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BIOL 0280 UNIT 4 > Lecture 20: Transcription & RNA Metabolism > Flashcards

Lecture 20: Transcription & RNA Metabolism Flashcards

1
Q

Intro Facts

Where does Transcription and Translation occur in prokaryotic cells?

Eukaryotic cells?

A

Prokaryotic - everything occurs in the cytoplasm

Eukaryotic
- Transcription & Processing/Transportation
= Nucleus

  • Translation = in the Cytoplasm
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2
Q

Intro Facts

What is transcription?

What is the purpose of transcription?

A
  • Transcription is the process of copying a segment of DNA into RNA
  • The purpose of transcription is to make RNA copies of genes
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3
Q

Intro Facts

How many DNA serve as a template in transcription?

A

Only one DNA will serve as a template

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4
Q

Intro facts

What is the template strand?

What is the non template/coding strand?

A

Template: The strand that is complimentary to RNA transcript

Non template: the same sequence as RNA (with T instead of U)

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5
Q

Transcription vs. DNA replication

How much of the genome is copied in Transcription vs. DNA replication?

A

Transcription: One/particular group of genes

DNA replication: whole genome

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6
Q

Transcription vs. DNA replication

What is the template used in Transcription vs. DNA replication?

A

Transcription: only one strand serves as the template

DNA replication: both strands will serve as template

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7
Q

Transcription vs. DNA replication

Which processes use primers?

A

Transcription does not use primers

DNA replication uses primers

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8
Q

Transcription Basics

Are polymers built continuously?

A

Yes, polymers are built continuously (processive)

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9
Q

Transcription Basics

In what direction is RNA synthesized?

Describe the relationship between the template and RNA strand

A

5’ -> 3’

  • They are anti-parallell
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10
Q

Transcription Basics

What is RNA Polymerase?

A

Enzymes that catalyzes the synthesis of RNA by reading the template strand

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11
Q

Transcription Basics

Does RNA polymerase require a primer?

What does RNA polymerase use?

A
  • Does not require a primer
  • Uses ribonucleotides
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12
Q

Essential Components of a Gene

What are the 4 essential components of a gene?

A

1) Promoter
2) Transcription factors
3) RNA Polymerase
4) Terminator

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13
Q

Essential Components of a Gene

What is the promotor region?

A

Sequence of DNA where transcription proteins assemble

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14
Q

Essential Components of a Gene

What specific machinery do we need at the promoter?

A
  • Transcription factors
  • RNA Polymerase
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15
Q

Essential Components of a Gene

What do transcription factors do?

A

They are proteins that bind to promoters to direct RNA synthesis

MAIN PURPOSE: Increase affinity of RNA polymerase at the promoter region

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16
Q

Essential Components of a Gene

What is the terminator region?

A

Sequence of DNA where transcription ends

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17
Q

Transcription Phases

What are the 4 phases of transcription?

What is its nemonic?

A

1) Assemble
2) Initiation
3) Elongation
4) Termination

Aliens Invade Earth Tonight

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18
Q

Transcription Phases

Which phases of transcription are very different in prokaryotes vs. eukaryotes?

A

Initiation & termination

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19
Q

Transcription Phases

What are transcription factors typically referred to as in prokaryotes?

A

Sigma factors

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20
Q

Transcription Phases

What are sigma factors?

A

Proteins that bind to specific DNA sequences in the promoter and increase affinity for RNA polymerase

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21
Q

Transcription Phases

What are consensus sequences?

A

DNA sequences that transcription factors and polymerases repeatedly assemble at

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22
Q

Transcription Phases

Where do transcription factors, sigma factors, and polymerases assemble?

A

They assemble at the promoter

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23
Q

Transcription Phases

What are the steps of initiation?

A

1) Initiation occurs at the promotor sequence

2) Transcription factors, sigma 70 & RNAPs assemble at the promoter region and forms a closed complex

3) Transcription bubble forms and RNAP forms open complex

4) Transcription is initiated

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24
Q

Transcription Phases

What are the main things that occur during elongation?

What are the steps of elongation?

A

Main Occurrences
- Clearing the promoter
- RNA polymerase builds the RNA strand by complementing the template strand

Steps
1) Sigma 70 disassociates
2) Sigma 70 is replaced by NusA
3) Transcription terminates and NusA disassociates

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25
Q

Transcription Phases

Super coils form during the elongation process

In what direction do they form?
What kind form?

A

Positive supercoils in the direction of transcription

Negative supercoils in the opposite direction of transcription

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26
Q

Transcription Phases

What is the cost of putting in a new nucleotide during elongation?

A

2 ATP equivalents

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27
Q

Transcription Phases

What is the main thing that occurs during termination?

A

RNA polymerase dissociates from DNA strand, releasing newly synthesized RNA polymer

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28
Q

Transcription Phases

What are the two different termination methods that occur in prokaryotes

A
  • ρ-independent
  • ρ-dependent
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29
Q

Transcription Phases

Describe the difference between ρ-independent and ρ-dependent

A

ρ-independent
- relies on a repeating palindromic sequences at the termination sequence
- RNA transcribed from these sequences fold onto itself

ρ-dependent
- relies on ρ helicase, a hexamer protein that binds to the rut element and travels fowards, disassociating the elongation machinery

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30
Q

Eukaryotic vs. Prokaryotic Transcription

What are the 3 key differences between the two transcription methods?

A

1) There are many MORE transcription factors
called general transcription factors in
assembly, not one signal sigma factor
- However their job is still to increase affinity of promoter to RNA polymerase

2) C-terminal Domain (CTD) on RNA polymerase
is phosphorylated to start elongation

3) In the same vein, elongation is terminated by
dephosphorylation of RNA polymerase by
termination factors 12

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31
Q

Eukaryotic vs. Prokaryotic Transcription

What are the RNAPs present in Bacteria/Archea?

A

only one form of RNAP

32
Q

Eukaryotic vs. Prokaryotic Transcription

What are the RNAPs present in Eukaryotes?

List them

A

Multiple different types
- RNAP I
- RNAP II
- RNAP III
- RNAP IV/RNAP V

33
Q

Eukaryotic vs. Prokaryotic Transcription

What are the each specific RNA polymerase responsible for?

A

For producing different types of RNAs

34
Q

Eukaryotic vs. Prokaryotic Transcription

What RNA(s) does RNAP I produce?

A

rRNA

35
Q

Eukaryotic vs. Prokaryotic Transcription

What RNA(s) does RNAP II produce?

A

mRNA, snRNA, miRNA

36
Q

Eukaryotic vs. Prokaryotic Transcription

What RNA(s) does RNAP II produce?

A

mRNA, snRNA, miRNA

37
Q

Eukaryotic vs. Prokaryotic Transcription

What RNA(s) does RNAP III produce?

A

tRNA

38
Q

Eukaryotic vs. Prokaryotic Transcription

What RNA(s) does RNAP IV/V produce?

A

siRNA (plants)

39
Q

Eukaryotic vs. Prokaryotic Transcription

How many genes are transcribed in Prokaryotes?

A

polycistronic
- Transcribe multiple genes at once
- Genes are transcribed as operons

40
Q

Eukaryotic vs. Prokaryotic Transcription

How many genes are transcribed in Eukaryotes?

A

Monocistronic (ONE gene at a time)

41
Q

Eukaryotic vs. Prokaryotic Transcription

What is unique about RNA polymerase II?

A

Phosphorylation state of the C-terminal domain (CTD) changes with each stage of transcription

42
Q

Post Translational Processing

What are the 3 Steps to Process RNA?
When do these steps occur?

A

1) 5’ end is capped
2) Exon sequences are spliced together
3) Poly-adenylated tail (Poly-A) is added to the 3’ end

These steps occur alongside transcription, they are co-transcriptional

43
Q

Post Translational Processing

Why are Eukaryotic mRNAs capped?

A

1) To protect from ribonuclease
2) Enable binding to cap-binding proteins and ribosomes

44
Q

Post Translational Processing

What occurs during the capping of the 5’ end?

What are the results?

A
  • Add 7-methylguanosine to 5’ end
  • Enhanced stability
  • Roles in processing/translation
45
Q

Post Translational Processing

What do Poly (A) tails contribute to?

A

Roles in stability and translation

46
Q

Post Translational Processing

What are primary transcripts spliced into?

A

a continuous coding sequence

47
Q

Post Translational Processing

What are introns?

A

Sequences of RNA in between expressed regions of the transcript

48
Q

Post Translational Processing

What are exons?

A

Sequences of RNA transcript that code for protein

49
Q

Post Translational Processing

How do introns and exons vary in length?

A

Introns vary widely between 50-700,00 nucleotides
~1800 nucleotides on average

Exons are relatively short less than 1000 nucleotides
~100-200 on average

50
Q

Post Translational Processing

What are the 2 splicing mechanisms?

A

1) Self-Catalyzed splicing
2) Spliceosome Mediated Splicing

51
Q

Post Translational Processing

Describe self catalyzed splicing
- Where does it occur?
- What are the 4 steps?

A

Occurs in RNA but NOT DNA
1) Driven by the higher reactivity of RNA given by the 2’ OH

2) 2’ OH specific Adenosine performs a nucleophile attack of the 5’ splice site, forming lariat

3) Exposed 3’ OH of 5’ exon (the base at the 5’ splice site was “stolen”) now attacks the other side of the lariat

4) The lariat disassociated and the two ends of the exon are ligated together

52
Q

Post Translational Processing

What do Eukaryotes utilize on order to splice transcripts?

A

Spliceosome

53
Q

Post Translational Processing

What is a Spliceosome?

A

A large complex made up of small nuclear ribonucleo Proteins (snRNPs)

54
Q

Post Translational Processing

What do snRNPs contain?

A

specific small nuclear RNA to interact with specific sequences within pre-mRNA

55
Q

Post Translational Processing

What do snRNPs do?

A

Binds to intron and assemble to entire spliceosome to cut that region out

56
Q

Post Translational Processing

What do poly-A binding proteins regulate?

A

They regulate RNA export, breakdown, and aid in translation to protein

57
Q

Post Translational Processing

What 2 steps occur during the poly a tail addition?

A

1) Primary transcript is trimmed, leaving a consensus sequence at the 3’ end

2) Polyadenylate polymerase adds 30 (yeast) to 100 (animals) A residues

58
Q

Post Translational Processing

What is alternate splicing?

What does it result in?

A

the same “raw” transcript can result in two different mature transcripts

these two different transcripts translate into two different protein sequences with different downstream effects

59
Q

Post Translational Processing

What does alternative splicing allow for?

A

Usually allows for tissue specific responses of a gene

Allows more possible “outputs”

60
Q

Post Translational Processing

How is alternative splicing used to treat Spinal Muscular Atrophy?

A
  • SMN1 protein is produced to mainatin motor neurons
  • SMA patients have a mutation in SMN1 gene - no protein produced leads to degeneration of neurons
  • Nusinersen effectively hides SMN2 RNA, affecting alternative splicing, producing a functional protein
61
Q

Different forms of RNA

Why is rRNA processed?

A

Ribosomal RNA is processed to produce proper ribosome structure

62
Q

Different forms of RNA

How is rRNA formed?

A

1) Modify many RNA nucleotides
2) Cut it into smaller pieces
1) Pieces come together to form 3 dimensional structure

63
Q

Different forms of RNA

Why is tRNA processed?

A

tRNA is processed to produce specific structures

64
Q

Different forms of RNA

How is tRNA formed?

A

A single strand of RNA forms intramolecular base pairs to fold into specific shapes

65
Q

Different forms of RNA
What helps govern the level of gene expression?

A

the cellular concentration of mRNA

66
Q

Different forms of RNA

Describe the relationship between the rate of mRNA synthesis and the rate of mRNA degredation

A

When synthesis = degradation, mRNA remains constant (steady state)

67
Q

Different forms of RNA

What

A
68
Q

Different forms of RNA

What does miRNA do?
What is it utilized for?

A

Fine tunes gene expression

Utilized to target specific sequences for either silencing or actual degradation

69
Q

Different forms of RNA

What is miRNA made of?
Who utilizes miRNA?

A

22 nucleotide RNA oligomers
Plants and Animals

70
Q

Different forms of RNA

What is unique about the structure of miRNA?

A

Near perfect complement mRNA sequences and can inhibit translation or degrade mRNA

71
Q

RNA Degradation

How is mRNA degraded in prokaryotes?

A

It is degraded by ribonuclease.
Essentially: Individual proteins responsible for cleaving the RNA backbone and chewing up the polymer

  • They utilize enzymes known as endonuclease
  • They cut in the middle portion of the mRNA
  • Exonuclease’s can come from the end and completely digest these particles themselves
72
Q

RNA Degradation

How is mRNA degraded in Eukaryotes?
What enzyme do they utilize?

A

They utilize a very specific enzyme - endonuclease

73
Q

RNA Degradation

What is endonuclease?
What does it do?

A

It’s a large protein complex

It unwinds and feeds mRNA through a nuclease for cleavage

74
Q

RNA and the origins of life

What is a Ribozyme?

What does it suggest?

A

RNAs that fold into highly specific 3D shapes and catalyze reactions

Suggests that perhaps there was an old version of enzymes made of RNA

75
Q

RNA and the origins of life

What is in the catalytic core of ribosomes composed of?

A

The catalytic core is composed of RNA

BIOL 0280 UNIT 4 (6 decks)

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