Transposition 3 Flashcards
what is the retrovirus life cycle and its RNA to DNA genome transitions.
- RNA goes to DNA and then back to RNA
- The infectious particle is a single strand RNA
- Must be turned into DNA and integrate into host genome before turned back into RNA
- RNA to DNA is facilitated by the protein called reverse transcriptase - viral enzyme, DNA polymerase, uses RNA as a template
- Becomes linear DNA
- Integrated into host genome - enzyme that puts it into host genome is called integrase
- When inside host genome, uses host RNA pol 2 to make a mRNA and follows normal DNA transcription to make the RNA
What is provirus?
When linear DNA integrates into host genome
what are the components of the retrovirus?
RU5
U3R
complete long terminal repeat
gag
structural matrix protein in the core of the viral particle
pol
reverse transcriptase; where all enzymes are encoded
env
encodes what is exposed to environment, spike and stem on the spike
where does the membrane come from in retroviruses?
from the host
what occurs in the cytoplasm? nucleus?
- C: RNA reverse transcription into linear DNA
- N: linear DNA integration into nucleus, provirus made, and transcription into RNA from host RNA polymerase
what protein products does gag produce?
- matrix
- capsid
- nucleocapsid
what protein products does pol produce?
- protease
- reverse transcriptase
- integrase
what protein products does env produce?
- surface protein
- transmembrane
what are LTRs?
long terminal repeats
- RU5 and U3R
what activities does retrovirus emply?
- splicing
- suppression and frameshifting (suppress termination, rRNA code for amino acid instead of stop)
- processing of initial protein into several
How are LTRs and direct repeats generated during the retrovirus life cycle?
- Linear form is the only reconstituted form - not the integrated because there are some bases missing there
U3 and U5 loses 2 bp - DNA has to turned into an RNA infectious particle
- Host RNA polymerase
- Need a promoter in DNA to start transcription
- Promoter does not end up in RNA, needs to have stop
- LTR is the mechanism in which the ends are generated in order to make a transcription product
- Positive strand is missing start and stop
- Reverse transcriptase from RNA generates linear DNA with enact ends with same beginning and ends
- Integrase deletes two bases on both ends, so the provirus is also defected
synthesis of the minus strand
- Infectious +RNA, missing U3 and U5 on each ends
- Starts DNA synthesis on the edge of 5’ and makes a copy of LTR and jumps to the other strand of the DNA molecule
- Cellular tRNA (uncharged glutamate from the host), 10 bp homology so tRNA will form a hybrid
- Primes off the 3’ end of the tRNA - strong stop minus DNA
- Discrete size - starts and stops at specific place
- Strong stop region and reverse transcriptase changes template
- Jumps because there is degradation in positive RNA and is a signal for new piece of minus DNA to jump to the other end
- This is now the primer and goes all the way across the DNA
- Minus strand is complete on the right LTR
synthesis of plus strand
- Minus strand DNA made
- tRNA primer cut off
- Replace positive strand with plus strand DNA
- RNAH is packaged in nucleic acid and randomly regrades RNA and leaves gaps
- The remaining RNA acts as a primer to synthesize DNA
- Partial copy of strand and jumps and synthesizes the rest of the way from the opposite side
- Hybridizes the R and U5 and synthesizes it to the other side so it is complete
- Minus strand DNA completed
- Double stranded retrovirus ready for integration
what is the primer in the minus strand?
a specific cellular tRNA acts as a primer of reverse transcriptase
what is the primer in the positive strand?
RNA is degraded by RNaseH and the random spaces leftover are primers for partial synthesis
copy choice
- jumping of reverse transcriptase onto other templates to form recombination events in RNA
- since the virus particle contains two copies of the positive RNA, recombination is always possible by jumping to another template
- not thought to occur in regular host-mediated recombination
integrase
- catalyzes all the stages of integration
- the small inverted repeats at the ends of the provirus DNA are recognized by integrase as sites to cut 2 bases from the 3’ end
can you “catch” cancer?
- retroviruses may acquire cellular ONC genes and form defective viruses that need helper viruses to replicate
- the v-onc proteins are initially expressed as gag0fusion proteins
- transducing viruses often cause unrestricted cell growth in the infected tissues
what is the complete form of a retrovirus?
linear DNA
How are 2 nt lost during the integration process? From which strand?
the small inverted repeats at the ends of the provirus DNA are recognized by integrase as site to cut
- 2 bases from the 3’ ends
Which genes are missing in a transducing retrovirus?
pol and env
what are some examples of retrotransposons?
- Ty in yeast
- copia in drosophila
what type of element has no infectious particle, but still produces a particle?
Retrotransposon
how are retrotransposons produced?
- two RNAs are produced: TyA protein (gag-related) encodes a capsid protein
- TyB (pol related) encodes protease, RT and integrase
- Ty A and TyB ORF overlap by 13 amino acids, ribosome has to frameshift to synthesize TyAB proteins
- no env because it does not make an infectious particle
what does the yeast Ty particles contain?
capsid protein packages viral RNA and transports it to the nucleus
- full length RNA
- reverse transcriptase
- dsDNA
- TyB (integrase activity)
- capsid
arc protein
related to gag capsid protein
- involved as connection hub in neural function plasticity mammals
- required for formation of mRNA containing extracellular vesicles in neurons
LTR-retrotransposons
- encode reverse transcriptase and integrase
- no infectious particle
- copia and Ty are examples
what are the three classes retroelements fall into?
- LTR retrotransposons
- non-LTR retrotransposons
- SINES
non-LTR retrotransposons
- encode reverse transcriptase, nucleic acid binding protein, and endonuclease
- no LTR
- derived from RNA pol 2 transcripts
- example: LINES, L1
SINES
- short-interspersed nuclear elements
- no LTRs
- derived from RNA pol 3 transcripts
- always nonautonomous
- encode tRNAs, 7SL RNA, an d5S RNA
- example: Alu
- may use LINES to replicate
SINES are derived from the transcripts of which host RNA polymerase?
RNA pol 3
how large and how many Alu repeats are there?
- over 1 million in human genome
- 200 bp (7SL RNA)
what gene block is missing from every type other than retroviruses?
env
what gene does Ty and Copia have that is missing in the LINES?
long terminal repeats
what is the unique protein present only in LINES?
nuclear binding protein
what is the advantage of having a cis-acting mechanism for protein binding?
To ensure propagation
what is the role of the endonuclease activity?
Endonuclease ensures the nuclear binding protein and RNA can integrate back into the hot nucleus and DNA to be copied again
Which two classes of Retroelements are most abundant in the human genome?
LINES (17%) and SINES (15%)
Exaptation
adapt something that originally had a function into something asa completely different function
- arc protein
Exaptation
adapt something that originally had a function into something asa completely different function
- arc protein
