Transcription Flashcards
(186 cards)
1
Q
serves as the instruction manual for making proteins
A
DNA
2
Q
Going from instructions to product occurs in 3 main steps
A
- Transcription
- mRNA editing
- Translation
3
Q
The information contained in the DNA is copied
into a complementary strand of RNA (ribonucleic acid)
A
Transcription
4
Q
RNA copy
A
messenger RNA or mRNA
5
Q
The mRNA copy needs to be modified (cleaned-up) before it leaves the nucleus
A
mRNA editing
6
Q
does mRNA editing happen in
prokaryotic cells
A
no
7
Q
Ribosomes read the mRNA and make a protein
A
Translation
8
Q
is Transcription gene specific
A
yes
9
Q
when Transcription is on what happens
A
Each gene provides instructions for a single protein (usually)
- When turned on, only that one gene is transcribed (unlike DNA rep)
10
Q
types of signals that tells a cell to start transcribing a specific gene
A
1) Constituitive expression
2) Regulated expression
11
Q
Some genes are transcribed continuously
independent of cell health or environmental conditions
A
Constituitive expression
12
Q
what proteins uses Constituitive expression–
A
seen for those proteins that are needed 100% of the time
Example: ATP synthase, actin, tubulin
13
Q
Allows for much tighter control of gene
expression
A
Regulated expression
14
Q
what does Regulated expression do
A
Signal from outside or inside the cell
starts the whole process
15
Q
what kind of pathways does Regulated expression use
A
signal transduction pathways
16
Q
All genes unique region of DNA sequence
upstream of the transcriptional start site that serves as a binding site for the RNA polymerase enzyme
A
promoters
17
Q
General functions of promoters
A
- Provides specificity
- Tell RNA polymerase where to start
- Indicate which strand will be transcribed and the direction of transcription
18
Q
Tells the cell where the gene of interest is
A
Provides specificity
19
Q
Each gene has a unique
A
promoter sequence
20
Q
Although each promoter is unique, most share a few common sequence elements called
A
consensus sequences
21
Q
if consensus sequences was mutated what happens
A
transcription does not take place
22
Q
are consensus sequences evolutionarily conserved
A
yes
23
Q
Prokaryotic promoters share 2 main types of consensus sequences
A
TATA box (aka Pribnow box)
TTGACA
24
Q
found 10 nucleotides upstream from the start site (-10 -> -15), sequence is usually TATAAT
A
TATA box (aka Pribnow box)
25
found 35 nucleotides upstream
from start (-35)
TTGACA
26
RNA polymerase enzyme makes what with the promoter at these consensus sequences
direct contact
27
Binding of RNA polymerase to the promoter
Initiation
28
Prokaryotic cells only produce 1 type of RNA polymerase that produces
mRNA, tRNA, and rRNA in the cell
29
Prokaryotic RNA polymerase structure
2 alpha (α),
1 beta (β),
1 beta prime (β'),
and 1 sigma (σ) subunit
30
β, β’, α subunits interact and form the
CORE ENZYME
31
Together they have the basic catalytic function of producing new RNA
β, β’, α subunits
32
controls binding to the promoter (PROMOTER RECOGNITION)
σ factor
33
After initial binding has been successful, the σ factor does what
comes off the core
34
Bacteria make many different types of
sigma factors
35
Once it binds to the specific promoter, RNA polymerase does what
positions itself over the transcriptional start site
36
positions itself over the transcriptional start site
RNA polymerase
37
Chooses the start site based on its what
distance from the consensus sequences
38
RNA pol unwinds the DNA near the start site called what
transcription bubble
39
what starts the transcription bubble
RNA pol
40
RNA pol reads 1st ~5-10 DNA nucleotides and brings in
complimentary RNA nucleotides
(remember: U inserted across from A)
41
how does RNA pol connect DNA nucleotides and RNA nucleotides
phosphodiester bonds
42
Connects them via phosphodiester bonds
RNA pol
43
how many strands does RNA pol transcribes
one strand
44
in what direction foes RNA polymerase move
5' -> 3'
45
RNA polymerase changes shape after how many nucleotides
~10
46
why does RNA polymerase changes shape
a) Cause it to lose its attraction for the consensus sequences and allows it to begin moving along the gene
b) Cause the sigma factor to fall off
47
RNA pol continues producing a complimentary RNA molecule until it transcribes what
a terminator signal (part of the sequence)
48
why do prokaryotic transcriptional terminator signals exist
a) Slow down the RNA polymerase
b) Weaken the interaction between the DNA and RNA in the bubble
49
Properties of the two major prokaryotic terminator signals
1. Rho-independent terminators
2. Rho-dependent terminators
50
The DNA sequence contains an inverted repeat followed by a string of 6 adenines
Rho-independent terminators
51
Following their transcription, the inverted repeats hydrogen bond with each other within the RNA and form what
hairpin loop
52
Loop formation causes the RNA pol to
pause
53
The pause combined with weak what between
6 straight A-U pairs cause the RNA to totally fall off of the DNA template
hydrogen bonding
54
The pause combined with weak hydrogen bonding between 6 straight A-U pairs
cause the RNA to totally fall off of the DNA template
55
Also contain inverted repeats that cause hairpin loop formation in the new RNA
Rho-dependent terminators
56
Rho-dependent terminators slows down what
polymerase
57
how does Rho-dependent terminators Rho's move
towards the 3’ end
58
Rho-dependent terminators Catches up to the RNA polymerase and unzips what
the RNA from the DNA
59
what does Rho-dependent terminators act as
Acts as a RNA/DNA helicase
60
These do not contain adenine rich region after
the inverted repeat
Rho-dependent terminators
61
Eukaryotic transcription steps
Activation,
initiation,
elongation,
termination
62
General features of transcriptional activation is similar in what
prokaryotic and eukaryotic cells
63
Eukaryotic DNA is tightly coiled around proteins called
histones
64
DNA is negatively charged (phosphate groups),
histones are
very positively charged
65
Every 146 base pairs are wrapped around
a histone octamer (8 histone proteins in a complex) Each group is called a
nucleosome
66
To initiate transcription, DNA in the area of the gene has to be
loosened/unwound from histones and other proteins
67
DNA is freed from histones in 2 major ways:
1) Histone acetylation
2) Chromatin remodeling
68
Enzymes called histone acetyl transferases (HATs) add an acetyl group (CH3CO) to histones neutralizing their positive charge and causes
them to lose their attraction for DNA
Histone acetylation
69
add an acetyl group (CH3CO) to histones
histone acetyl transferases (HATs)
70
neutralizes their positive charge and causes
them to lose their attraction for DNA
histone acetyl transferases (HATs) add an acetyl group (CH3CO) to histones
71
where does this occur: histone acetyl transferases (HATs) add an acetyl group (CH3CO) to histones
only in the area to be transcribed
72
remove acetyl groups after transcription
Deacetylases
73
Eukaryotic promoters
TATA box
CAAT box
GC box
74
located -25-30.
a DNA sequence that signals the start of transcription in genes
TATA box
seen as (TATAAA)
75
Located -70-80
-Mutation of this region usually significantly lowers rate of transcription
a sequence of nucleotides that signals the binding site for RNA transcription factors in eukaryotic genes
CAAT box-
seen in dna as CAAT or CCAAT
76
a DNA sequence that regulates transcription in eukaryotic genes
GC box
seen as GGGCGG
77
Sequences between and around the three common consensus sequences are
different in each promoter
78
recognizes and binds directly to consensus sequences in prokaryotic promoters
A single type of RNA polymerase
79
Eukaryotic cells have multiple RNA
polymerases and none of them recognize
promoter sequences directly
80
Eukaryotic cells have 3 RNA polymerases
1) RNA polymerase I
2) RNA polymerase II
3) RNA polymerase III
81
Produces ribosomal RNA (rRNA)
RNA polymerase I
82
Produces messenger RNA (mRNA)
RNA polymerase II
83
Produces some types of rRNA and all tRNAs
RNA polymerase III
84
All function to transcribe a gene (DNA) into what
RNA
85
Binding of RNA polymerases to the promoter
Initiation
86
If eukaryotic RNA polymerases do not recognize promoter sequences directly,
how do they bind to the correct promoter
WITH THE HELP OF TRANSCRIPTION FACTORS
87
Class of proteins that bind to DNA and help to recruit RNA polymerase enzymes to promoters
Transcription factors
88
Two main classes of Transcription factors
1) Basal transcription factors
2) Regulatory transcription factors
89
(similar function as the sigma factor) in
bacteria)
Transcription factors in eukaryotes
90
Common set of proteins needed to get
transcription started (all promoters use these) – give low levels of transcription
Basal transcription factors
91
Provide more specific transcriptional
control
Regulatory transcription factors–
92
Provide more specific transcriptional
control
Regulatory transcription factors
93
steps for Binding of basal transcription factors to the promoter (and recruitment
of RNA pol II)
1) The TFIID complex binds to the TATA box through its TBP subunit
2) This binding alters the shape of the DNA and allows for binding of
TFIIA and TFIIB
3) RNA polymerase (escorted by TFIIF) then comes in and interact with the
preassembled complex
4) Other factors then come in and help
RNA pol II to gain direct access to
the promoter
94
what does TBP stand for
TATA-binding protein
95
binds to the TATA box through its TBP subunit
The TFIID complex
96
The TFIID complex binds to the TATA box through its
TBP subunit
97
This binding alters the shape of the
DNA and allows for binding of
TFIIA and TFIIB
98
Stabilizes interaction between
TBP and the DNA
TFIIA
99
Helps find the start site
TFIIB
100
RNA polymerase (escorted by
TFIIF
101
(escorted by TFIIF)
then comes in and interact with the
preassembled complex
RNA polymerase
102
Serves as a helicase to separate the strands during transcription
TFIIH
103
will leave most of the other proteins
behind and start making mRNA
RNA pol II
104
when does RNA pol II leave most of the other proteins behind and start making mRNA
Once this giant complex of proteins is
assembled on the TATA box,
105
Binding of basal transcription factors to the promoter (and recruitment
of RNA pol II) occur during all eukaryotic
gene transcription and provides just
basal levels of transcription
106
When a cell wants more or less than just
basal levels of transcription, it will use what
regulatory transcription factors
107
is essentially the same in prokaryotic and
eukaryotic cells
Basic mechanism of elongation
108
Basic mechanism of elongation is what?
- Transcription bubble
- Ribonucleotides added onto the 3' end of the growing RNA
109
Each type of RNA pol utilizes a different
termination mechanism
110
has no No clear termination signal
RNA pol II
111
Transcription continues well beyond the end of the gene
RNA pol II
112
Termination is coupled to mRNA processing (see later)
RNA pol II
113
Eukaryotic and prokaryotic cells have the ability to control BLANK specific genes are transcribed
how often
114
Transcriptional regulation in prokaryotic cells
1) Promoters and sigma factors
2) Gene grouping (operons)
115
recognize different promoters
Different σ factors
116
When a cell needs to transcribe specific genes, it does what
adds the appropriate σ factor to the core components
117
which organize their genes much more efficiently
Bacteria
118
Bacteria typically organize all genes that have a related function together into what is known as an
operon
119
what does an Operons contain
A promoter region
Set of related genes found in tandem (back-to-back)
120
which contains the operator region
A promoter region
121
Binding site for a repressor protein (ON/OFF switch)
Operator
122
all genes that share the same promoter do what
turn on once one of them is
123
steps of Gene grouping (operons)
1) Positive regulation of operons (inducible operons
2) Negative regulation of operons (repressible operons
124
Positive regulation of operons has what kind of operons
inducible operons
125
Negative regulation of operons has what kind of operons
repressible operons
126
Transcription is normally turned off (because the repressor is active when
it is unbound by anything)
Positive regulation of operons (inducible operons
127
example of Positive regulation of operons (inducible operons)
Lac operon
128
comes along, binds to the repressor, and inactivates it
inducer
129
With the repressor shut down what happens
transcription of all genes in the operon is
allowed to occur
130
Transcription is normally on
Negative regulation of operons (repressible operons)
131
example of Negative regulation of operons (repressible operons)
Trp operon
132
comes along and activates the repressor,
corepressor
133
corepressor comes along and activates the what
repressor
134
binds to the operator
The activated repressor then
135
what happens when The activated repressor then binds to the operator
Transcription of all genes in the operon is shut off
136
Some mechanisms of eukaryotic transcriptional control
1) Regulatory transcription factors
2) DNA methylation
137
Basal TFs only provide baseline levels of transcription
Regulatory transcription factors
138
sole function of Regulatory transcription factors
Get the RNA polymerase onto the DNA
139
function to alter transcription above or below basal levels
- Provides fine control over transcription
- Cells have 100s of them
Regulatory TFs
140
Regulatory TFs bind to 2 types of DNA seq
- Promoters
and
- Enhancers
141
Function to increase rates of transcription
Enhancers
142
where are Enhancers located
located far from the promoter (up or down)
143
TF binding causes DNA bending so that
enhancer/promoter close in 3-D
144
Regulatory transcription factors do their job by
a) Binding to DNA and directly altering recruitment/binding/movement of
basal TF and RNA pol
b) Others bind to DNA and recruit or block HATs
145
Increasing or decreasing histone acetylation can alter what
how well basal TF get to the DNA
146
End results of Regulatory transcription factors
Clear the way for basal TF (or increase their affinity) ----> enhancement Block basal TF in any way ----> reduce transcription
147
A methyl group is enzymatically added to
cytosines via specific
methyltransferases
148
A methyl group is enzymatically added to
cytosines via specific methyltransferases
DNA methylation
149
is usually the recognition sequence
CG
150
Methylated bases serve as binding sites for what
methyl-CpG-binding domain proteins (MGDs)
151
These block transcription factor assembly, block chromatin remodeling,
and also recruit histone deacetylases
methyl-CpG-binding domain proteins (MGDs)
152
DNA methylation usually functions to
block transcription
153
Methylation is what specific
cell and tissue-specific
154
different genes become methylated in
different cell types
During cell differentiation
155
Total collection of DNA structural alterations
epigenetics
156
- Take nucleus out of a donor somatic cell
- Inject it into an empty egg and implant into a
surrogate mom
- Baby born will have identical genome as the
donor animal
Somatic cell nuclear transfer procedure
157
Two problems of Somatic cell nuclear transfer procedure
a) Demethylation is never complete in the egg
b) Methylation is affected by the environment
158
what actually happpens with Somatic cell nuclear transfer
A few genes fail to be demethylated
- Start a new animal with certain genes in
the “off” position permanently
Most clones die very young!!
159
what happens because Methylation is affected by the environment
Clone can have different
appearance and personality as the donor
160
Specific case of DNA methylation whereby a given gene is methylated differently depending on what parent it came from
Imprinting
161
which DNA strands are repaired much more efficiently Transcribed DNA strands or non-transcribed strands
non-transcribed strands
162
DNA repair mechanism General steps
1) RNA pol II (eukaryotes) is reading a DNA
strand and making a complimentary RNA
strand when it comes to a damaged nt
2) The damage causes the RNA pol II to
stall (but stay on the DNA)
3) Certain proteins recognize stalled RNA pol II
molecules and bind to it
4) Those molecules recruit DNA repair enzymes to the damage site
5) Damage is fixed and RNA pol II continues
163
- Have defects in the above repair mechanism
- Sufferers have a short stature, age prematurely, and have muscle/bone defects
Cockayne syndrome
164
How do we measure how often a gene is transcribed in a selected cell type
1) Measure promoter/enhancer activity
2) Quantitation of specific mRNA species
165
how to Measure promoter/enhancer activity
1) Insert the promoter/enhancer of a gene of interest in front of a reporter gene
2) Introduce this chimera into a cell of interest and measure how much reporter protein is made
166
how to measure Quantitation of specific mRNA species
1) Northern blot
2) Microarray analysis
167
A more direct approach for measuring transcription is to determine how
much of a
given mRNA species inside the cell
168
If a cell has a ton of mRNA, the gene is being transcribed how
at a high rate
169
If a cell has no mRNA, the gene is being transcribed how
no gene transcription
170
Allows one to determine how large a given mRNA molecule is and how much of that mRNA is in different cell types (Analyze 1 gene at a time)
Northern blot
171
procedure of Northern blot
1. Extract total mRNA from different cell types
2. Separate the different mRNA molecules by gel electrophoresis
3. Transfer separated mRNA molecules from
the gel to an artificial membrane
(nitrocellulose)
4. Add a gene-specific probe that is labeled with radioactivity
- Should only bind to the complimentary mRNA band that is on the membrane
5. A single radioactive band should appear
172
Small, representative portion of a gene (single-stranded)
probe
173
Allows us to obtain a relative quantitation of ALL the mRNA molecules in the cell (Not just one)
(used to determine the effect of cellular changes on global gene expression)
Microarray analysis
174
procedure of Microarray analysis
1. Isolate total cytoplasmic mRNA from cells and convert to single-stranded cDNA (labeled with a dye)
2. Add the labeled cDNA to each spot of a
DNA microarray (aka gene chip)
175
complimentary DNA produced when the
enzyme reverse transcriptase (from HIV) reads
the mRNA and makes a complimentary strand
of DNA
cDNA
176
is the reverse of transcription
mRNA --> DNA
177
Glass chip that has single-
stranded DNA fragments of every gene
spotted individually (species-specific)
DNA microarray
178
how many copies are in No hemoglobin (Hg) mRNA
(0 copies)
179
how many copies are in Little actin (An) mRNA
(50 copies)
180
how many copies are in Med. keratin (Kn) mRNA
(500 copies)
181
how many copies are in High ATP synthase (AS) mRNA
(5,000 copies)
182
Hundreds of different diseases are a direct result of abnormal
transcription of 1 or more genes. Some examples are
1. Cancer
2. Fragile X syndrome
3. Prader-Willi and Angelman syndromes
183
- involves the abnormal activation of 1 or more transcription factors
- Results in the abnormal production of certain proteins, which cause the cell to divide uncontrollably or spread (metastasis)
Cancer
184
- One of most common inherited causes of mental retardation and autism
- Mutation causes the abnormal expansion of the sequence CGG (repeated
hundreds of times)
Fragile X syndrome
185
Abnormal imprinting on chromosome 15 leads to severe neurological problems
Prader-Willi and Angelman syndromes
186
Mom’s gene normally methylated, father’s copy is abnormally methylated (shut-down) – as if they have no copies
Prader Willi
