up regulates viral transcription

Function of the Rev protein

Reenters the nucleus and is critical for exporting larger mRNAs

Function of the Nef proteins

Maintains T-cell activity and prevents defense against the virus

Incompletely spliced HIV mRNAs

1. Vif disrupts antiviral activity

2. Vpr is necessary for the pre integration complex

3. Vpu releases immature HIV particles from the host cell membrane and is incorporated into the golgi

4. Env polyprotein is broken down into gp 120 and gp 41

Host proteases process Env to create gp 120 and gp 41 to make up which structure

The spike

Unspliced HIV mRNAs

1. Translated to make Gag and Gag-pol

2. Rarely the stop codon on Gag is not recognized resulting in the full Gag-pol polyprotein

HIV assembly

Which virus has killed more humans than any other virus?

Influenza epidemics

Influenza Structure

What are H spikes?

Fusion glycoprotein for entering host cells

What are N spikes

Receptor-destroying enzyme for exiting host cells

Influenza cap snatching

Oythomoxyviruses likely originated in

Invertebrates

Influenza host are…..

Ducks, shorebirds, gulls, pigs, bats

How many subtypes of influenza are there in birds?

16 HA and 9 NA

Reassortment of RNA segments results in altered surface proteins, such an event is called

Antigenic shift

The defining process of influenza virus a in humans are mutations to the -ssRNA code, such an event is known as

Antigenic drift

What type of selection exists for the following mutants: HA spikes that allow the virus to enter, NA spikes that allow the virus to exit, and viruses that evade host immune systems

Positive directional selection

What does SARS-CoV-2 stand for?

Severe acute respiratory syndrome coronavirus 2

Coronavirus structure

Coronavirus Genome

Hallmark of coronaviruses

The translation of two large non structural poly proteins through a programmed -1bp ribosomal frame shift

During a programmed -1bp ribosomal frame shift, what happens?

A sequence has an adenine that is used twice, once as the last nucleotide of a codon and the 2nd time as the first nucleotide of the next codon.

What is essential for the frame shift to occur?

The RNA pseudoknot

Polyproteins pp1a and pp1ab undergo what process to become 16 enzymes?

Proteolysis

Genes ORF1 and ORF 1b encode many of the proteins that make the

Replicase-transcriptase complex

+ / - gRNA vs + / - sgRNAs

+ / - gRNAS are replicated and the +ssRNA is packaged into virions

- sgRNAs are created from +gRNA through discontinuous synthesis. -sgRNA eventually make +sgmRNAs that are translated into structural proteins

What is discontinuous synthesis?

+ssRNA genome is used as a template by the replicaser to from replicative forms

Reverse transcriptase mutation rate is highest in which virus?

HIV

HIV is diploid, why is that important?

Heterozygous strands can recombine

If you scanned your own genome which viral sequence would you expect to find?

HIV

Who discovered transposable elements

Barbara McClintock

What are transposable elements

Nucleic acids capable of moving among locations within the genome

Retrotransposons vs Transposons

Retrotransposons: RNA elements

Transposons:DNA elements

Eukaryotes vs Prokaryotes

Eukaryotes: Type 1 Retrotransposons which are typically mutated retroviruses and reverse transcribe themselves into the genome through a copy and paste method. Type 2 transposons move by the enzyme transposase which uses a cut and paste method

Prokaryotes: Insertion sequences move by the enzyme transposase through cut and paste method. Transposons have complex end sequences; cut and paste method.

50% of the genome is … and … make up more than 40% of the genome

Transposons and Retrotransposons

What is the most common element in the genomes of many species?

Transposable elements

What are restriction endonucleases

Introduce double stranded breaks into foreign DNA of viruses and plasmids

What does CRISPR stand for?

Clustered Regularly Interspersed Short Palindromic Repeats

What are spacers?

Sequences used as the host cell as memories of previous infections, and are acquired from phages or plasmids that don’t lyse the cell.

What are Cas genes and how many are there?

CRISPR associated genes and there are roughly 93

What is Cas9?

Bacterial RNA-guided DNA endonuclease that defends against foreign viruses and plasmids

The development of CRIPSR was recognized by the Nobel prize which was awarded to

Emmanuelle Charpentier and Jennifer Doudna

CRISPR-Cas system in eukaryotes

Eukaryotes natural repair double strand breaks, NHEJ can produce indels through a single strand break which results in a loss of function. HDR uses a desirable DNA sequence as a template for repair and may be incorporated into the genome

In humans what protein recognizes DNA-RNA retrovirus hybrids?

APOBEC3G

How does APOBEC3G deal with proviruses?

APOBEC3G causes mutations to accumulate in the provirus by deaminating the DNA from cytosines to uracil