HRR: Fundamentals of Gene Expression: Transcription I

Question 1

Transcription Template

Answer 1

RNA synthesis is template directed.

The template strand of DNA
is always transcribed in 3´ to 5´ direction by RNA Polymerase

Question 2

Transcription Enzyme:

Answer 2

RNA Polymerase

Question 3

Enzymatic Reaction

Answer 3

(RNA)n + NTP —(Mg2+)–> (RNA)n+1 + PPi

NTP= ATP, CTP, UTP, GTP

Question 4

Processivity***

Answer 4

RNA Polymerases are processive enzymes that transcribe the template strand at rate of 50 nucleotides/sec.

Question 5

Main Steps of Transcription

Answer 5

• Termination
• Initiation
• Elongation

Question 6

do RNA have proofreading?

-why does this matter?

Answer 6

Polymerases do not have

3´ to 5´ exonuclease (proofreading) activity

Question 7

t/f both RNA and DNA synthesis, require a primer ed.

Answer 7

RNA polymerase initiates RNA synthesis de novo by joining 2 ribonucleotides together to form the first 3´to 5´ phosphodiester bond.

Unlike DNA synthesis, a primer is not required.

Question 8

Elongation Reaction of RNA Polymerases

Answer 8

Elongation Reaction of RNA Polymerases

Question 9

Elongation Reaction of RNA Polymerases

Answer 9

note

Question 10

t/f RNA has many different RNA polymerase

Answer 10

FALSE

RNA Polymerase: Prokaryotes have one type of RNA Polymerase

Question 11

Subunit Composition Number
a
b
b´
s
2
1
1
1
Function
Binds regulatory proteins with w subunit
Phosphodiester bond formation, Grasps DNA
Grasps DNA template
Promoter recognition and initiation

Answer 11

Subunit Composition Number
a
b
b´
s
2
1
1
1
Function
Binds regulatory proteins with w subunit
Phosphodiester bond formation, Grasps DNA
Grasps DNA template
Promoter recognition and initiation

Question 12

Gene Promoter: DNA sequence in the template strand of a gene that binds to RNA Polymerase.
In prokaryotes, the promoter of most genes has 2 consensus sequences as shown below.

Answer 12

. Gene Promoter: DNA sequence in the template strand of a gene that binds to RNA Polymerase.
In prokaryotes, the promoter of most genes has 2 consensus sequences as shown below.

Question 13

A consensus sequence is derived by

Answer 13

A consensus sequence is derived by determining the base found most frequently
at each position in the promoter region of the gene.

Question 14

3. Initiation Step of Transcription

RNA Polymerase holoenzyme binds to DNA nonspecifically

Answer 14

3. Initiation Step of Transcription

RNA Polymerase holoenzyme binds to DNA nonspecifically

Question 15

RNA Polymerase searches for ____ site
by “sliding” along double-stranded DNA.
-stops when :

Answer 15

RNA Polymerase searches for promoter site
by “sliding” along double-stranded DNA.
The s subunit binds SPECIFICALLY to the promoter
sequences to form the closed promoter complex

Question 16

closed promotor complex

Answer 16

bold

Question 17

RNA Polymerase unwinds 17 base pairs of DNA around

the initiation site to form the open promoter complex.

Answer 17

RNA Polymerase unwinds 17 base pairs of DNA around

the initiation site to form the open promoter complex.

Question 18

open promotor complex

Answer 18

the sigma subunit is released

Question 19

The first phosphodiester bond

is formed during initiation.

Answer 19

The first phosphodiester bond

is formed during initiation.

Question 20

unwound DNA

Answer 20

location

Question 21

_____ subunit is released upon

completion of the initiation step.

Answer 21

sigma subunit is released upon

completion of the initiation step.

Question 22

Elongation by core RNA Polymerase

Answer 22

Elongation by core RNA Polymerase

Question 23

. Initiation Step of Transcription

Answer 23

. Initiation Step of Transcription
RNA Polymerase Holoenzyme
• There is only one RNA Polymerase in prokaryotes. However, the RNA Polymerase
holoenzyme can initiate transcription of specific genes via the s subunit.
• Multiple s subunits exist, each one binds preferentially to a consensus sequence in the
promoter of a certain gene. Thus, specificity of the RNA Polymerase for a given promoter
is dependent on the s subunit that is part of the holoenzyme.

TTGACA TATAAT
-35 -10 +1
s70
TNNCNCNCTTGAA CCCATNT
\+1
s32
CTGGGNA TTGCA
\+1
s54
Standard
Promoter
Promoter for
heat shock genes
Promoter for nitrogenstarvation genes
Once the s subunit binds specifically to the consensus sequence, helicase activity in the
RNA Polymerase holoenzyme unwinds 17 bp of DNA to form an open promoter complex.

Question 24

Open Promoter Complex

Answer 24

Once the s subunit binds specifically to the consensus sequence, helicase activity in the
RNA Polymerase holoenzyme unwinds 17 bp of DNA to form an open promoter complex.

Question 25

Functions of the s Subunit

Answer 25

• RNA Polymerase holoenzyme is directed to the promoter due to high affinity binding
between the s subunit and the promoter sequence.
• The s subunit has much lower affinity for non-specific DNA sequences, which enables
RNA Polymerase holoenzyme to “slide” along the DNA and find promoter.

Question 26

Elongation Step of Transcription

G-C base-pairing in newly synthesized
RNA forms hairpin, which disrupts its
association with DNA template strand
and halts the transcription reaction.

b) Rho-directed Termination: Rho protein
binds to specific sequences in newly
synthesized RNA. Rho protein has
ATP-dependent helicase activity that
dissociates RNA from template strand.
a) Template-directed Termination: Sequence in
DNA template strand of certain genes contain a
G-C rich repeat region followed by a region of A-T base pairs.
Transcription I [7]
Core Polymerase
Elongation
Core RNA Polymerase
The core RNA Polymerase generates
a transcription bubble that moves along
the chromosome. It consists of the
following:
• Unwound region = 17 bp of DNA
• RNA/DNA duplex = 12 bp

Answer 26

Elongation Step of Transcription

G-C base-pairing in newly synthesized
RNA forms hairpin, which disrupts its
association with DNA template strand
and halts the transcription reaction.

Question 27

Transcription Bubble

Answer 27

The core RNA Polymerase generates
a transcription bubble that moves along
the chromosome. It consists of the
following:
• Unwound region = 17 bp of DNA
• RNA/DNA duplex = 12 bp

Question 28

Termination Step of Transcription (prokaryotes)

Answer 28

a) Template-directed Termination: Sequence in
DNA template strand of certain genes contain a
G-C rich repeat region followed by a region of A-T base pairs.

b) Rho-directed Termination: Rho protein
binds to specific sequences in newly
synthesized RNA. Rho protein has
ATP-dependent helicase activity that
dissociates RNA from template strand.

Weak bonding between
A-U of RNA-DNA duplex
causes dissociation of RNA
from DNA template.

Question 29

Weak bonding between
A-U of RNA-DNA duplex
causes dissociation of RNA
from DNA template.

Answer 29

Weak bonding between
A-U of RNA-DNA duplex
causes dissociation of RNA
from DNA template.

Question 30

RNA Polymerase I

Answer 30

Types of RNA Polymerases

products: 45S Pre-ribosomal RNA [5.8S, 18S, 28S rRNA]
location: Nucleolus

Question 31

RNA Polymerase II

Answer 31

RNA Polymerase II

RNA products: Pre-mRNAs, Primary miRNAs
snRNAs, lncRNAs

Location: Nucleus

Question 32

RNA Polymerase III

Answer 32

products: tRNAs, 5S rRNA,
some snRNAs
location: Nucleus

Question 33

Mitochondrial

Answer 33

RNA products: All mitochondrial RNAs

location: Mitochondria

Question 34

Basic Components of Eukaryotic Transcription by RNA Polymerases

Answer 34

Core RNA Polymerase: Transcription enzyme consisting of multiple protein subunits.
b) General Transcription Factors: Proteins required for basal transcription of all genes.
c) Specific Transcription Factors: Proteins that regulate transcription of specific target genes.
The composition and number of specific transcription factors that regulate each gene differ,
depending on its regulatory sequences. The following terminology is often used for specific
transcription factors:
• Activators (Transactivators) and Repressors (Transrepressors): Regulate transcription
by binding directly to DNA regulatory sequences in a gene. They can also bind to other
proteins including general transcription factors and mediator proteins.
• Mediator Proteins (Coactivators and Corepressors): Do not bind DNA directly, rather
they bind to and interact with general and/or specific transcription factors to regulate their
activity. Many mediator proteins function in modifying chromatin by epigenetic mechanisms.

Question 35

Core RNA Polymerase: Transcription enzyme consisting of multiple protein subunits.

Answer 35

Core RNA Polymerase: Transcription enzyme consisting of multiple protein subunits.
b) General Transcription Factors: Proteins required for basal transcription of all genes.
c) Specific Transcription Factors: Proteins that regulate transcription of specific target genes.
The composition and number of specific transcription factors that regulate each gene differ,
depending on its regulatory sequences. The following terminology is often used for specific
transcription factors:
• Activators (Transactivators) and Repressors (Transrepressors): Regulate transcription
by binding directly to DNA regulatory sequences in a gene. They can also bind to other
proteins including general transcription factors and mediator proteins.
• Mediator Proteins (Coactivators and Corepressors): Do not bind DNA directly, rather
they bind to and interact with general and/or specific transcription factors to regulate their
activity. Many mediator proteins function in modifying chromatin by epigenetic mechanisms.

Question 36

b) General Transcription Factors: Proteins required for basal transcription of all genes.

Answer 36

c) Specific Transcription Factors: Proteins that regulate transcription of specific target genes.
The composition and number of specific transcription factors that regulate each gene differ,
depending on its regulatory sequences. The following terminology is often used for specific
transcription factors:
• Activators (Transactivators) and Repressors (Transrepressors): Regulate transcription
by binding directly to DNA regulatory sequences in a gene. They can also bind to other
proteins including general transcription factors and mediator proteins.
• Mediator Proteins (Coactivators and Corepressors): Do not bind DNA directly, rather
they bind to and interact with general and/or specific transcription factors to regulate their
activity. Many mediator proteins function in modifying chromatin by epigenetic mechanisms.

Question 37

c) Specific Transcription Factors: Proteins that regulate transcription of specific target genes.
The composition and number of specific transcription factors that regulate each gene differ,
depending on its regulatory sequences. The following terminology is often used for specific
transcription factors:
• Activators (Transactivators) and Repressors (Transrepressors): Regulate transcription
by binding directly to DNA regulatory sequences in a gene. They can also bind to other
proteins including general transcription factors and mediator proteins.
• Mediator Proteins (Coactivators and Corepressors): Do not bind DNA directly, rather
they bind to and interact with general and/or specific transcription factors to regulate their
activity. Many mediator proteins function in modifying chromatin by epigenetic mechanisms.

Answer 37

c) Specific Transcription Factors: Proteins that regulate transcription of specific target genes.
The composition and number of specific transcription factors that regulate each gene differ,
depending on its regulatory sequences. The following terminology is often used for specific
transcription factors:
• Activators (Transactivators) and Repressors (Transrepressors): Regulate transcription
by binding directly to DNA regulatory sequences in a gene. They can also bind to other
proteins including general transcription factors and mediator proteins.
• Mediator Proteins (Coactivators and Corepressors): Do not bind DNA directly, rather
they bind to and interact with general and/or specific transcription factors to regulate their
activity. Many mediator proteins function in modifying chromatin by epigenetic mechanisms.

Question 38

Transcriptional Regulation: cis-Acting Elements and trans-Acting Factors

Answer 38

a) cis-Acting Elements: Regulatory DNA sequences specific to each gene, e.g., Core Promoter,
Proximal Promoter Elements, Enhancers, Silencers, Downstream Promoter Elements.

b) trans-Acting Factors: Regulatory proteins derived from genes other than the target gene,
e. g., General and Specific Transcription Factors (activators, repressors, mediator proteins)

Question 39

Transcriptional Regulation

cis-Acting Elements:

Answer 39

Regulatory DNA SEQUENCES specific to each gene, e.g., Core Promoter,
Proximal Promoter Elements, Enhancers, Silencers, Downstream Promoter Elements.

b) trans-Acting Factors: Regulatory proteins derived from genes other than the target gene,
e. g., General and Specific Transcription Factors (activators, repressors, mediator proteins)

Question 40

Transcriptional Regulation:

trans-Acting Factors:

Answer 40

Regulatory PROTEINS derived from genes other than the target gene,
e.g., General and Specific Transcription Factors (activators, repressors, mediator proteins)

Question 41

RNA Polymerase I: Enzyme that transcribes rRNA genes in the nucleolus.

Answer 41

(transcription in eukaryotes)

RNA Polymerase I: Enzyme that transcribes rRNA genes in the nucleolus.

Question 42

rRNA Genes

• The core promoter for rRNA genes is located:

• A human diploid cell contains approximately __#___ rRNA genes arranged in _____……
Each rRNA gene is separated by:

Answer 42

a) rRNA Genes

• The core promoter for rRNA genes is located in nontranscribed spacer DNA and spans the first 167 nt upstream of the transcription start site (+1).
• A human diploid cell contains approximately 400 rRNA genes arranged in tandem repeats
along the 5 acrocentric chromosomes [13, 14,15, 21, 22] in the nucleolus. Each rRNA gene is separated by nontranscribed spacer DNA

Question 43

• RNA Polymerase I transcribes rRNA genes to generate 45S pre-rRNA. This precursor
transcript is processed into 18S, 28S and 5.8S rRNA.

Answer 43

• RNA Polymerase I transcribes rRNA genes to generate 45S pre-rRNA. This precursor
transcript is processed into 18S, 28S and 5.8S rRNA.

Question 44

EM of nucleolar chromatin showing 3 tandem repeats of the rRNA gene separated
by nontranscribed spacer DNA. Each gene is transcribed by multiple RNA Polymerase I
complexes as indicated by the growing array of pre-rRNA transcripts

Answer 44

• Below: EM of nucleolar chromatin showing 3 tandem repeats of the rRNA gene separated
by nontranscribed spacer DNA. Each gene is transcribed by multiple RNA Polymerase I
complexes as indicated by the growing array of pre-rRNA transcripts

Question 45

Components of RNA Polymerase I Transcription Complex

Answer 45

• Core RNA Polymerase I: Multimeric transcription enzyme consisting of 10-12 subunits
• Selectivity Factor 1 (SL1): Complex of 4 general transcription factors that binds to DNA
sequence in the core promoter of rRNA gene
• Upstream Binding Factor (UBF): Specific transcription factor that binds to DNA sequence
in the core promoter of rRNA gene and interacts with SL1 to promote assembly of a
transcriptionally-active RNA Polymerase I complex

Question 46

RNA Polymerase I Complex

Answer 46

RNA Polymerase I Complex

Question 47

• Core RNA Polymerase I: Multimeric transcription enzyme consisting of 10-12 subunits

Answer 47

• Core RNA Polymerase I: Multimeric transcription enzyme consisting of 10-12 subunits
• Selectivity Factor 1 (SL1): Complex of 4 general transcription factors that binds to DNA
sequence in the core promoter of rRNA gene
• Upstream Binding Factor (UBF): Specific transcription factor that binds to DNA sequence
in the core promoter of rRNA gene and interacts with SL1 to promote assembly of a
transcriptionally-active RNA Polymerase I complex

Question 48

• Selectivity Factor 1 (SL1): Complex of 4 general transcription factors that binds to DNA
sequence in the core promoter of rRNA gene

Answer 48

• Selectivity Factor 1 (SL1): Complex of 4 general transcription factors that binds to DNA
sequence in the core promoter of rRNA gene
• Upstream Binding Factor (UBF): Specific transcription factor that binds to DNA sequence
in the core promoter of rRNA gene and interacts with SL1 to promote assembly of a
transcriptionally-active RNA Polymerase I complex

Question 49

• Upstream Binding Factor (UBF): Specific transcription factor that binds to DNA sequence
in the core promoter of rRNA gene and interacts with SL1 to promote assembly of a
transcriptionally-active RNA Polymerase I complex

Answer 49

• Upstream Binding Factor (UBF): Specific transcription factor that binds to DNA sequence
in the core promoter of rRNA gene and interacts with SL1 to promote assembly of a
transcriptionally-active RNA Polymerase I complex

Question 50

RNA Polymerase II: Enzyme that transcribes most genes to synthesize mRNA

Answer 50

RNA Polymerase II: Enzyme that transcribes most genes to synthesize mRNA

Question 51

Core Promoter of Eukaryotic Genes
-def

The core promoter of most genes ranges between _____ bp in length and contains a combination of consensus sequence elements as shown below.

Genes vary greatly with respect to types of sequence
elements that are present in the core promoter.

Answer 51

Core Promoter of Eukaryotic Genes
The core promoter of a gene is defined as the minimal stretch of DNA sequence that is
sufficient to direct accurate initiation of transcription by RNA polymerase II. The core promoter
of most genes ranges between 60-120 bp in length and contains a combination of consensus
sequence elements as shown below. Genes vary greatly with respect to types of sequence
elements that are present in the core promoter.

Question 52

• Initiator Sequence (Inr):

• TATA Box =
• TFIIB Recognition Element (BRE):
• Motif 10 Element (MTE):
• Downstream Promoter Element (DPE):

Answer 52

• Initiator Sequence (Inr): 6 bp sequence that spans the transcription start site (+1)
• TATA Box = TATA(A/T)A: Located -25 to -30 bp upstream of the transcription start site
• TFIIB Recognition Element (BRE): Approximately -35 bp upstream of transcription start site
• Motif 10 Element (MTE): Approximately +18 to +27 downstream of transcription start site
• Downstream Promoter Element (DPE): Approximately +28 to +32 downstream of the
transcription start site

Question 53

Components of RNA Polymerase II Pre-initiation Complex
Transcription of all genes by RNA Polymerase II is dependent on assembly of the following
components into a pre-initiation complex on the core promoter of the gene. It also is called
the basal transcription complex.

• Core RNA Polymerase II:
• General Transcription Factors (TFs):
• Mediator:

The role of specific transcription
factors

Answer 53

Components of RNA Polymerase II Pre-initiation Complex
Transcription of all genes by RNA Polymerase II is dependent on assembly of the following
components into a pre-initiation complex on the core promoter of the gene. It also is called
the basal transcription complex.
• Core RNA Polymerase II: Multimeric enzyme consisting of 12 subunits. The C-terminal tail
domain (CTD) contains multiple Ser residues that are phosphorylated.
• General Transcription Factors (TFs): TFIID, TFIIA, TFIIB, TFIIE, TFIIF, TFIIH.
TFIID consists of: TATA box binding protein (TBP) and 13 TBP-associated factors (TAFs).
• Mediator: Large coactivator complex that is required for the initiation step of transcription.
It consists of approximately 26 proteins that interact with the CTD of RNA Polymerase II,
general transcription factors and specific transcription factors.
Schematic of a pre-initiation complex
assembled on core promoter of a gene.
Specific transcription factors are not
shown. The role of specific transcription
factors is to promote assembly of this
complex on the core promoter of
specific genes to initiate transcription
by RNA Polymerase II above basal
levels.

Question 54

RNA Polymerase II

Answer 54

C. TRANSCRIPTION IN EUKARYOTES
• Initiation of transcription in all genes is dependent on recruitment of RNA Polymerase II to the
the core promoter through assembly of a pre-initiation complex with general transcription
factors [TFIID, A, B, E, F, H] and the mediator complex.

Question 55

Core promoter of any gene:
t/f Sequence elements in the
core promoter are the same among
individual genes.

Answer 55

false

Core promoter of any gene:
Sequence elements in the
core promoter varies among
individual genes.

Question 56

• Current models hold that TFIID directs assembly of the pre-initiation complex, TFIID binds
first to the core promoter along with TFIIA. The TBP subunit binds directly to the TATA box if
present in the core promoter. However, most genes do not have a TATA box sequence in
the core promoter, thus, they are called “TATA-less”. As shown below, TAF subunits function
as a “molecular ruler” that positions TBP at the proper site in the core promoter.

Answer 56

• Current models hold that TFIID directs assembly of the pre-initiation complex, TFIID binds
first to the core promoter along with TFIIA. The TBP subunit binds directly to the TATA box if
present in the core promoter. However, most genes do not have a TATA box sequence in
the core promoter, thus, they are called “TATA-less”. As shown below, TAF subunits function
as a “molecular ruler” that positions TBP at the proper site in the core promoter.

Question 57

Binding of TFII to core promoter:
what ensures that
TBP is positioned correctly relative to
the transcription start site?

Answer 57

Binding of TFII to core promoter:
Note that the TAF subunits form a
C-shaped bridge that facilitates
interactions between upstream and
downstream promoter elements such
as MTE and DPE. This ensures that
TBP is positioned correctly relative to
the transcription start site.

Question 58

• TFIIB, TFIIF, TFIIE and TFIIH are recruited along with the core enzyme of RNA polymerase II.
This mechanism of sequential assembly ensures correct placement of RNA polymerase II
holoenzyme relative to the transcription start site.

Answer 58

• TFIIB, TFIIF, TFIIE and TFIIH are recruited along with the core enzyme of RNA polymerase II.
This mechanism of sequential assembly ensures correct placement of RNA polymerase II
holoenzyme relative to the transcription start site.

Question 59

TFIIB:*

Answer 59

TFIIB:
Binds to BRE if present in the
core promoter.

Question 60

TFIIF & TFIIE:
Assembly and stabilization of
initiation complex. You don’t
need to memorize functions
of TFIIF or TFIIE.

Answer 60

TFIIF & TFIIE:
Assembly and stabilization of
initiation complex. You don’t
need to memorize functions
of TFIIF or TFIIE.

Question 61

TFIIH: *

Answer 61

-when activated it unwinds

TFIIH: Composed of multiple subunits for 2 main functions
1) Helicase activity- unwinding DNA strands in the core promoter
2) Protein kinase activity- phosphorylating CTD of RNA Polymerase II, which facilitates
promoter release for transcription of template DNA

Question 62

Mediator: Protein complex required for initiation step by functioning as a link between
general and specific transcription factors. Mediator proteins are released from the initiation
complex upon phosphorylation of the CTD domain of RNA Polymerase II

Answer 62

Mediator: Protein complex required for initiation step by functioning as a link between
general and specific transcription factors. Mediator proteins are released from the initiation
complex upon phosphorylation of the CTD domain of RNA Polymerase II

Question 63

) Elongation Step: Phosphorylation of CTD facilitates release of the RNA Polymerase II core
enzyme from the promoter. Additional elongation factors are recruited and activated

Answer 63

) Elongation Step: Phosphorylation of CTD facilitates release of the RNA Polymerase II core
enzyme from the promoter. Additional elongation factors are recruited and activated

Question 64

Termination Step: Termination of transcription is coupled to synthesis of a polyadenylation
sequence in the 3´end of pre-mRNA. Cleavage and polyadenylation specificity factor (CPSF)
binds to this polyadenylation sequence, which results in downstream cleavage of the 3´-end.
An exonuclease is recruited that leads to dissociation of RNA Polymerase II from the template.

Answer 64

Termination Step: Termination of transcription is coupled to synthesis of a polyadenylation
sequence in the 3´end of pre-mRNA. Cleavage and polyadenylation specificity factor (CPSF)
binds to this polyadenylation sequence, which results in downstream cleavage of the 3´-end.
An exonuclease is recruited that leads to dissociation of RNA Polymerase II from the template.

Question 65

Transcription overview

Answer 65

DNA (genes) –> RNA

Question 66

RNA Polymerases are ____ enzymes that transcribe the template strand at rate of _____nucleotides/sec.

Answer 66

RNA Polymerases are processive enzymes that transcribe the template strand at rate of 50 nucleotides/sec.

Question 67

A pharmaceutical Co develops an antibiotic that blocks transcription by specifically inhibiting activity of the sigma subunit of bacterial RNA Polymerase. Which of the following functions of RNA polymerase would be inhibited directly?

Answer 67

Recognition of consensus promoter sequence

Question 68

Alice when walking in the woods eats some wild mushrooms =- the next day she vomits and has diarrhea. She goes to the hospital with liver failure and needs a transplant. It is found that the mushrooms contain a toxin that specifically inhibits RNA polymerase II.

Which of the following proces was most likely affected by the toxin?

Answer 68

Synthesis of pre-mRNA

bc RNA polymerase II

Question 69

REGULATION OF TRANSCRIPTION IN PROKARYOTES
1. lac Operon Model (Jacob & Monod)

A single polycistronic mRNA is generated that codes for all of the protein products.

Answer 69

note

Question 70

1. lac Operon Model (Jacob & Monod)

Answer 70

An operon is a set of coordinately expressed genes.
A single polycistronic mRNA is generated that codes for all of the protein products.

lactose—-(B-galactosidase)—> glucose + galactose

Question 71

operon- def

Answer 71

An operon is a set of coordinately expressed genes.

Question 72

Basic Components of Operon

a) Regulator Gene (i): Codes for repressor or activator protein
b) Operator (o): DNA regulatory sequence of the operon
c) Operon: Set of coordinately regulated target genes (z, y, a)
d) Inducer: Nutrient or metabolite such as lactose

Answer 72

Basic Components of Operon

a) Regulator Gene (i): Codes for repressor or activator protein b) Operator (o): DNA regulatory sequence of the operon
c) Operon: Set of coordinately regulated target genes (z, y, a) d) Inducer: Nutrient or metabolite such as lactose

Question 73

lac Operon Model (Jacob & Monod)

• when remove B-galactosidase (in presence of lactose)

Answer 73

stop creating bacterial protein

Question 74

T/F both DNA and RNA can be transcribed

Answer 74

false - only DNA!

Question 75

T/F RNA is usually synthesized in 5´ to 3´ direction.

Answer 75

FALSE

RNA is ALWAYS synthesized in 5´ to 3´ direction.

Question 76

RNA grows in what direction?

Answer 76

5’–>3’

Question 77

a core enzyme vs holoenzyme **

Answer 77

need a sigma subunit to make a holoenzyme

core enzyme : alpha2, W (omega i think), B and B’

holoeznyme: alpha2, W (omega i think), B - B’ - sigma

Question 78

B and B’ subunits

Answer 78

form “pincers” that grasp the DNA template during transcription

Question 79

core enzyme subunits

Answer 79

alpha2, W (omega i think), B and B’

Question 80

(transcription in prokaryotes the initiation step)

Once the s subunit binds specifically to the consensus sequence, helicase activity in the
RNA Polymerase holoenzyme unwinds ____ bp of DNA to form an open promoter complex.

Answer 80

Once the s subunit binds specifically to the consensus sequence, helicase activity in the
RNA Polymerase holoenzyme unwinds 17 bp of DNA to form an open promoter complex.

Question 81

**specificity of RNA Polymerase for a promoter is determined by the ___ subunit.

Answer 81

You don’t need to memorize these sequences or specific s subunits, rather, understand the
concept that specificity of RNA Polymerase for a promoter is determined by the s subunit.

Question 82

transcription buble
• Unwound region =
• RNA/DNA duplex = bp

Answer 82

• Unwound region = 17 bp of DNA

* RNA/DNA duplex = 12 bp

Question 83

Role of Topoisomerases (in prokaryote transcription)
The transcription bubble causes overwinding of the DNA ahead of it. To prevent formation of positive supercoils, prokaryotes use gyrase (Type II topoisomerase) to create negative supercoils ahead of the bubble. As DNA rewinds, negative supercoils can form. Prokaryotes use Type I topoisomerases to relax negative supercoils behind the transcription bubble.

Answer 83

Role of Topoisomerases
The transcription bubble causes overwinding of the DNA ahead of it. To prevent formation of positive supercoils, prokaryotes use gyrase (Type II topoisomerase) to create negative supercoils ahead of the bubble. As DNA rewinds, negative supercoils can form. Prokaryotes use Type I topoisomerases to relax negative supercoils behind the transcription bubble.

Question 84

b) Rho-directed Termination: Rho protein
binds to specific sequences in newly
synthesized RNA. Rho protein has
ATP-dependent helicase activity that
dissociates RNA from template strand.

Answer 84

b) Rho-directed Termination: Rho protein
binds to specific sequences in newly
synthesized RNA. Rho protein has
ATP-dependent helicase activity that
dissociates RNA from template strand.

Question 85

which RNA polymerase take place in the nucleus?

Answer 85

RNA polymerase I and III

Question 86

Core Promoter is an example of ____ element

Answer 86

cis

Question 87

t/f there is only one promoter in a rRNA gene

Answer 87

FALSE

there can be multiple

Question 88

pre-mRNA only contains exons

Answer 88

false contains both introns and exons

gets spliced to the mature mRNA which is only exons

Question 89

t/f every gene has a TATA box

Answer 89

false- only about 15% do

Question 90

TFIID **

Answer 90

TFIID consists of: TATA box binding protein (TBP) and 13 TBP-associated factors (TAFs).

“directs”

Question 91

lac operon

Operator (o):

Answer 91

b) Operator (o): DNA regulatory sequence of the operon

Question 92

Operon: **

Answer 92

Operon: Set of coordinately regulated target genes (z, y, a)

Question 93

Absence of Lactose:

lac operson

Answer 93

transcription is repressed

Question 94

presence of lactose

Answer 94

lactose is an inducer
-protein is inactive

“de-press the gene”

Question 95

when is the lac operon DEPRESSED

Answer 95

when lactose is present

because promotoer is accessible to RNA

Question 96

low glucose and presence of lactose

Answer 96

result

Question 97

Positive Regulation of lac Operon by cAMP
a) Presence of Lactose and Glucose: Transcription of the lac operon does not occur if glucose is still present. This is termed catabolite or glucose repression. Although the repressor
has been inactivated by addition of lactose, RNA Polymerase does not bind to the promoter. High glucose inhibits the expression of enzymes involved in breakdown of alternative sugars such as lactose by lowering cAMP.

Answer 97

Positive Regulation of lac Operon by cAMP
a) Presence of Lactose and Glucose: Transcription of the lac operon does not occur if glucose is still present. This is termed catabolite or glucose repression. Although the repressor
has been inactivated by addition of lactose, RNA Polymerase does not bind to the promoter. High glucose inhibits the expression of enzymes involved in breakdown of alternative sugars such as lactose by lowering cAMP.

Question 98

t/f the core promoter can vary

Answer 98

TRUE

Question 99

cis-Acting Regulatory Elements of a Gene

Transcription II [5]

Answer 99

a) Core Promoter: The minimal stretch of DNA that is sufficient to direct accurate initiation of transcription by RNA polymerase II. It encompasses the transcription start site (+1) and ranges between 60-120 bp in length for most genes. Each core promoter contains a combin- ation of consensus DNA sequence elements such as Inr, TATA box, BRE, MTE and DPE.
b) Proximal Promoter Elements (PPE): cis-Acting sequences located -200 bp or closer to the transcription start site of the gene. They contain DNA sequences that are binding sites for specific transcription factors that function as either activators or repressors of basal transcription. The CCAAT box and GC box are PPEs found in many genes and are located frequently about 75 to 100 bp upstream of the transcription start site.
c) Enhancers: cis-Acting sequences that serve as DNA binding sites for specific transcription factors that function as activators of basal transcription. Enhancers regulate transcription independent of distance from the core promoter and can be located as far as 50 kilobases either upstream or downstream of the transcription start site. Enhancers can still activate transcription when polarity of DNA strands is flipped experimentally in the reverse orientation.
d) Silencers: cis-Acting sequences that serve as DNA binding sites for specific transcription factors that function as repressors of transcription. Like enhancers, silencers can function independent of distance from the core promoter and can be located many kilobases either upstream or downstream of the transcription start site.

Question 100

***A single gene is regulated by multiple types of cis-acting elements.

Answer 100

A single gene is regulated by multiple types of cis-acting elements.

Question 101

a) Core Promoter:

Answer 101

The minimal stretch of DNA that is sufficient to direct accurate initiation of transcription by RNA polymerase II. It encompasses the transcription start site (+1) and ranges between 60-120 bp in length for most genes. Each core promoter contains a combin- ation of consensus DNA sequence elements such as Inr, TATA box, BRE, MTE and DPE.

Question 102

b) Proximal Promoter Elements (PPE): cis-Acting sequences located -200 bp or closer to the transcription start site of the gene. They contain DNA sequences that are binding sites for specific transcription factors that function as either activators or repressors of basal transcription. The CCAAT box and GC box are PPEs found in many genes and are located frequently about 75 to 100 bp upstream of the transcription start site.
c) Enhancers: cis-Acting sequences that serve as DNA binding sites for specific transcription factors that function as activators of basal transcription. Enhancers regulate transcription independent of distance from the core promoter and can be located as far as 50 kilobases either upstream or downstream of the transcription start site. Enhancers can still activate transcription when polarity of DNA strands is flipped experimentally in the reverse orientation.
d) Silencers: cis-Acting sequences that serve as DNA binding sites for specific transcription factors that function as repressors of transcription. Like enhancers, silencers can function independent of distance from the core promoter and can be located many kilobases either upstream or downstream of the transcription start site.

Answer 102

-upstream regulation very close to core promoter

Question 103

c) Enhancers: cis-Acting sequences that serve as DNA binding sites for specific transcription factors that function as activators of basal transcription. Enhancers regulate transcription independent of distance from the core promoter and can be located as far as 50 kilobases either upstream or downstream of the transcription start site. Enhancers can still activate transcription when polarity of DNA strands is flipped experimentally in the reverse orientation.

Answer 103

• can be anywhere

- independent of distance and orientation

Question 104

d) Silencers: cis-Acting sequences that serve as DNA binding sites for specific transcription factors that function as repressors of transcription. Like enhancers, silencers can function independent of distance from the core promoter and can be located many kilobases either upstream or downstream of the transcription start site.

Answer 104

• independent of distance
  d) Silencers: cis-Acting sequences that serve as DNA binding sites for specific transcription factors that function as repressors of transcription. Like enhancers, silencers can function independent of distance from the core promoter and can be located many kilobases either upstream or downstream of the transcription start site.

Question 105

how many types of PPE

Answer 105

2

Question 106

trans-Acting Factors: Proteins that are derived from a gene other than the target gene. trans- acting factors regulate transcription via cis-acting DNA sequences.
• Basal transcription of all genes is dependent on assembly of a pre-initiation complex on the
core promoter. In consists of RNA Polymerase II, general transcription factors and the mediator.
• Transcription of individual genes is regulated by specific transcription factors that function as activators to increase transcription of target genes above basal levels or as repressors to repress or silence transcription of target genes. Activators and repressors bind directly to DNA sequences in cis-acting elements, but they also have domains for recruiting other trans- acting factors that function in transcriptional regulation such as coactivators and corepressors.

Answer 106

trans-Acting Factors: Proteins that are derived from a gene other than the target gene. trans- acting factors regulate transcription via cis-acting DNA sequences.
• Basal transcription of all genes is dependent on assembly of a pre-initiation complex on the
core promoter. In consists of RNA Polymerase II, general transcription factors and the mediator.
• Transcription of individual genes is regulated by specific transcription factors that function as activators to increase transcription of target genes above basal levels or as repressors to repress or silence transcription of target genes. Activators and repressors bind directly to DNA sequences in cis-acting elements, but they also have domains for recruiting other trans- acting factors that function in transcriptional regulation such as coactivators and corepressors.