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Retrotransposons & Retrovirus. CA García Sepúlveda MD PhD. Laboratorio de Genómica Viral y Humana Facultad de Medicina, Universidad Autónoma de San Luis Potosí. Session #27 Retroposons Introduction. Retroposon or Retro-transposons are transposons that mobilize via an RNA form.

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CA García Sepúlveda MD PhD

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Ca garc a sep lveda md phd

Retrotransposons &

Retrovirus

CA García Sepúlveda MD PhD

Laboratorio de Genómica Viral y HumanaFacultad de Medicina, Universidad Autónoma de San Luis Potosí


Session 27 retroposons introduction

Session #27 RetroposonsIntroduction

  • Retroposon or Retro-transposons are transposons that mobilize via an RNA form.

  • The DNA element is transcribed into RNA and then reverse-transcribed into DNA, which is inserted at a new site in the genome.

  • Transposition that involves an obligatory intermediate of RNA is unique to eukaryotes.

  • Some eukaryotic transposons are related to retroviral proviruses in their general organization.

  • Retroviruses insert DNA copies (proviruses) of an RNA viral genome into the chromosomes of a host cell.


Session 27 retroposons introduction1

Session #27 RetroposonsIntroduction

  • The life cycle of retroviruses and retroposons involve alternation of reverse transcription from RNA to DNA with transcription from DNA to RNA.

  • Only retroviruses can generate infectious particles.

  • Retroposons are confined to an intracellular cycle.

  • Retroposons & retrovirus share features with other transposons

    • short direct repeats of target DNA at the site of an insertion.


Session 27 retroposons retroviruses

Session #27 RetroposonsRetroviruses

dsRNA

ssRNA+

dsDNA

ssRNA-

RNAVirus

RT

DNAVirus

ssDNA

Virus


Session 27 retroposons retrovirus

Session #27 RetroposonsRetrovirus

  • Belong to the Retroviridae family of single-stranded RNA viruses that Reverse Transcribe (ssRNA-RT).

  • Enveloped viruses possessing an RNA genome.

AKA Pararetrovirus

dsDNA-RT

ssRNA-RT


Session 27 retroposons retrovirus1

Session #27 RetroposonsRetrovirus

  • Belong to the Retroviridae family of single-stranded RNA viruses that Reverse Transcribe (ssRNA-RT).

  • Enveloped viruses possessing an RNA genome.

  • Its members include:

ssRNA-RT


Session 27 retroposons retrovirus2

Session #27 RetroposonsRetrovirus

  • Belong to the Retroviridae family of single-stranded RNA viruses that Reverse Transcribe (ssRNA-RT).

  • Enveloped viruses possessing an RNA genome.

  • Its members include:

ssRNA-RT

  • Avian leukosis virus


Session 27 retroposons retrovirus3

Session #27 RetroposonsRetrovirus

  • Belong to the Retroviridae family of single-stranded RNA viruses that Reverse Transcribe (ssRNA-RT).

  • Enveloped viruses possessing an RNA genome.

  • Its members include:

ssRNA-RT

  • Mouse mammary tumour virus


Session 27 retroposons retrovirus4

Session #27 RetroposonsRetrovirus

  • Belong to the Retroviridae family of single-stranded RNA viruses that Reverse Transcribe (ssRNA-RT).

  • Enveloped viruses possessing an RNA genome.

  • Its members include:

ssRNA-RT

  • Murine & feline leukemia virus


Session 27 retroposons retrovirus5

Session #27 RetroposonsRetrovirus

  • Belong to the Retroviridae family of single-stranded RNA viruses that Reverse Transcribe (ssRNA-RT).

  • Enveloped viruses possessing an RNA genome.

  • Its members include:

ssRNA-RT

  • Human T-lymphotropic virus


Session 27 retroposons retrovirus6

Session #27 RetroposonsRetrovirus

  • Belong to the Retroviridae family of single-stranded RNA viruses that Reverse Transcribe (ssRNA-RT).

  • Enveloped viruses possessing an RNA genome.

  • Its members include:

ssRNA-RT

  • Walleye dermal sarcoma virus


Session 27 retroposons retrovirus7

Session #27 RetroposonsRetrovirus

  • Belong to the Retroviridae family of single-stranded RNA viruses that Reverse Transcribe (ssRNA-RT).

  • Enveloped viruses possessing an RNA genome.

  • Its members include:

ssRNA-RT

  • Human immunodeficiency virus 1


Session 27 retroposons retrovirus8

Session #27 RetroposonsRetrovirus

  • Belong to the Retroviridae family of single-stranded RNA viruses that Reverse Transcribe (ssRNA-RT).

  • Enveloped viruses possessing an RNA genome.

  • Its members include:

ssRNA-RT

  • Chimpanzee foamy virus


Session 27 retroposons retrovirus9

Session #27 RetroposonsRetrovirus

  • Rely on reverse transcriptase to perform the reverse transcription of its genome from RNA into DNA.

  • cDNA is integrated into the host's genome with the help of an integrase.

The virus then replicates as part of the cell's DNA, which means:

mRNA

Positive sense

5'-Capped

3'-Poly-A


Session 27 retroposons retrovirus10

Session #27 RetroposonsRetrovirus

  • These retrovirus are exogenous.

  • A retroviral sequence that is integrated in the germline remains in the cellular genome as an endogenous PROVIRUS.

  • Like a lysogenic bacteriophage, a provirus behaves as part of the genetic material of the organism.

  • They are inherited to the progeny.

ssRNA-RT


Session 27 retroposons retrovirus11

Session #27 RetroposonsRetrovirus

  • Retrovirus that have been incorporated to the genome are called ENDOGENOUS (ERV).

  • They are derived from ancient infections of germ cells in humans, mammals & other vertebrates.

  • ERVs make up 5-8% of the human genome (98,000 elements).

  • Most insertions have no known function (junk DNA) but some play important roles in host biology:

    • Control of gene transcription.

    • Control of cell fusion during placental development.

    • Resistance to exogenous retroviral infection.

    • Immunosuppresion


Session 27 retroposons retrovirus12

Session #27 RetroposonsRetrovirus

  • ERVs are activated during pregnancy in viviparous mammals (not Monotremes) and particularly during the implantation of the embryo.

They act as immunosupresors protecting the embryo from its mother's immune system.

Viral fusion proteins involved in the formation of placental syncytium limits cell migration (something an epithelium will not do well, as certain blood cells are able to diapedize).


Session 27 retroposons retrovirus13

Session #27 RetroposonsRetrovirus

  • ERV's are similar to HIV (which causes AIDS in humans).

  • The immunodepressive action was the initial normal behavior of the virus, as in HIV.

  • The fusion proteins were a way to spread the infection to other cells by merging them with the infected one, as in HIV.

  • It is believed that the ancestors of modern vivipary mammals evolved after an infection by this virus, enabling the fetus to survive the immune system of the mother.


Session 27 retroposons retrovirus14

Session #27 RetroposonsRetrovirus

  • 24 ERV families identified by Human Genome Project (HGP).

  • Broadly classified into 3 classes, on the basis of relatedness to exogenous genera:

  • Class I are similar to the gammaretroviruses

  • Class II are similar to the betaretroviruses & alpharetroviruses

  • Class III are similar to the spumaviruses


Session 27 retroposons retrovirus15

Session #27 RetroposonsRetrovirus

  • Most ERVs appear to be defective, containing nonsense mutations or major deletions which prevent them from producing infectious virus particles.

  • However, there is one family of viruses that have been active since the divergence of humans and chimpanzees [ hERV-K(HML2)].

  • Make up less than 1% of hERV elements but is one of the most studied.

  • There are indications it has been active in the past few hundred thousand years, as some humans carry more copies of the virus than others.


Session 27 retroposons retrovirus16

Session #27 RetroposonsRetrovirus

  • Human ERVs (hERVs) are suspected of involvement in some autoimmune diseases (multiple sclerosis).

  • Especially human endogenous retrovirus W known (MSRV).

  • Also a possible hERV involvement in the HELLP (Hemolytic anemia, Elevated Liver enzymes & Low Platelet count) syndrome & pre-eclampsia.

  • hERVs very likely associated with some types of schizophrenia.


Session 27 retroposons retrovirus17

Session #27 RetroposonsRetrovirus

  • Retroviruses have ssRNA genomes.


Session 27 retroposons retrovirus18

Session #27 RetroposonsRetrovirus

  • Retroviruses have ssRNA genomes.

  • Genomes are replicated through a dsDNA intermediate.


Session 27 retroposons retrovirus19

Session #27 RetroposonsRetrovirus

  • Retroviruses have ssRNA genomes.

  • Genomes are replicated through a dsDNA intermediate.

  • The life cycle involves an obligatory stage of integration in which the proviral dsDNA is inserted into the host genome by a transposition-like event that generates short direct repeats of target DNA.


Session 27 retroposons retrovirus20

Session #27 RetroposonsRetrovirus

  • The enzyme responsible for generating the initial DNA copy of the RNA is reverse transcriptase.

  • Converts the RNA into a linear (or circular) duplex of DNA in the cytoplasm of the infected cell.


Session 27 retroposons retrovirus21

Session #27 RetroposonsRetrovirus

  • The enzyme responsible for generating the initial DNA copy of the RNA is reverse transcriptase.

  • Converts the RNA into a linear (or circular) duplex of DNA in the cytoplasm of the infected cell.

  • The enzyme responsible for integrating the DNA copy is integrase.


Session 27 retroposons retrovirus22

Session #27 RetroposonsRetrovirus

  • Implications of proviral DNA integration:

    • A retroviral sequence that is integrated in the germline remains in the cellular genome as an endogenous provirus (behaves as part of the genetic material of the organism).

    • Cellular sequences may be transposed with proviral DNA and inserted into the genome as duplex sequences in new locations.

    • Retrovirally transposed cellular sequences may change the properties of the cell that becomes infected with the virus.


Session 27 retroposons hiv as an example

Session #27 RetroposonsHIV as an example

  • Tropism

  • Fusion

  • RT

  • Integration

  • Transcription

  • Assembly

  • Budding

  • Release

  • Maturation


Session 27 retroposons hiv as an example1

Session #27 RetroposonsHIV as an example

  • Tropism

  • Fusion

  • RT

  • Integration

  • Transcription

  • Assembly

  • Budding

  • Release

  • Maturation

Integration mediated by integrase.

Can occur at one or more sites.


Session 27 retroposons hiv as an example2

Session #27 RetroposonsHIV as an example

  • Tropism

  • Fusion

  • RT

  • Integration

  • Transcription

  • Assembly

  • Budding

  • Release

  • Maturation

Proviral DNA is transcribed by the host machinery to produce viral RNAs:

- mRNAs

- genomic RNA.


Session 27 retroposons hiv as an example3

Session #27 RetroposonsHIV as an example

  • Tropism

  • Fusion

  • RT

  • Integration

  • Transcription

  • Assembly

  • Budding

  • Release

  • Maturation

The enzymes reverse transcriptase and integrase are carried with the genome in the viral particle.


Session 27 retroposons hiv as an example4

Session #27 RetroposonsHIV as an example

Two copies of the RNA genome are packaged into each virion (diploid virus).

When a cell is infected by two different viruses,

it is possible to generate heterozygous virus particles carrying one genome of each type.


Session 27 retroposons retroviral translation processing

Session #27 RetroposonsRetroviral translation & processing

A typical retroviral sequence contains 3 or 4 "genes".

Gag-Pol-Env

Coding regions give rise to multiple proteins by processing reactions.

Retroviral mRNA has a conventional structure; it is capped at the 5′ end and polyadenylated at the 3′ end.


Session 27 retroposons retroviral translation processing1

Session #27 RetroposonsRetroviral translation & processing

Retroviral mRNAis represented by two subgenomic species.

1. Gag-Pol-Env (Full length)

2. Env


Session 27 retroposons retroviral translation processing gag

Session #27 RetroposonsRetroviral translation & processing (Gag)

The Gag product is translated by reading from the initiation codon to the first termination codon.

Gag derived products p6, p7, p17 & p24 are generated by post-translational processing.

Gag derived products have structural functions.


Session 27 retroposons retroviral translation processing pol

Session #27 RetroposonsRetroviral translation & processing (Pol)

To translate the Pol product, the first termination codon must be bypassed.

Depends on the “reading frame correspondence” between Gag and Pol.

If Gag and Pol are in the same reading frame...

Termination codon usage is suppressed by a glutamyl-tRNA, generating a Gag-Pol peptide.


Session 27 retroposons retroviral translation processing pol1

Session #27 RetroposonsRetroviral translation & processing (Pol)

Gag-Pol peptide is then processed to yield:

1- Gag derived products (p6, p7, p17 & p24).

2- Pol derived products (p10, p32, p51, p66).

Pol derived products have enzymatic functions.


Session 27 retroposons retroviral translation processing pol2

Session #27 RetroposonsRetroviral translation & processing (Pol)

When Gag & Pol are in different reading frames, a ribosomal frameshift occurs to generate a single protein.

Readthrough is ~5% efficient, so Gag protein outnumbers Gag-Pol protein by 20x.

←20x

←1x


Session 27 retroposons retroviral translation processing env

Session #27 RetroposonsRetroviral translation & processing (env)

The Env polyprotein is expressed by post-transcriptional splicing generating a shorter subgenomic messenger that is translated into the Env product.

Env products gp120 & gp41 are envelope proteins.


Session 27 retroposons retroviral translation processing2

Session #27 RetroposonsRetroviral translation & processing


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