lecture 1,2,3

Question 1

why cells are so different?

Answer 1

although most cells in an organism contain the entire genome, only a small proportion will be expressed and this will govern the function of that cell

Question 2

central dogma of molecular biology

Answer 2

DNA (gene) makes RNA makes PROTEIN, transcription then translation;
upstream 5’, downstream 3’

Question 3

where RNA is made

Answer 3

nucleus; transcription

Question 4

steps of eukaryotic transcription

Answer 4

1. unwind dna and expose the basis on each strand (usually H bonded)
2. one of dna strands acts as template
3. transcription initiation complex
4. addition of nucleotides on rna
5. dna helix reforms behind

Question 5

transcriptions initiation complex contains (3 parts)

Answer 5

rna pol 2 (enzyme, forms phosphodiester bonds and adds ribonucleotide) bound to a promoter (short dna sequence) and transcription factors (proteins)

Question 6

rna pol 1, 2, 3 functions

size/weight

Answer 6

pol 1: synthesises rRNA
pol2: synthesises mRNA
pol3: synthesises tRNA, 5S rRNA, small nuclear RNAs
8-12 subunits, 500 kDa

Question 7

template strand

Answer 7

3’ to 5’ that what rna polymerase uses

Question 8

rna pol functions

Answer 8

unwinds and copies dna

more that 1 rna can be made from 1 dna

Question 9

mRNA, rRNA, tRNA functions

Answer 9

mRNA (3%), code for proteins
rRNA(71%), forms the basic structure of the ribosome and catalyses protein synthesis
tRNA, adaptor between mRNA and aa (used in translation)

Question 10

how does the cell know where on the dna to form the transcription initiation complex?

Answer 10

promoters and GTF

Question 11

promoters, general characteristics and example

Answer 11

-short sequence found upstream of a coding region
-rna pol binds to dna
-like a little flag saying “im a gene, come and form initiation complex on me”
-TATA box (25-35 bp form the start site of transcription)
-people believed it was ridiculous that sequence was too short; it always starts with T (83%)
WHERE rna pol will bind

Question 12

GTF

Answer 12

rna pol cannot directly bind to dna, GTF are needed
-transcribed by pol3, multimeric and highly conserved
HOW rna pol will bind
-gtf bind rna pol 2 at promoter site, resulting in TRANSCRIPTION INITIATION COMPLEX (TIC)

Question 13

TIC

Answer 13

• GTF (proteins) + promoter (TATA box, short dna sequence) = TIC

Question 14

assembly of TIC and start of transcription

Answer 14

1. TBP (TATA-binding protein) binds to tata box in dna and bends the dna
2. GTF: TF2D, TF2B, recruitment of rna pol 2+TF2F+CTD tail, TF2E, TF2H
3. TF2H contains kinase and helicase activity-leads to phosphorylation of CTD
4. rna pol 2 leaves the promoter and starts elongation of nascent dna

Question 15

pre-mRNA/hpRNA

processing steps

Answer 15

gene copied with introns (non coding) and exons (coding) into rna
steps:
-5' capping 
-3' polyadenylation
-splicing
end: mature mRNA

Question 16

advantages of polyadenylation

Answer 16

• stabilizing rna
• longer polyA tails=longer half life
• nuclear export
• translation

Question 17

what is cDNA

Answer 17

copy of mRNA

Question 18

elements for polyA (many A)

Answer 18

• polyA signal AAUAAA
• polyA site (rna will be cleaved)
• G/U region

Question 19

factors for polyA

Answer 19

• CPSF with CStF, CF1 CF2 by bending pre-mRNA
• PAP (poly A polymerase), cleavage (split) 10-35 upstream of polyA signal causing slow polyadenylation (3’ polyadenylation); ENERGY DEPENDENT PROCESS (atp to ppi)
• after, CPSF CstF CF1 CF2 factors leave, and PAB2 binds. This increased the addition rate of A by PAP (rapid polyadenylation)
• 200-250 A: PAB2 signals PAP to stop

Question 20

problems with gene expression lead to diseases

Answer 20

• cancer: absence of transcription off switch
• development disease: problems in expression of early proteins; like, kidneys when cannot be produced at all
• chronic disease: like, obesity; can cause inflammation and increase the risk of other diseases

Question 21

where gene expression is regulated the most?

Answer 21

transcription initiation

Question 22

two types of genes

Answer 22

constitutive (house-keeping): genes that most cells will need
highly regulated: selectively expressed under specific conditions (eg. cyclins-cell division) or in specific tissues

Question 23

U1A

Answer 23

• nuclear protein
• involved in splicing of rna
• U1A binds its own mRNA, leading to inhibition of the polyadenylation meaning that pre-mRNA never matures