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Flow of genetic information BIOL2010 > Transcription > Flashcards

Transcription Flashcards

1
Q

Where is DNA found in eykaryotes and how does it send messages to create proteins?

A

found in the nucleus and uses mRNA

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2
Q

Where does transcription occur in prokaryotes?

A

In the cytoplasm.

Replication, transcription, and translation all occur in the cytoplasm for prokaryotes.

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3
Q

Why is the intermediate mRNA needed for the transcription of proteins?

A

better regulation of the protein production process.

Prevents interference and potential additional mistakes.

proteins can still be made when DNA is being replicated.

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4
Q

What is RNA?

A

Single stranded linear polymer like DNA but has a ribose sugar. has the bases A,G,C and U.

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5
Q

What types of RNA are there?

A

rRNA: ribosomal
tRNA: transfer RNA which acts as the adaptor between mRNA and amino acids
non coding RNA: e.g microRNA, long non coding RNA.
mRNA: messenger RNA which codes for 3-5% of proteins

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6
Q

What is the most common form of control of protein production?

A

Regulation of Transcription. (pre-mRNA) need for things such as splicing of introns etc.

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7
Q

What is the structure and function of RNA polymerase?

A

Has a crab claw structure. seperates 2 strands of DNA and uses one strand as the template for RNA synthesis

does not require a primer and transcribes in the 5’ to 3’ direction.

synthesises a complementary RNA copy of the template DNA.

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8
Q

What are the 3 main processes in transcription?

A

Inititation, Elongation, Termination

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9
Q

Many RNA copies are made at the same time? True/False

A

True! multiple RNA polymerases per gene

multuple transcripts per gene. as soon as the first RNA pol starts to move down the gene the next POL can bind and initiate RNA transcription

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10
Q

How big is the RNA transcription bubble?

A

Length of the bubble is around 12-14 base pairs and has a reaction rate of approx 40bp per second.

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11
Q

What is the step of initiation?

A

polymerase locates the promoter,
polymerase unwinds the DNA
Transcription begins

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12
Q

what is elongation?

A

POL places the RNA in the exit hole

once RNA is 10 or more bases long there is a conformational change in the RNA POL that closes binding to itself securing it to the DNA.

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13
Q

What is the step of termination?

A

once the termination sequence is reached the POL seperates and the RNA is released.

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14
Q

WHat type of genes regulate transcriptional control in bacteria?

A

-Consstutitive genes (hosuekeeping genes) the are transcribed the whole time and regulate everyday function of the bacterial cell.

  • regulated genes which turn on and are transcribed when there is certain conditions
    e. g. change in food sources where a gene encoding for an enzyme might be needed to metabolise a sugar

or change in enviromental stresses such as pH and temp where genes are turned on to help survive.

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15
Q

What is an Operon?

A

Genes encoding for proteins in the same pathway that are located adjacent to one another and are controlled as a single unit that is transcribed into a polycistoronic RNA (no introns)

e.g Lac Operon

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16
Q

What does the lac operon control?

A

changes in lactose which switches genes on that encode for enzymes which are needed to metabolise that sugar.

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17
Q

what is the lac operon made of?

A

LacZ, LacY, LacA and then lac terminator

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18
Q

What do the genes in the lack operon encode?

A

LacZ encodes for beta Galactosidase
LacY encodes for Permease
LacA encodes for Transacetylase

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19
Q

What are bacterial promotors needed for?

A

Control of transcription most commonly occurs at initaiton. Promotors are the site of initiation.

The promotors are recognised by RNA POL by having a consensus (common pattern) DNA sequence.

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20
Q

What is the consensus DNA sequence of promotors?

A

hexamer (6bp) at -35 upstream of start point of transcription.
TATAAT (Pribnow box) at -10 upstream of start point of transcription.

Only found on one strand so RNA POL knows the direction of transcription.

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21
Q

What type of mutations can decrease or increase promotor effectiveness?

A

Down mutations decrease by reducing conformation to the consensus sequence.

up mutations have the opposite effect.

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22
Q

What is the bacterial RNA POL made of?

A

it is a holoenzyme that consists of 4 types of subunits (5 units total)

2 alpha
1 beta
1 beta prime
1 sigma

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23
Q

What is the function of the alpha subunits in baterial RNA POL

A

needed for enzyme assembly, promotor region binds some activators

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24
Q

What is the core enzyme of RNA POL

A

the two alpha, beta and beta prime. the core enzyme has a general affinity for DNA. this is known as loose binding.

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25
Q

What is the sigma subunit in the RNA POL for?

A

ensures RNA polymerase only binds at promotor sequences.

1000x binding strength with it and overall there is only enough sigma subunits for 1/3 of all the RNA POL.

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26
Q

What are some of the alternative sigma factors used for?

A

sigma70 is for general use.
sigma32 is for high temperatures.
sigma54 is for nitrogen

when conditions change there is induction of these alternative sigma factors that recognise different -35 and -10 consensus sequences.

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27
Q

What type of proteins control transcripton?

A

Regulatory proteins called transcription factors.

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28
Q

Negative regulation of transcripton is controlled by what?

A

Transcriptional repressors. They bind to sites known as operator sites that stop RNA polymerase binding.

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29
Q

Positive regualtion of transcription is controlled by what?

A

Transcriptional activators. They bind to specific site that helps RNA polymerase bind.

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30
Q

What does beta galactosidase do?

A

cleaves lactose into its component sugars

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31
Q

what does permease do?

A

transport lactose into cells

32
Q

what does transacetylase do?

A

covalently modifies lactose

33
Q

What is the trancriptional activator in the Lac operon?

A

CAP. (catabolite activator protein)

34
Q

what is the lac Repressor?

A

a transcriptional repressor. is always on when lactose is absent. is produced by the gene LacI

35
Q

When lactose is present how can lac repressor be turned off?

A

when lactose is present, alloactose is produced that binds to the lac repressor that induces a conformational change in LacI so the repressor cannot bind.

36
Q

What is the structure of the lac repressor?

A

it is a tetramer that contains 2 dimers.
each dimer within the tetramer can bind to one operator site( operator sites overlap with the start site of transcription)

37
Q

How many operator sites does the lac operon have?

A

3.

38
Q

What operator sites must be bound to by lac repressor for repression to occur?

A

MUST BIND TO 2 out of 3 sites.

can bind to O1 + O2 OR O1 + O3

BUT NOT O2 + O3

this is due to the loops that can form between the repressor and operon that are needed to prevent the attachment of RNA polymerase.

39
Q

what occurs with the lac operon in enviroment of low glucose and high glucose?

A

low glucose > high levels of cAMP. this activates CAP promoting transcription of the lac Operon.

high glucose > low levels cAMP. this mean CAP remains inactive and transcription of lac Operon is repressed

40
Q

Glucose is the preffered sugar. true or false?

A

TRUE

41
Q

what is an intrinsic terminator?

A

A run of A-Ts in the template strand

42
Q

How does the stem loop strucutre in the RNA cause termination?

A

formed by inverted repeats. Causes RNA POL to pause and then unravel from the weakly bound A:U terminal region.

43
Q

What deterimines the efficency of a terminator?

A

the sequence of the hairpin and the length of the U rich region.

44
Q

genes that are transcribed and expressed by mRNA use what polymerase?

A

RNA polymerase II

45
Q

What is the difference between Eukaryotic and bacterial RNA POL II

A

eukaryotic RNA POL II has no sigma factor so cannot initiate transcription

46
Q

What is the role of transcrpitional activators?

A

they help attract RNA POL II to the promoter and help reguate the rate and tissue specificity of gene expression.

47
Q

how do the transcrpitonal activators recognise parts of DNA?

A

They look at the major and minor groves of the DNA. binding tends to occur in the MAJOR groove which allows for specicifity.

48
Q

What is the first transcription factor to bind?

A

TFIID. it is made up of TBP (TATA binding protein) is also associated with TAF’s (TBP associated factors) that are involved in binding other TF’s to help attach RNA POL to the DNA and attaching proteins to unwind the chromatin structure.

49
Q

What is the order of transcription factors binding?

A

First TFIID
second TFIIA
third TFIIB

once these three have bound then RNA POL II and TFIIF binds as a complex.

next TFIIE (ATPase) binds
followed by TFIIH (helicase)

this whole complex is known as the PIC (Pre initiation complex)

50
Q

what are enhancers?

A

they can be placed thousands of Base pairs away from the promotor but still enhance the rate of transcription.

51
Q

What are the 3 general structures of DNA binding domains?

A

Zinc fingers
Helix turn helix
Basic binding domains

52
Q

What is a zinc finger binding domain and its structure?

A

contains a loop of 23 amino acids and usually has multiple zinc fingers.
the linker between the fingers is 7 to 8 amino acids long.

the ion doesn’t not directly interact with the DNA but is essential for the folding of the finger

zinc fingers bind both the major and minor grooves

53
Q

what is an example of a zing finger containing transcription factor?

A

steroid hormone receptors ( cys2-cys2 fingers)

54
Q

what is a helix turn helix? (homeodomain)

A

they are 60aa and contain 3 alpha helices.

the c terminal helix number 3 is 17aa and lies in the major groove.

helice 1 and 2 point away from the DNA.

55
Q

what are the basic binding domains?

A

transcription factors with basic bidning domains cannot bind to DNA alone. They must dimerise (become a dimer) to become specific.

example of this is a leucine zipper.

56
Q

what do transcription factors do to modify histones?

A

recruit co-activators such as histone acetylates (HAT) that acetylates the N terminal tail lysine of the histone units which neutralises their +ve charge.

opens up the DNA allowing TF and RNA POL to gain acess to the DNA. (loosens DNA histone complex)

57
Q

how do TF inhibitory domains work?

A

bind to DNA and block TF with activator domains from binding.

they bind to the PIC and block transcription with its inhibitory domain.

co repressors can also be recruited which interact with the PIC and tighten the chromatin structure.

58
Q

how do Co repressors alter the chromatin structure?

A

they modify the histones. Histone de-acetylase (HDAT) removes acetyl groups of histone units and restores their +ve charge.

closes down the DNA which shuts off transcription.

59
Q

In elongation of the RNA what is required to happen to the PIE?

A

TFIIH phosphorylates of the c terminal domain of RNA POL. causes a conformational change where the grip tightens and the general TF dissociate. leads to the acquisition of new proteins including elongation factors.

60
Q

What is the C terminal domain in RNA POL made of?

A

is 52 tandem repeats of 7 amino acids. each contain 2 serines (Ser2 and Ser5)

the phosphoryation of it allows for the loading of RNA processing machinery. like modification enzymes, elongation factors, splicing proteins etc

61
Q

What are some of the post transcriptional processing modifications that can occur?

A
  1. addition of a 5’ cap to protect and mark the mRNA
  2. 3’ processing and polyadenylation which marks the end of the mRNA and protects it.
  3. splicing to remove non coding regions
  4. editing which is similar to mutating the mRNA to produce an alternative final product but is not random
  5. transport to cytoplasm for translation.
62
Q

What are the steps to RNA splicing?

A

step 1. cleavage at the 5’ splice site. OH of the A nucleotide within the branch point acts as a nucleophile and attacks the 5’ splice site. at the same time the intron folds back on itsel and forms a phosphodiester bond between A at branch point and G at the splice site.

step 2. cleavage at the 3’ splice site and then the joining of the exons. the intron gets released in the form called a lariat and degraded in the cell.

63
Q

What is a spliceosome?

A

complexes made up of RNA and protein. the RNA component is made of small nuclear RNA (snRNA)

each RNA is complexed with 6-10 proteins to form small nuclear ribonuclear protiens (snRNPs)

different complexes come in at different stages

use ATP.

64
Q

in what order does the spliceosome form?

A

first U1 snRNP binds to the 5’ splice site.
then BBP (branch point binding protein) binds to the branch point.
U2AF35 binds to the 3’ splice site
U2AF65 to the polypyrimidine tract.

next U2 snRNP binds to the branch site which is aided by the U2AF proteins and displaces BBP.

next the U4, 5 and 6 snRNPs bind.
the U2AF proteins are displaces and this brings the 5’ splice site close to the branch point and 3’ splice site.

65
Q

What are SR proteins?

A

proteins rich in serine and arginine. they bind to sequences called exonic splicing enhancers (ESE’s)

SR proteins recruit the splicing machinery to the nearby splicing sites.

66
Q

What are the 5’ capping enzymes

A

RNA triphosphatase
Guanylyltransferase
Methyltransferase

67
Q

What are the steps involved in the addition of the 5’ cap.

A

there is the addition of the 7 methyl-guanosine to the 5’ end of the RNA that is done in 2 steps.
first with guanylyltransferase and the RNA triphosphaste that removes one phosphate from pppRNA, converts GTP to GMP and then links them together to make GpppRNA.

the next step is the addition of the methyl group at the 7th position of the guanine base by methyltransferase. to make 7MeGpppRNA

68
Q

What is the purpose of the 5’ cap?

A

helps distingush mRNA from other RNA
helps mRNA be properly processed and exported from the nucleus
Protects it from degregation in the cell.

69
Q

What does viral mRNA do in order to be translated by ribsosomes?

A

Viral cap snatching. steals the 5’ cap of the host cell.

70
Q

What happens to the 3’ end?

A

the 3’ end is polyadenylated which involves the addition of a string of adenosines ( ~200) to the end of the transcript. the signal for polyadenylation is encoded for in the DNA ( conserved 3’ control region)

proteins bind to the poly A tail. (PolyA binding protiens (PAB)) and help stabaise the RNA.

71
Q

What set of proteins recoginse the seqence at the 3’ end for polyadenylation?

A

first the end has to be recognised by a set of proteins.
CPSF = clevage and polyadenylation specicfity factor
PolyA pol = Pol A polymerase
cstF = Clevage stimulation factor
CFI and CFII = clevage factors I and II.

72
Q

How is RNA transported out of the nucleus?

A

via nuclear pores. Are brought to the pores by REF proteins found at exon splicing junctions. where they associate with other proteins TAP and MEX that allow them to be transported to the nuclear pore and dock.

73
Q

what is micro mRNA and how is is synthesised?

A

first pre Micro mRNA is produced in the nucleus by the protein DROSHA. then the pre micro mRNA is synthesised to micro mRNA in the cytoplasm by DICER.

74
Q

what is the role of micro mRNA?

A

used to make an RNA induced Silencing complex to prevent the translation of the mRNA into proteins. the micro mRNA tend to be around 22 bp long and bind to the mRNA to form an RNA duplex that prevents translation. q

75
Q

what protein helps micro mRNA bind to the correct gene mRNA to silence?

A

argonaut protein. (Ago) helps line up the micro mRNA to the genes mRNA and leads to translational repression or mRNA degradation.

Flow of genetic information BIOL2010 (4 decks)

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