Transcription

Question 1

what do all cells have…

Answer 1

• All cells have the same DNA content (with the exception of gametes, RBCs, T cells and B cells which have their antibody genes rearranged)

Question 2

why is the control of gene expression important

Answer 2

• The control of gene expression is fundamental to the development and maintenance of multicellular organisms, the correct temporal and spatial expression of specific genes governs the identity of a cell and defines its function

Question 3

what control elements are in the nucleus

Answer 3

• There is transcriptional control between converting DNA to pre – mRNA
• There is processing control between converting primary transcript to mRNA

Question 4

what control elements are in the cytoplasm

Answer 4

• mRNA moves out through the nuclear membrane pores this is controlled by transport control
• mRNA is then either degraded which happens by mRNA stability
• or turned into an active protein this happens by translation control
• the active protein is then converted into an inactive protein by protein activity control

Question 5

what are the control elements are there overall

Answer 5

In the nucleus – this is what this lecture is about
- There is transcriptional control between converting DNA to pre – mRNA
- There is processing control between converting primary transcript to mRNA
In the cytoplasm
- mRNA moves out through the nuclear membrane pores this is controlled by transport control
- mRNA is then either degraded which happens by mRNA stability
- or turned into an active protein this happens by translation control
- the active protein is then converted into an inactive protein by protein activity control

Question 6

name the types of RNA

Answer 6

• mRNA
• rRNA
• tRNA
• others they are mainly regulatory

Question 7

what is the RNA type and function of RNA polymerase I

Answer 7

rRNA

- ribosomes - structural and catalytic subunits of ribosomes

Question 8

What is the RNA type and function of RNA polymerase II (has 12 subunits)

Answer 8

mRNA - messenger (proteins) - code for proteins
miRNA - regulate mRNA
snRNA- regulate splicing
snoRNA - guide to methylation on RNA
IncRNA - chromatin binding and protein scaffolding

Question 9

What is the RNA type and function of RNA polymerase III

Answer 9

tRNA - Amino acid carriers in translation
miRNA - regulate mRNA
snRNA - regulate splicing

Question 10

Name and briefly explain the steps in mRNA synthesis

Answer 10

1. initiation – the polymerase binds to gene
2. elongation – polymerase transcribes the gene
3. termination – the polymerase stops transcribing the gene
4. processing – the mature mRNA is formed by splicing
5. export – the mRNA leaves the nucleus to be translated

Question 11

describe pre-initiation and initiation

Answer 11

1. exon 1 contains the codon ATG which is an start codon and this is what the RNA polymerase II binds to
2. in the upstream of the coding region of a gene is a region of DNA called a proximal promoter region
3. promoter has a pre- initiation complex formed on it which attracts RNA Polymerase II to the gene
4. TATA Box is the code which enables RNA polymerase II to bind to the coding region of the gene, (repeat of TA bases over and over again)
5. transcription factors are proteins that bind to a specific sequence of DNA and either recruit or block the binding of RNA polymerase
6. the polymerase, when bound, can then transcribe the gene – it does the stages of initiation, elongation and termination

• Distal regulatory elements – regions to which transcription factors bind

Question 12

describe transcription factors in initiation

Answer 12

Controlling rate – operator regions

• DNA proteins act as monomers and bind to enhancers and silencers
• Enhancers – bind activator proteins to increase the rate of transcription
• Silencers bind repressors to decrease the rate of transcription by preventing activator binding
• The rate of assembly of the preinitiation complex determines the rate of transcription of the gene
• Transcription factors interact with components of the basal machinery either directly or more generally via bridging proteins called co-factors

Question 13

Describe elongation

Answer 13

Elongation
1. RNA pol II adds RNA bases from 5’ to 3’
- Some factors have helical processes this allows the DNA to be unwound at the front and rewound at the back and therefore forms a transcriptional bubble
- As polymerase adds RNA bases in a 5’ to 3’ direction
- RNA polymerase stops transcription once it reaches termination signal
- There is a coding strand and a non coding strand, RNA polymerases reads of the non coding strand, by base pairing with that it creates mRNA that is identicial to the coding strand
the antisense strand of DNA is read by RNA polymerase from the 3 prime end to the 5 prime end during transcription, and the complementary RNA is created in the opposite direction in the 5 prime to 3 prime direction matching the sense strand

Question 14

describe termination and processing

Answer 14

• Basically RNA polymerase stops transcription once it reaches the stop codon
• AAUAAA is recognition sites for two factors called CPSF and CstF which are associated with RNA polymerase, CstF also binds to GUU region
• When MRNA goes through the factors and binds, this attract an endnuclease that cuts the growing mRNA
• other associated proteins such as PAP, synthesis 200 adenine molecules that get added to the end of it, this is required for mRNA stability – 3’poly-A-tail
• X nucleases attack from the end of the mRNA molecules
• Then the polymerase doesn’t stop and carries on, end is attacked and degrades the mRNA that is not used, this moves faster then the polymerase synthesises, and knocks the polymerase of the template

Question 15

what makes up the process of processing

Answer 15

• RNA capping
• 3’ poly-A-tail
• processing splicing

Question 16

describe the process of RNA capping and the function of it

Answer 16

At start of transcription there is a modification of the 5 prime end this is to stop it being degraded and subject to attack from endonucleases.
To stop this a cap is added to the 5 prime end this is 7-methylguanosine
This is attached via the 5’-5’ triphosphate bridge
In order to attach the 7-methylguanosine the 5 prime end is dephosphorylated and then guanosine transferase adds the modified guanine
Functions of this
- Prevents degradation
- Assists splicing
- Assists mRNA export out the nucleus
- Assists mRNA binding to the ribosome

Question 17

describe the process of 3 poly A tail and the function of it

Answer 17

3’ poly-A-tail
- Polyadenylation of the 3’end ( this means that lots of As are added)
- cleavage factors bind to polyadenylation signal and GU-rich sequence
- PAP attaches the As to the 3’end of mRNA
- PolyA binding protein binds PABP to the tail to accelerate the process
Function
- This prevents degradation
- Assists mRNA export
- Assists RNA translation initiation

Question 18

describe the processing of splicing

Answer 18

• Introns need to be removed
• 5’ splice site – GU
• 3’ splice site – AG
• Spliced by a protein RNA complex called snRNPs = small nuclear ribonucleoprotein polypeptide n
• 5 snRNP complex (U1, U2, U4, U5, U6) cleaves GU and AG (which are the splice sites) this connects the exons and forms an intron ring via GpA bond

Question 19

describe Nuclear export of mRNA

Answer 19

• Cap-binding complex binds to 5’ cap of mRNA an recognises if the mRNA has been correctly capped
• NXF1 binds to mRNA – 5’Cap, PolyA tail, and PABP are necessary
• mRNA is recognised by the nuclear pore complex(proteins in membrane) – mRNA export occurs

Question 20

what happens when general transcription and chromatin regulators go wrong

Answer 20

1. Mediator (a co-activator) in fibroids and prostate cancer, as well as developmental disorders
2. Loss of function of nucleosome modellers in a number of cancers

Question 21

what happens when transcription goes wrong

Answer 21

1. cMyc-most frequently amplified oncogene
2. AIRE-gene defects lead to autoimmune disorders
3. HNF1a/b, HNF4a- maturity onset diabetes of the young
4. Nuclear receptor e.g. VitD receptor, thyroid receptor, PPARy-endocrin problems
5. Huge number of transcription factors are important for development

Question 22

what happens when splicing is misrelated

Answer 22

• Mutated splice sites: cancer(BRAC1 and 2) spinal muscular atrophy (SMN2), Frasier syndrome(WT1) atypical cystic fibrosis(CFTR)
• Mutated splicing machinery: retinitis pigmentosa (spliceosome components), spinal muscular atrophy(snRNP assembly), myotonic dystrophy and glioblastoma (alternative splicing factors)

Question 23

what is the proteome

Answer 23

the entire complement of proteins that is or can be expressed by a cell tissue or organism

Question 24

the proteome of a cell defines…

Answer 24

its biological activity

Question 25

the information to code for the proteome is stored …

Answer 25

in the DNA

Question 26

what is the difference between RNA and DNA

Answer 26

• DNA is more stable than RNA
• RNA has uracil instead of thymine
• ribose has one more oxygen than deoxyribose

Question 27

what is the antisense strand

Answer 27

this is the template strand that is transcribed (non coding)

Question 28

what is the sense strand

Answer 28

this is the strand that is identical to the mRNA and is not transcribed (coding)

Question 29

what are the antibiotics in RNA transcription

Answer 29

rifampicin
rigamycin
streptolydigin
- these target RNA polymerase