Topic 5 Flashcards
1
Q
Genotype dictates ___________
A
Phenotype
2
Q
The molecular hierarchy of genetic information
A
DNA (Genotype) - RNA - Protein (Phenotype)
3
Q
Central Dogma
A
DNA - RNA - Protein
4
Q
Simplified process of transcription
A
DNA - RNA
5
Q
Simplified process of translation
A
RNA - Amino Acid Chain
6
Q
Simplified Process of folding
A
Amino acid - Protein
7
Q
Main processes in the Dogma (In order)
A
- Transcription
- Translation
- Folding
8
Q
What does DNA encode
A
mRNA
9
Q
What does mRNA encode
A
Protein
10
Q
What did Francis Crick original “flow of information” propose
A
- Information stored in DNA is unidirectionally transferred to RNA molecules during transcription and to proteins during translation
11
Q
What are the post - model complexities
A
RNA viruses introduced new ways to transduce genetic information across polymers
12
Q
2 post model complexities
A
- RNA can replicate itself
- RNA can be retro transcribed to DNA
13
Q
- RNA can Replicate itself
A
RNA - dependent RNA Polymerase (RdRP) activity in retroviruses
14
Q
Example of RNA replicating itself
A
- RNA viruses that use antisense (non - coding) as genetic material
- For these viruses to complete their life cycle in a cell and produce new virons, they need to use their use their RNA is 2 ways:
1. As a template to generate sense mRNA for protein synthesis
2. As template to replicate its genome material - Both are accomplished without DNA via the activity of RdRP (RNA - dependent RNA polymerases) enzymes
15
Q
- RNA can be Retrotranscribed to DNA
A
Retrotranscriptase (RT) activity in retroviruses
16
Q
Protein + Infection =
A
Prion
17
Q
A
How we understand Dogma Today
18
Q
What is the original self replicator molecule
A
RNA
19
Q
What did RNA do as a self replicator molecule?
A
- Can store information (today’s DNA primary function)
- Can transduce DNA messages into protein (today’s RNA primary function)
- Can read information and catalyze reactions as ribozymes (Today’s proteins primary function)
20
Q
What aspect does an addition of a Nucleus do to transfer of genetic information?
A
Regulates it
21
Q
Transfer of Genetic Information of Prokaryotes
A
- Transcription and Translation happen in the cytoplasm and therefore cannot be timely separate
- This reduced the opportunity to evolve regulatory mechanisms to uncouple these events
22
Q
Transfer of Genetic Information in Eukaryotes
A
- Transcription and Translation happen in different cells compartments and are physically separated
- Synthesized mRNA does not have to be used for translation immediately or at all
- mRNA processing in the nucleus and export needs to occur prior to translation
- This resulted in the evolution of different layers or regulation of gene expression post transcription in eukaryotes
**** See how complex and regulated the transfer is with prensnce of nucleus and membrane bound organelles
23
Q
Steps of the Big Daddy
A
- Gene transcription
- mRNA processing
- mRNA Export
- Escape from RNAi
- mRNA Translation
24
Q
Steps of mRNA processing
A
- 5’ capping
- Polymerization of poly A tail
- Intron splicing
25
Where is RNA synthesized
Nucleus
26
Where is RNA transported to
Cytoplasm
27
What came from the Pulse Chase Labelling
RNA is synthesized in the nucleus and then transported to the cytoplasm
28
What are phages
Phages are DNA viruses that infect bacteria
29
When would be an infection by a phage successful
It must hijack BOTH the transcriptional and translational machineries to make new virons
30
What happens at early infection
Virus takes over bacterial transcriptional machinery to make viral mRNAs
31
What happen at peak of infection or infection
Bacterial mRNA levels drop as the viral genome is used for transcription
32
What happens at late infection
Transcriptional of the bacterial genome increases as virus completes its cycle
33
Timeline of mRNA syntheses
Rapidly induced and rapidly uses up before its degradation in the cell
34
Characterizes of RNA molecules
- RNA is single stranded allowing it to from secondary structures : The shape that RNA molecules that (intra molecular folding) can be as important as its base composition
- Has a OH group in position 2 of the ribose, instead H in DNA
- Utilizes Uracil instead of Thymine. U can pair with A during transcription, but when RNA folds U can pair A or G, which allows more options in forming secondary structures
35
Coding RNAs
mRNA
36
Non Coding RNAS
SnRNAs
tRNA
rRNA
MiRNAs
37
mRNAs
Intermediates that carry genetic information from DNA to the ribosomes
38
SnRNAs
Structural components of sliceosomes
39
tRNA
Adaptors between amino acids and the codons in mRNA
40
rRNA
Structural and catalytic components of ribosomes
41
MiRNAs
Short Single Stranded RNAs that Block expression of complimentary mRNAs
42
General features of RNA synthesis
- RNA synthesis in catalyzed by RNA polymerases and ALWAYS proceeds in the 5’ - 3’ direction
- Only one strand will be used as template (the coding strand)
- Uracyl will the incorporated instead of Thymine
- The RNA molecule will be complementary to the DNA template (antisense) strand and identical to the DNA non template (sense) strand
43
RNA is complimentary to
DNA template strand
44
RNA is identical to ________ (Except U instead of T)
Non template strand
45
Transcription is a _________ event
Dynamic
46
Stages of Transcription in Prokaryotes
1. RNA chain initiation
2. RNA elongation
3. RNA termination
47
Main functions of RNA polymerases
Enzymes responsible to decode DNA into RNA
Aka RNA making machines
RNA polymerases are highly conserved proteins across all life forms
48
How many subunits does RNA polymerase have
5 subunits
49
Phi subunit
Initiation of transcription (released after)
50
Alpha subunit
Assembly of tetrameric core
51
Beta subunit
Ribonucleotide triphosphate binding site
52
Beta ‘ Subunit
DNA template binding protein
53
Omega subunit
Chaperone activity
Regulates correct folding of beta ‘
54
(A) RNA polymerase binding to promoter
- phi subunit associates with the free hold enzyme and recognizes promote (DNA) sequences
- The activity of the phi subunit is responsible for the specificity of RNA polymerase binding
- Bacteria have different phi genes that are specific to controlling initiation of transcription of different target genes
55
(B) Initiation
Phi subunit must leave the holoenzyme to license initiation of transcription (RNA polymerase moves along the template)
56
Initiation of RNA chains
- Binding of RNA polymerase holoenzyme directly to a region in the template strand.
- Localized unwinding of two strands of DNA by RNA polymerases to provide a single stranded template
- Formation of phosphodiester bonds between the first few nucleotides in the nascent RNA chain
57
How does the polymerase know the strand to be used as template?
- The site of RNA polymerase binding is guided by the recognition of conserved cis elements (sequences of DNA) in the promoter region of genes located 5’ of the transcription start site (TSS)
- Promoters are composed of cis elements that are recognized by the RNA polymerase by the sigma (phi) subunit
- Promoter sequences are only present in the template strand, which ensure the proper sense mRNA is made (the polymerase cannot recognize the non template strand)
58
Stage 2 of Transcription: Elongation
- RNA chains grow from 5’ to 3’ (phosphodiester bonds between growing P-5’ and PH - 3’ of free nucleotide)
- RNA polymerases with intrinsic helical activity (to unwind DNA)
- DNA re - winding happens without energy input by reforming hydrogen bonds between the two DNA strands
59
Stage 3 of Transcription: Termination
- RNA polymerase arrives at termination site (TS), decouples from DNA template and the RNA strand in released
- DNA template fully re - anneals by complimentary
60
2 types of transcription termination in bacteria
- pho dependent
- pho independent (requires cis elements in the end of transcript
61
Steps Of rho dependent termination
- RNA pauses at TS
- p binds rut sequence
- RNA polymerase / RNA displacement from template
- RNA release
62
What is this
Pho dependent termination
- at stop codon
- towards the rut sequence
63
What is this?
Intrinsic pho independent termination
64
What does intrinsic termination require?
Stem loop structure in the RNA that is rich in GC upstream of poly - U sequence
65
Steps of intrinsic termination
- RNA polymerase pauses at poly - U
- Stem loop encourages the disassociation of the RNA strand and RNA polymerase
66
Differences between prokaryotic and eukaryotic transcription
67
68
Genome is much bigger in ____________
Eukaryotes
69
Special challenges in Eukaryotic transcription
- Harder to locate a promoter because the genome is much bigger and consequently genes are more spaced out
- Transcription and Translation are decoupled, feedback that regulates transcriptional rates relies on communication across the nuclear membrane
- Access of RNA polymerase to DNA more difficult. Eukaryotic DNA is wrapped up around proteins - histones - that need to be removed / dislodged for transcription to occur.
- Eukaryotic transcription is more complex: the holoenzyme has more subunits, Transcription factors are required to recruit the polymerase (polymerases alone about find and associate to promoters) and there are several types of RNA polymerases
70
What does RNA I polymerase do
Transcribes mRNA and some functional (non - coding) RNAs
71
RNA polymerase II
- Transcribes mRNA and some functional (non coding) RNAs
- Cannot Associate with DNA
- RNA polymerase II is assisted by transcription factors - protein complexes that help it recognize and initiate transcription at the promoter
- Many RNA pol. Promoters contain a TATA box, but not all promoters. These “TATA” - less promoters use other elements to direct polymerase II
72
73
How many subunits does RNA polymerase have
10-12
74
Steps of transcription initiation in Eukaryotes
1. First TFIID binds the promoter at the TATA box via TBP (part of TFIID)
2. Preinitiation complex (PIC) is formed by association of TFIIG, TFIII and TFIIF which bring the polymerase to the site of transcription
3. TFIIF, H, I leave PIC, licensing elongation. The RNA polymerase starts RNA synthesis and leaves the promoter leaving TFIID behind
4. This is coupled to the phosphorylation of RNA polymerase at the C terminal domain
75
When does RNA processing happen
When RNA is being transcribed with the help of RNA polymerase
76
Why does phosphorylation happen at CTDs?
To ensure capping happens first, followed by splicing, and finally poly adenylation of the 3’ end
77
When does RNA processing occur
During Elongation
78
What is the 7 Methyl Guanosine (7-MG) Cap
Unusual 5’-5’ phosphodiester bond between a methylated guanine nucleotide to the first transcribed nucleotide in the RNA
79
3 main purposed of the (7-MG) Cap
1. Transport across the nuclear envelope
2. Protects mRNA from cytoplasmic nucleases
3. Recognition signal for translation machinery
80
The 3’ poly (A) tail formation
1. Recognition of an end of transcriptional signal in the 3’ of the (GT rich) - polymerase stalls
2. Endonuclease activity cleaves transcript downstream of an AU rich region
3. Poly A polymerase recognizes cleaved and processed transcripts as templates to add poly A tail (200 adenine’s added without a DNA template)
81
Purpose of Poly A tail
1, Enhances mRNA stability in the cytoplasm
2. Promoted translation (deadenylated mRNA show reduced translation)
82
Introns
Non coding sequences
83
R loop consists of
RNA - DNA non template strand (Hetroduplex)
84
Introns are only in
Eukaryotes
85
Introns are removed from ______
PremRNA
86
Splicing
- Removal of introns must be very precise
- Conserved sequenced for removal of nuclear mRNA genes are minimal
87
Two main mechanisms of splicing evolved in eukaryotes
1. Self Spl
