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B3 Biochem > Transciption > Flashcards

Transciption Flashcards

1
Q

Action of RNA Polymerase

How does it begin?
How does it read the template?
How does it synthesize the transcript?
What does it recognize?

A

Begins transcription WITHOUT a primer

Reads the template strand from 3’ to 5’

Synthesizes RNA transcript 5’ to 3’ using ribonucleotides

Recognizes transcription termination signals

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2
Q

What is an NTP?

A

Ribonucleotide

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3
Q

Transcription begins with..

A

The RNA polymerase identifies a genes’ PROMOTER region

Interacts with the DNA template strand

Initiates synthesis of a complementary , anti parallel RNA transcript

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4
Q

Transcription ends when…

A

A termination signal is reached

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5
Q

What is described by the sequence of the coding strand?

A

Genes.

However, the template strand is actually the strand of DNA read during synthesis.

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6
Q

The DNA coding sequence is identical to:

A

The RNA transcript sequence, instead of T’s it uses U’s

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7
Q

How are base sequences labeled (downstream to the right)?

And upstream DNA bases (to the left)?

A

+1, being the first base transcribed, then +2 +3, etc in the 3’ direction

Upstream -1, -2, -3, etc in the 5’ direction

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8
Q

Using what method, how do you determine the other two strands being transcribed/translated?

A

Watson-Crick base pairing

ATGGGG -> TACCCC -> AUGGGG -> Met Gly

DNA -> transcribed strand -> translated strand -> protein

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9
Q

What type of molecule is a Prokaryotic RNA polymerase?

A

Holoenzyme

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10
Q

What makes up the holoenzyme?

A

Core enzyme + sigma factor

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11
Q

What makes up the core enzyme of the holoenzyme?

A

4 subunits.

2 alpha subunits
1 beta subunit
1 beta-prime subunit

α2ββ’

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12
Q

The importance of the core enzyme and sigma factor parts of the holozenzyme/RNA polymerase

A

Core enzyme- has RNA polymerase activity

Sigma factor- is required for recognizing and binding promoter sequences

Looks like an upside down chef’s hat. The base is the sigma factor, the top is the core enzyme

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13
Q

Role of the sigma factor (of the holoenzyme)

A

Different sigma factors recognize different types of promoter sequences

They recruit the core enzyme to the DNA promoter (since they’re attached)

Dissociates from the core enzyme after transcription begins

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14
Q

Initiation of transcription

A

Scanning and recognition of promoter sequence cues by the holoenzyme trigger transcription initiation.

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15
Q

What do the promoters contain?

A

Contains two consensus sequences that are recognized by sigma factor.

  1. TATA box
  2. -35 sequence
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16
Q

TATA box

A

Part of the promoter region

Consensus sequence of 6 nucleotides

TATAAT

Aka Pribnow box, ~7 nucleotides upstream from the +1 transcriptional start site (~-7)

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17
Q

Another name for the TATA box

A

Pribnow box

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18
Q

What is the -35 sequence?

A

Part of the promoter region

Consensus sequence of 6 nucleotides

TTGACA

~35 bases upstream from the +1 transcriptional start site

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19
Q

What is elongation?

A

When transcription begins at the +1 transcriptional start site.

Sigma factor is released and the core enzyme continues

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20
Q

The elongation process contains the core enzyme which does what?

A

Creates a temporary “melt” between the double stranded DNA

-forming a transcription “bubble”

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21
Q

What can supercoils of the DNA strand be relieved by?

A

The action of topoisomerases

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22
Q

Net reaction of Prokaryotic RNA Synthesis

A

Addition of a ribonucleotide to the growing RNA chain, and the release of pyrophosphate

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23
Q

Pyrophosphate is further cleaved by:

A

Pyrophosphatase.

Irreversible coupled reaction.

2 high energy bonds are cleaved.

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24
Q

What are the 2 termination mechanisms of Prokaryotic RNA synthesis?

A

Rho-dependent termination requires an additional protein.
(Rho factor: displaces the DNA template strand from RNA polymerase)

Rho-independent termination requires a G-C rich stem loop followed by a poly-U stretch
(Causes RNA polymerase to dissociate)

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25
Q

Prokaryotic mRNAs can be polycistronic..what does that mean?

A

One mRNA can code for several proteins

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26
Q

Polycistronic mRNAs often do what?

A

Code for multiple enzymes in the same biosynthetic pathway

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27
Q

Eukaryotic mRNAs are always polycistronic.

T/F

A

FALSE

They’re always MONOcistronic

28
Q

What is rifampin?

A

Aka rifamycin.

An antibiotic

Binds to prokaryotic RNA polymerase (changes its confirmation) and prevents transcription initiation/elongation

29
Q

What is rifampin/rifamycin used to treat?

A

TB

30
Q

Prokaryotic synthesis of tRNA and rRNA, how is it different from eukaryotic RNA synthesis?

A

Similar to model for Eukaryotes, except in prokaryotes the same RNA polymerase produces all types of RNA

31
Q

Eukaryotic DNA is highly condensed into __________.

But for gene transcription, the genes must be exposed.

This “exposure” occurs in the ____________. Why?

A

Chromatin.

Euchromatin, less condensed area, more accessible to RNA polymerases.

32
Q

What causes the chromatin to revert to nucleosomes?

A

Acetylation of histone H1

33
Q

Heterochromatin vs Euchromatin

A

Euchromatin- less condensed

Heterochromatin - appears more dense, DNA is relatively inaccessible due to highly condensed structure

34
Q

What is special about DNA in heterochromatin?

A

Its highly methylated

35
Q

DNA methylation characteristics is the foundation of..

A

Epigenetics

36
Q

Eukaryotic RNA polymerases

A

I, II, and III

37
Q

Transcribes precursors to rRNA in the nucleolus

A

RNA polymerase I

38
Q

Transcribes the precursor to mRNA (hnRNA) in the nucleoplasm (some snRNAs as well)

A

RNA polymerase II

39
Q

Transcribes tRNA precursors (also small rRNA (5S) and some snRNAs)

A

RNA polymerase III

40
Q

Transcription initiation in eukaryotes begins with what RNA polymerase?

A

RNA polymerase II

41
Q

RNA polymerase II recognizes two consensus sequences which are…

For transcription initiation in eukaryotes

A

TATA box (Hogness box)

  • similar to TATA box in prokaryotes
  • approximately 25 nucleotides upstream (-25)

CAAT box
-approximately 70 nucleotides upstream (-70)

42
Q

What is different about elongation and termination in Eukaryotic RNA synthesis compared to prokaryotes?

A

Elongation is essentially the same.

Termination in eukaryotes requires a signal…that isn’t exactly understood. Hmm.

43
Q

rRNA Synthesis and Processing

A

3 rRNAs are transcribed as a single large precursor (by rRNA pol I in the nucleolus)

Individual rRNAs cleaved apart by RNases

Prokaryotic rRNA synthesis is the same as in eukaryotes

44
Q

What transcribes the three rRNAs from a ribosomal RNA gene?

What cleaves the 3 RNAs from a chain to individual rRNAs?

A

RNA polymerase I

RNases

45
Q

Where is RNA pol III?

A

In the nucleoplasm

46
Q

What is RNA pol III responsible for?

A

In tRNA synthesis and processing:

Removal of intron loops.

Trimming the 5’ and 3’ ends

Base modifications

Addition of the 3’ -CCA sequence

47
Q

Where does eukaryotic mRNA synthesis occur?

Transcription and translation

A

Transcription - in the nucleus

Translation - in the cytoplasm

48
Q

Eukaryotic mRNAs vs Prokaryotic mRNAs

A

Euk mRNAs are extensively processed.
-hnRNA to mRNA

Pro mRNAs begin translation before transcription finished.

49
Q

What are the MAJOR differences between Pro/Euk RNA synthesis?

A

Pro- promoter regions, 5’ and 3’ UTRs, coding region

Euk- 3’ terminus (and 5’ terminus), exons and introns are present

50
Q

What protects eukaryotic mRNA from nuclease degradation?

A

5’ capping addition of 7-methylguanosine to the 5’ end of the mRNA

5’ to 5’ linkage

51
Q

What is the 3’-poly-A tail on euk mRNA?

What does it do? What does it serve as? Is it encoded in the gene?

A

The 3’ terminus contains a polyadenylation signal - Poly-A polymerase, a special enzyme that adds a variable stretch of adenine residues (~40-200)

Serves as a signal for transport out of the nucleus, stabilizes the mRNA from nuclease attack.

Not encoded in the gene. The SIGNAL sequence is encoded, but not the adenine additions.

52
Q

Intervening sequences

A

Introns

53
Q

Expressed sequences

A

Exons

54
Q

What is the fate of introns?

A

Must be removed from the coding region for proper translation

55
Q

The removal of introns and the joining of exons by snRNPs

A

Splicing

56
Q

Composed of snRNAs and a group of proteins

A

snRNPs

57
Q

snRNPs combined with the primary transcript

The molecular machine that performs the splicing reaction

A

Spliceosome

58
Q

What are splice sites?

A

Have very specific sequences, mutations at these sites can cause incorrect splicing

59
Q

Exons are joined together during splicing to form..

A

Mature mRNA

60
Q

What is a lariat?

A

The excised intron

61
Q

Eukaryotes can contain how many intron sequences?

A

0-50

62
Q

Certain hnRNAs may undergo alternative splicing. What is that? What’s the point?

A

Joining of different exons together to form different mRNAs

It permits the production of proteins with some common domains, but overall the function is different.

63
Q

Systemic Lupus Erythematosus

What is it?
Who does it usually affect?
Why is it a concern?

A

Autoimmune disease.

Late-teen females

Antibodies are produced that recognize host proteins, including snRNPs.
Other pts can produce antibodies that recognize nucleic acids (anti-dsDNA)
**anti-nuclear antibody panel is performed to counter this

64
Q

Eukaryotic mRNA Processing summary

A
  • processing events
  • 5’ capping
  • 3’ poly-adenylation
  • splicing
  • after all processing events are complete, mature mRNA is transported to the cytoplasm
65
Q

Mushroom Poisoning

What type of mushroom? What does it do?

A

Amanita phalloides, the death cap mushroom. OOOOoooOOoo

Alpha-amantin binds to RNA polymerase II and inhibits mRNA synthesis.

Accounts for 95% of all mushroom fatalities

B3 Biochem (4 decks)

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