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MBB164 - Molecular Biology > RNA Structure and Processing > Flashcards

RNA Structure and Processing Flashcards

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1
Q

What is the difference between RNA and DNA nucleotide interactions?

A

RNA nucleotide interactions are more diverse

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2
Q

What are cellular RNAs processed from?

A

larger precursors

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3
Q

What generates small stem-loops structures within RNA?

A

base-pair interactions

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4
Q

What are the 4 nitrogenous bases found in RNA?

A
  • Uracil (pyrimidine)
  • Cytosine (pyrimidine)
  • Adenine (purine)
  • Guanine (purine)
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5
Q

What is a pyrimidine?

A

a nitrogenous base with one ring

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6
Q

What is a purine?

A

a nitrogenous base with two rings

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7
Q

What is RNA?

A

a liner polymer of nucleotides

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8
Q

What is an RNA nucleotide made of?

A
  • ribose sugar
  • mono (or more) phosphate(s)
  • nitrogenous base
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9
Q

What is found at the 5’ end of an RNA polynucleotide?

A

a monophosphate group

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10
Q

What is found at the 3’ end of an RNA polynucleotide?

A

a hydroxyl group

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11
Q

What causes the formation of short helices in RNA polynucleotides?

A

intramolecular base-pairing interactions within the single stranded RNA

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12
Q

What do stem-loops typically have?

A
  • an irregular structure

- caused by interruptions by non-base-paired nucleotides

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13
Q

What type of base-pairing can occur within RNA polynucleotides?

A
  • canonical (Watson/Crick)

- non-canonical (“wobble” base-pairing)

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14
Q

What are the 3 edges of an RNA nucleotide?

A
  • the Watson/Crick edge
  • the Hogsteen (purines) or h/c (pyrimidines) edge
  • the sugar edge
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15
Q

What do the 3 edges allow for?

A

non-canonical base interactions within RNA

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16
Q

How is hydrogen bonding via a nucleotide mediated?

A
  • via ring nitrogen atoms and the attached oxygen atoms or amine groups
  • electronegative ring N atoms and ketone O atoms accept H bonds
  • H atoms bound to the electronegative ring donate H bonds
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17
Q

What makes up an RNA nucleoside?

A

a ribose sugar and nitrogenous base

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18
Q

How is RNA synthesised?

A
  • covalently linking the alpha phosphate at the 5’ position of the incoming nucleotide
  • with the 3’ hydroxyl group of the ribose unit of another nucleotide
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19
Q

In what direction is the RNA extended?

A
  • 5’ to 3’

- RNA retains a 3’ OH group and a 5’ phosphorylated end

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20
Q

What does the RNA polynucleotide originally have?

A
  • a 5’ triphosphate end

- after processing the RNA typically has a 5’ monophosphate group

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21
Q

How do G-U nucleotides base-pair?

A
  • via the same chemical groups that mediate Watson/Crick base-pairing
  • the geometry of the base-pair is unusual
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22
Q

What shape doe heterocyclic bases have?

A

flat and rigid

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23
Q

Which edges can be involved in noncanonical base-pair interactions within RNA?

A

the hogsteen edge and the sugar edge

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24
Q

What does non-canonical base-pairing allow RNA to do?

A
  • allows RNA to adopt unique folding patterns

- which are difficult to predict

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25
Q

What are Tetraloops?

A
  • short loops

- containing 4 nucleotides

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26
Q

How are tetraloops stabilised?

A

through base-staking interactions

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27
Q

What do base-stacking interactions do?

A

contribute significantly to the stability of nucleic acid helices

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28
Q

What is the Sarcin/Ricin Loop?

A
  • an irregular stem-loop structure
  • within large ribosomal RNA
  • critical for the process of translation
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29
Q

What is the Function of the Sarcin/Ricin Loop?

A
  • recognised by elongation factor which binds to the ribosome and drives translocation of the ribosome along the mRNA
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30
Q

What is Ricin?

A
  • well known toxic substance
  • extremely effective at blocking protein synthesis
  • recognises the sarin/ricin loop and removes a single base within the terminal tetra loop structure
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31
Q

How are RNA molecular analogous to proteins?

A
  • have specific 3D shapes

- largely determined by interactions between the nucleotides

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32
Q

What is the secondary structure of RNA?

A
  • a 2D map

- reflects the base-pair interactions which form small helices and stem-loops

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33
Q

What stabilises stem-loops?

A
  • base-pair interactions

- base-stacking interactions

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34
Q

What stabilises small helices?

A
  • ionic base-pairing interactions

- hydrophobic base-stacking interactions

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35
Q

What is coaxial stacking of helices?

A
  • adjacent helices are aligned end to end and stacked on top of each other
  • short helices aligned along the same longitudinal axis
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36
Q

What does coaxial stacking allow for?

A
  • a greater degree of base-stacking interaction than seen in 2 separate helices
  • contributes to the stability of the folded RNA
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37
Q

Are short helices and stem-loops enough?

A
  • may not provide sufficient thermodynamic stability

- RNA may not fold correctly

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38
Q

Similarities between DNA and RNA helices

A

geometries are similar

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39
Q

Differences between DNA and RNA helices

A
  • DNA double helix has a broader “major” groove and a narrower “minor” groove
  • RNA “major” groove is deeper than DNA’s
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40
Q

What is a “long range” tertiary interaction within RNA?

A

interactions between sequences which are well separated in the primary sequence of the RNA

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41
Q

What is a “short range” tertiary interaction within RNA?

A

interactions between closely positioned nucleotides within the primary sequence of the RNA

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42
Q

What is the most common tertiary interaction within RNA?

A

A-minor motif

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43
Q

What is the A-minor motif?

A
  • 2 adjacent A residues interact with adjacent base-pairs in the minor groove
  • through sugar edge interactions
  • long range tertiary structure interaction
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44
Q

What are pseudoknots?

A
  • localised tertiary structure interaction
  • tightly bound structure
  • can be resolved into a single strand by “pulling” the ends apart
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45
Q

How are pseudoknots formed?

A
  • nucleotides within a stem-loop base-pair with nucleotides adjacent to he base of the stem
  • two helical elements are coaxially stacked
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46
Q

What does a pseudoknot result in?

A

tightly folded & compact region of RNA

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47
Q

What type of RNA is predominantly found within a cell?

A

rRNA

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48
Q

What is the most abundant RNA within cells?

A

tRNA

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49
Q

How are RNAs generated from primary transcripts?

A

via processing reactions

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50
Q

What process in the cell do rRNAs and tRNAs function in?

A

translation

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51
Q

What are the molecular events leading to the production of mature, functional RNA called?

A

RNA processing reactions

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52
Q

What analysis technique is used to analyse nucleic acids

A

agarose gel electrophoresis

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53
Q

What type of electrophoresis can shorter nucleic acids be detected on?

A
  • acrylamide gel electrophoresis

- can show rRNA (25S, 18S, 5.8S, 5S) or tRNA

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54
Q

What % does rRNA comprise of cellular RNA?

A

75-80%

55
Q

What % does tRNA comprise of cellular RNA?

A

10%

56
Q

What % does mRNA comprise of cellular RNA?

A

5%

57
Q

What do small nuclear RNAs (snRNA) function as?

A

pre-mRNA splicing

58
Q

What do small nucleolar RNAs (snoRNA) function as?

A

involved in rRNA production

59
Q

What is an example of a regulatory RNA?

A

microRNAs

60
Q

What is cellular RNAs processed from?

A
  • longer precursor molecules

- generated from transcription

61
Q

How are 3/4 of rRNAs in eukaryotic cells processed?

A
  • via the action of endo- and exonuclease
  • cleave RNA within the polynucleotide sequence
  • or shorten the RNA from the 5’ to 3’ end
62
Q

Why is the processing of mRNA transcripts of particular interest?

A
  • sequences are removed through a mechanism not involving RNA cleavage
  • additional RNA sequence is typically added in the form of a poly(A) tail
63
Q

What type of transcripts are eukaryotic mRNA?

A

monocistronic

64
Q

What type of transcripts are prokaryotic mRNA?

A

polycistronic

65
Q

What is a single polycistronic mRNA translated into?

A

multiple functionally related proteins

66
Q

What is a single monocistronic mRNA translated into?

A

a single polypeptide

67
Q

What can the up or down regulation of polycistronic mRNAs lead to?

A
  • the coordinated regulation of all the components within a given biological pathway
  • in a prokaryote
68
Q

What modifications are made to eukaryotic mRNAs?

A
  • structural modifications
  • the 5’ end is capped by the addition of a single conceded guanosine nucleotide
  • the 3’ end is extended by the addition of multiply adenylate residues
69
Q

What do the structural modifications made to eukaryotic mRNAs do?

A
  • 5’ cap and 3’ poly(A) tail protect the mRNA from degradation by ribonuclease
  • stimulate translation of the mRNA
70
Q

What do polycistronic mRNAs allow for?

A
  • the coordinated expression of all functionally related proteins required for a particular biological pathway
  • in one transcript
71
Q

How are functionally related proteins in eukaryotic cells regulated?

A
  • through regulatory control circuits

- act independently on all the gens required for a particular process

72
Q

What do ribosomes recognise in oder to start translation?

A
  • nucleotide sequences

- recognised as translation start and stop signals

73
Q

The translated nucleotide sequence of an mRNA is known as what?

A

an open reading frame (ORF)

74
Q

How many open reading frames do polycistronic mRNAs contain?

A

multiple

75
Q

How many open reading frames do monocistronic mRNAs contain?

A

singular one

76
Q

How are ORFs flanked?

A
  • 5’ leader region

- 3’ untreated region (UTR)

77
Q

What do the 3’ UTR and 5’ leader region contain?

A
  • important regulatory sequence information

- required for the translation and stability of the mRNA

78
Q

What do all RNA Polymerase II transcripts receive?

A
  • a noncoded guanosine nucleotide “cap”

- at their 5’ end

79
Q

How is the 5’ cap attached to the mRNA?

A
  • covalently attached to the 1st transcribed nucleotide

- by a 5’-5’ triphosphate linkage

80
Q

What is different about the noncoded guanosine nucleotide?

A
  • universally methylated at the N7 position

- referred to as an m7G cap

81
Q

What modification is observed in most/all eukaryotic mRNAs?

A
  • a sequence of 70 - 200 noncoded A residues
  • at the 3’ end
  • referred to as a poly(A) tail
82
Q

What modification is observed in complex eukaryote’s mRNAs?

A

methylation of the sugar or base unit of the 1st transcribed nucleotide

83
Q

What is initially required for the termination of RNA Polymerase II transcription?

A
  • a cleavage event

- releasing the mRNA transcript

84
Q

What is this cleavage event dependent on?

A
  • recognition of specific nucleotide sequences

- found at the end of protein coding genes

85
Q

Give an example of a recognition sequence required for RNAP II termination?

A

AAUAAA

86
Q

What event is coupled to the cleavage reaction of RNAP II termination?

A
  • polyadenylation of the 3’ end

- via the enzyme poly(A) polymerase (PAP)

87
Q

What does the enzyme poly(A) polymerase do?

A
  • adds contemplated A residues to the 3’ end of a released mRNA transcript
  • protecting the RNA from degradation
88
Q

What are noncoding sequences in the DNA called

A

introns

89
Q

How are introns removed from mRNA transcripts to produce complete open reading frames?

A

pre-mRNA splicing

90
Q

What carries out pre-mRNA splicing?

A
  • large ribonucleoprotein complex

- called spliceosome

91
Q

What is involved in the pre-mRNA splicing process?

A
  • 2 transesterification reactions

- the elimination of the intron in the form of a lasso/lariat structure

92
Q

What catalyses the splicing reaction?

A

RNA subunits of the spliceosome

93
Q

What are enzymes containing a catalytic RNA subunit called?

A

ribozymes

94
Q

How can the nature of split or interrupted gens be visualised?

A

electronmicroscopy

95
Q

Typically how long are introns?

A

kilobases long

96
Q

Typically how long are exon sequences?

A

a few hundred nucleotides long

97
Q

Approximately what percentage of the primary transcript is intronic?

A

85%

98
Q

How is the accuracy of pre-mRNA splicing achieved?

A
  • recognition of conserved 3’ and 5’ splice sites
  • structural features of the intron
  • the assembly of RNA-binding proteins on sequences found across exons
99
Q

What are common structural features of introns?

A
  • branch point adenosine

- polypyrimidine tract at the 3’ end

100
Q

How does pre-mRNA splicing occur?

A
  • via two consecutive transesterification steps
  • OH group of an alcohol attacks a phosphodiester linkage
  • generating a new phosphodiester linkage
  • releasing another OH group
101
Q

In detail describe the 1st step of the transesterification steps

A
  • 2’ OH group at the branch point Adenosine residue attacks the phosphodiester linkage at the 5’ splice site
  • releasing the 5’ exon
  • generating a lariat-3’ exon
102
Q

In detail describe the 2nd step of the transesterification steps

A
  • OH group at the 3’ end of the 5’ exon attacks the phosphodiester linkage at the 5’ end of the 3’ exon
  • splicing the 2 exon together
  • releasing the intron lariat
103
Q

What 2 nucleotides are found at the 5’ splice site?

A

GU

104
Q

What structural features are found at the 3’ splice sit?

A
  • polypyrimidine tract

- AG nucleotides

105
Q

What carries out pre-mRNA splicing?

A
  • large ribonucleoprotein complex (RNP)

- called a spliceosome

106
Q

What is a ribonucleoprotein complex?

A

a complex containing both RNA and protein

107
Q

What is the differences between a spliceosome and a ribosome?

A
  • splicesosomes are larger than ribosomes

- splicesosomes are less easily visualised due to their dynamic structure

108
Q

What are splicesosomes assembled and disassembled on?

A

pre-mRNA transcripts during the splicing process

109
Q

What does a spliceosome consist of?

A
  • 5 distinct ribonucleoprotein particles

- each containing a single small nuclear RNA (snRNA) and a small number of proteins

110
Q

How are splicesosomes positioned correctly on the primary transcripts?

A
  • base pairing interactions between the snRNAs and the pre-mRNA
111
Q

What happens after splicing has occurred?

A
  • spliced exon and intron lariat is released

- spliceosome is disassembled

112
Q

What are ribozymes?

A

enzymes containing an RNA catalytic subunit

113
Q

What has nuclear pre-mRNA thought to have evolved from?

A

self-splicing introns

114
Q

What is another term for small nuclear RNPs (snRNPs)?

A

snurps

115
Q

Where does the process of transcription take place in eukaryotes?

A

the nucleus

116
Q

Where does the capping of the 5’ end of the transcript in eukaryotes occur?

A

the nucleus

117
Q

Where does the polyadenylation of the 3’ end of the eukaryotic transcript occur?

A

the nucleus

118
Q

Where does the pre-mRNA splicing in eukaryotes occur?

A

the nucleus

119
Q

What type of eukaryotic mRNA transcripts are transported from the nucleus to the cytoplasm?

A

fully processed, mature mRNA transcripts

120
Q

When do pre-mRNA processing events typically take place?

A
  • normally are coupled to transcription

- take place cotranscriptionally

121
Q

How are pre-mRNA processing and transcription coupled?

A

molecular machineries that carry out mRNA capping, splicing and polyadenylation are recruited to the transcript

122
Q

What do molecular machineries which carry out pre-mRNA processing interact with?

A
  • RNA polymerase

- at different points along the length of the gene

123
Q

When does the capping enzyme interact with RNA polymerase?

A

at the beginning of the gene

124
Q

When does the cleavage and polyadenylation complex interact with RNA polymerase?

A

towards the 3’ end of the gene

125
Q

What mediates the coupling of pre-mRNA processing and transcription?

A
  • changing patterns of post-translational modification of the catalytic subunit of RNA polymerase II
  • as the enzyme moves along the gene
126
Q

What are the 3 main steps in eukaryotic pre-mRNA processing that occurs in the nucleus?

A
  • capping of the 5’ end of the mRNA
  • removal of intronic sequences by pre-mRNA splicing
  • 3’ end processing (cleavage and polyadenylation)
127
Q

What do all RNAP II transcripts receive?

A

m7G cap

128
Q

What does the catalytic subunit of RNA Polymerase II contain?

A

an extended C-terminal domain (CTD)

129
Q

What does the CTD comprise of?

A

25-50 copies of a serine-rich heptapeptide sequence

130
Q

How is the CTD domain modified?

A
  • post-transcriptionally

- by the phosphorylation of the serine residues

131
Q

What happens to the pattern of phosphorylation of the CTD as the RNAPs moves along the gene?

A
  • it changes
  • allowing for various complexes required for pre-mRNA processing reactions to bind at different phosphorylated states
  • and so are recruited to the polymerase at different times as it moves along the gene
132
Q

What is the pattern of phosphorylation on the CTD known as?

A

the CTD code

133
Q

What is the serine-rich heptapeptide sequence found in the CTD?

A

YSPTSPS

MBB164 - Molecular Biology (10 decks)

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