Transcription I Flashcards
What are the differences btw RNA and DNA?
RNA
-
D-ribose instead of 2’-deoxyribose
→ can be hydrolyzed by alkalis to 2’, 3’ cyclic diesters due to presence of 2’-OH group - uracil instead of thymine
- usually single strand instead of double strand (except in certain viruses)
List different types of RNA.
protein coding RNAs
- mRNA: templates for protein synthesis
nonprotein coding RNAs
- rRNA: form ribosomes
- tRNA: adapter molecules for translation
- small RNA: see own card
- ribozymes: catalyticly active
List different types of small RNA.
- snRNA
- snoRNA
- miRNA
- siRNA
- piwiRNA
What is snRNA?
Where can it be found + function?
small nuclear RNA
- in eukaryotes, archae
- involved in splicing of pre-mRNA
What is snoRNA?
Where can it be found + function?
small nucleolar RNA
- in eukaryotes, archae
- involved in chemical modification of rRNA/tRNA/snRNA
What is miRNA?
Where can it be found + function?
micro RNA
- in eukaryotes
- inhibition of translation of mRNAs
What is siRNA?
Where can it be found + function?
small interfering/silencing RNA
- in eukaryotes
- cause degradation of RNA molecules due to hybridization (= silencing)
What is piwiRNA?
Where can it be found + function?
Piwi-interacting RNA
- in animals
- protection against transposons
What are differences btw RNA and DNA synthesis, hence transcription and DNA replication?
in RNA synthesis
- ribonucleotides used
- U instead of T
- no primer necessary, initiation starts de novo
- only portions transcribed, genes can overlap to code for different proteins
- no proofreading → lower fidelity
Differentiate btw coding, non-coding, and template strand.
-
template strand: DNA strand that is transcribed
= non-coding/antisense strand -
coding strand: DNA strand strand which has the same base sequence as the RNA transcript produced (obv T instead of U)
= non-template/sense strand
NOTE: both strands can be used as templates though
What is asymmetric transcription?
both DNA strands could be used as templates, but transcription would be happening in opposite directions (still both in 5’→3’ direction)
just like DNA replication: synthesized in 5’→3’ direction, template read in 3’→5’
Describe the basic mechanism of transcription.
- requires the unwinding of the double helix at gene that should be transcribed
-
RNA polymerase
- catalyzes the formation of phosphodiester bonds between nucleotides
- moves stepwise along the DNA extending the RNA chain as it goes
- as it moves it unwinds the next part of the helix
- helix behind RNA polymerase closes and the mRNA is displaced
What are different features of DNA/RNA polymerase?
RNA polymerase
- has intrinsic helicase activity
- only works on one strand
- does not require priming
- does not proofread
How many types of RNA polymerase (RNAP) are there in prokaryotes?
Describe its structure.
only one type
4 subunits:
- 2α: bind regulatory sequences
- β: forms phosphodiester bond
- β’: binds DNA template and σ factors
What are σ (sigma) factors, what do they do?
DNA-binding proteins, bind to β’ of prokarytoic RNAP to form RNAP holoenzyme Eσ
⇒ allow recognition of promoter
NOTE: σ factor considered an own subunit, so 5 subunits now
Describe the molecular mechanism how RNAP holoenzyme Eσ binds to the DNA.
recognizes atomic pattern in major groove of DNA (depending on base pairing)
- acts as H donor, or acceptor, forming H-bridges
- recognizes methyl groups, hydrophobic interactions
- recognizes H atom, unavailable for H-bonding
What does the +/- numbering system indicate?
- = upstream
- = downstream
⇒ way of defining the location of regulatory elements in a gene
How do you call the site on the DNA template where RNAP holoenzyme Eσ binds?
Structure.
promoter, upstream of transcription start site TSS
- -35 region w/ consensus sequence TTGACA
- 16-19bp spacer
- -10 region, Pribnow box
What is the Pribnow box?
AT-rich sequence in prokaryotic promoters, easier unwinding due to only 2 H-bonds
→ similar to TATA box in eukaryotes
Differentiate btw types of σ factors.
recognize different promoters + activate transcription of different genes
- sigma 70: housekeeping sigma factor (normal one)
- sigma 54: nitrogen-limitation sigma factor
- sigma 38: starvation phase sigma factor, activates gene involved in long term survival (i.e. peroxisomes)
- sigma 32: heat shock sigma factor
NOTE: variety of σ factors + competition for RNAP determines much of cell’s protein content
Which method is used to search for promoters on DNA?
footprinting