Transcription in Prokaryotes: RNA Polymerase [Part 1]

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Questions and Answers

What is the approximate molecular weight of RNA polymerase isolated from E. coli?

470,000 Da (correct)

670,000 Da

100,000 Da

220,000 Da

What is the function of the s-subunit of RNA polymerase?

It helps to unwind the DNA double helix

It is involved in the recognition of specific promoters (correct)

It is responsible for the synthesis of the RNA chain

It combines with other subunits to form the active site for polymerization

What is the direction of RNA synthesis by RNA polymerase?

5' to downstream

3' to 5' (correct)

upstream to 5'

5' to 3'

What is the name of the strand that is complementary to the RNA sequence produced?

Antisense strand Signup and view all the answers

What is the composition of the core enzyme of RNA polymerase?

a2vbb' Signup and view all the answers

What is the function of the b-, b'-, a-, and v-subunits of RNA polymerase?

They combine to make the active site for polymerization Signup and view all the answers

What is the name of the strand that has the same sequence as the RNA produced?

Coding strand Signup and view all the answers

What is the term for the RNA polymerase enzyme that includes all the subunits?

Holoenzyme Signup and view all the answers

What is the primary function of the core enzyme of RNA polymerase?

Providing catalytic activity for transcription Signup and view all the answers

What is the essential role of the s-subunit of RNA polymerase?

Recognition of the promoter Signup and view all the answers

How does RNA polymerase know where to begin transcription?

By recognizing specific DNA sequences Signup and view all the answers

What is the orientation of the promoter sequence relative to the coding strand?

Upstream, to the 5' side of the coding strand Signup and view all the answers

What is the term for the DNA sequence that signals the start of RNA transcription?

Promoter Signup and view all the answers

What is the name of the promoter element about 10 bases upstream of the transcription start site?

Pribnow box Signup and view all the answers

What is the term for the area from the 235 element to the transcription start site?

Core promoter Signup and view all the answers

What is a characteristic of promoter regions in prokaryotic genes?

They are A-T rich Signup and view all the answers

What is the function of the UP element in promoter regions?

Enhancing the binding of RNA polymerase Signup and view all the answers

What is the term for the sequences of bases that are common to many promoter regions?

Consensus sequences Signup and view all the answers

What is the primary function of the promoter base sequence?

To direct the RNA polymerase to the correct gene and control the frequency of transcription Signup and view all the answers

What is the name of the complex formed when RNA polymerase binds to the promoter?

Closed complex Signup and view all the answers

What is the significance of the s-subunit in chain initiation?

It bridges the 210 and 235 regions of the promoter to the RNA polymerase core Signup and view all the answers

What is the process called when RNA polymerase releases most chains near the beginning of transcription?

Abortive transcription Signup and view all the answers

What is the purpose of topoisomerases in transcription?

To relax the supercoils in front of and behind the transcription bubble Signup and view all the answers

What is the name of the region that moves down the DNA sequence to be transcribed during chain elongation?

Transcription bubble Signup and view all the answers

What is the energy source that allows RNA polymerase to break free from the promoter during chain elongation?

The torsional strain of the separated DNA strands Signup and view all the answers

What is the purpose of the s-subunit during chain elongation?

It dissociates after about 10 nucleotides have been incorporated Signup and view all the answers

What is the name of the process by which RNA polymerase moves quickly through some DNA regions and slowly through others?

Variable transcription rate Signup and view all the answers

What is the name of the specific sequences involved in the termination of RNA transcription?

Terminator sequences Signup and view all the answers

What is the characteristic feature of termination sites?

They are complementary and can loop back on themselves Signup and view all the answers

What is the role of uracils in intrinsic termination?

They lead to the dissociation of RNA from the template strand Signup and view all the answers

What is the function of the rho protein in rho-dependent termination?

It chases the RNA polymerase and facilitates the dissociation of the transcription machinery Signup and view all the answers

What is the primary way of controlling transcription in prokaryotes?

Control of transcription initiation Signup and view all the answers

What is the function of alternative σ factors?

They direct the RNA polymerase to different genes Signup and view all the answers

What is the role of enhancers in transcription regulation?

They are sequences that increase the binding of RNA polymerase to promoters Signup and view all the answers

What is the function of the gp28 protein in phage SPO1?

It is an alternative σ factor that directs the RNA polymerase to viral genes Signup and view all the answers

What is the response of E. coli to heat shock?

It produces a protein that directs the RNA polymerase to bind to different promoters Signup and view all the answers

What is the function of Fis sites in E. coli?

They are enhancers that increase the binding of RNA polymerase to promoters Signup and view all the answers

What is the mechanism of intrinsic termination?

The formation of a hairpin loop causes the RNA polymerase to dissociate from the template strand Signup and view all the answers

What is the main function of an enhancer?

To increase the level of transcription when bound to a transcription factor Signup and view all the answers

What is the difference between an enhancer and a promoter?

An enhancer is a DNA sequence that is bound by a transcription factor, while a promoter is a DNA sequence that is bound by RNA polymerase Signup and view all the answers

What is the term for the phenomenon where the production of proteins is triggered by the presence of a suitable substance?

Induction Signup and view all the answers

What is the function of the lacI gene in the lac operon?

To produce a protein that inhibits the expression of the structural genes Signup and view all the answers

What is the term for the DNA sequence that is bound by the repressor protein in the lac operon?

Operator Signup and view all the answers

What is the effect of the inducer on the lac operon?

It removes the inhibition of the structural genes by binding to the repressor Signup and view all the answers

What is the name of the enzyme that hydrolyzes the glycosidic linkage between galactose and glucose in lactose?

b-Galactosidase Signup and view all the answers

What is the function of the lacY gene in the lac operon?

To encode the enzyme lactose permease Signup and view all the answers

What is the term for the group of genes that are controlled as a unit and are often found close together on the DNA?

Operon Signup and view all the answers

What is the term for the type of regulation where the expression of genes is turned on unless something is present to turn it off?

Negative regulation Signup and view all the answers

What is the primary function of the TATA box in eukaryotic promoters?

To orient the RNA polymerase correctly Signup and view all the answers

What is the name of the protein that binds to the TATA box in eukaryotic promoters?

TATA-binding protein Signup and view all the answers

What is the term for the sequence of DNA that surrounds the transcription start site in eukaryotic promoters?

Initiator element Signup and view all the answers

What is the key difference between transcription in prokaryotes and eukaryotes?

The number of proteins associated with transcription Signup and view all the answers

What is the term for the complex formed during transcription initiation in eukaryotes?

Preinitiation complex Signup and view all the answers

What is the function of transcription factors in eukaryotic transcription?

To regulate transcription Signup and view all the answers

What is the term for the elements that are close to the core promoter in eukaryotic promoters?

Upstream elements Signup and view all the answers

What is the characteristic of promoters in eukaryotic genes that lack a TATA box?

They are called TATA-less promoters Signup and view all the answers

What is the function of the general transcription factor TFIID in eukaryotic transcription?

To recognize the TATA box Signup and view all the answers

What is the term for the proteins that are not part of the RNA polymerase complex but regulate transcription in eukaryotes?

Transcription factors Signup and view all the answers

What is the percentage of identical amino acids in the TBP protein across different species?

80% Signup and view all the answers

What is the role of the repressor in the lac operon?

To bind to the operator and prevent transcription Signup and view all the answers

What is the angle of the DNA bend when the TBP protein binds to the TATA box?

80° Signup and view all the answers

What is the function of TFIIF in the transcription initiation complex?

Suppression of nonspecific binding Signup and view all the answers

What is the function of the catabolite activator protein (CAP) in the lac operon?

To bind to the promoter and facilitate transcription Signup and view all the answers

What is the effect of glucose on the lac operon?

It represses the lac operon by reducing cAMP levels Signup and view all the answers

What is the role of TFIIH in transcription initiation?

Helicase activity Signup and view all the answers

What is the outcome of the assembly of the preinitiation complex?

Formation of the open complex Signup and view all the answers

What type of regulation does the lac operon exhibit?

Both positive and negative regulation Signup and view all the answers

What is the term for the genes that are always expressed?

Constitutive Signup and view all the answers

What is the function of TFIIS in transcription elongation?

Release of paused RNA polymerase Signup and view all the answers

What is the function of the lacI gene in the lac operon?

To encode the repressor protein Signup and view all the answers

What is the effect of P-TEF and N-TEF proteins on transcription elongation?

Both A and B Signup and view all the answers

What is the eukaryotic consensus sequence for termination?

AAUAAA Signup and view all the answers

What is the requirement for transcription of the lac operon?

The presence of lactose and the absence of glucose Signup and view all the answers

What is the outcome of termination in transcription?

Both A and B Signup and view all the answers

What is the role of the operator region in the lac operon?

To bind the repressor protein and prevent transcription Signup and view all the answers

What is the effect of cAMP on the lac operon?

It facilitates transcription by binding to the CAP site Signup and view all the answers

What is the function of TFIIF during transcription elongation?

Locking Pol II into an elongation-competent form Signup and view all the answers

What is the classification of the lac operon based on its response to inducers?

Inducible and repressible Signup and view all the answers

What is the primary function of mediator in transcription activation?

To bridge the promoter region with the enhancer region Signup and view all the answers

What is the role of chromatin remodeling complexes in transcription activation?

To mediate ATP-dependent conformational changes in nucleosome structure Signup and view all the answers

What is the effect of the presence of nucleosomes on transcription?

It represses transcription Signup and view all the answers

What is the role of histone-modifying enzymes in transcription activation?

To introduce covalent modifications into the N-terminal tails of histone proteins Signup and view all the answers

What is the function of the head and middle regions of mediator?

To bind to the CTD of RNA polymerase II Signup and view all the answers

What is the function of the tail region of mediator?

To bind to co-activators or co-repressors Signup and view all the answers

What is the ultimate regulator of eukaryotic transcription?

Mediator Signup and view all the answers

What is the role of mediator in basal transcription?

It is required for basal transcription Signup and view all the answers

What is the effect of chromatin remodeling complexes on nucleosome structure?

They loosen the DNA:protein interactions in nucleosomes Signup and view all the answers

What is the function of gene-specific transcription factors in eukaryotic transcription?

To produce a gene-specific level of transcription Signup and view all the answers

Which RNA polymerase is responsible for synthesizing precursors of most, but not all, ribosomal RNAs in eukaryotes?

RNA polymerase I Signup and view all the answers

What is the primary function of transcription factors in eukaryotic transcription?

To direct RNA polymerase to the correct gene Signup and view all the answers

What is the repeated sequence found in the C-terminal domain of RPB1?

PTSPSYS Signup and view all the answers

What is the function of the clamp near the active site of RNA polymerase II?

To lock the DNA:RNA hybrid to the polymerase Signup and view all the answers

How do eukaryotic RNA polymerases differ from prokaryotic RNA polymerase?

They have additional subunits Signup and view all the answers

What is the function of RNA polymerase III?

To synthesize tRNA molecules Signup and view all the answers

What is the significance of the homology between RNA polymerases from bacteria, yeast, and humans?

It suggests that RNA polymerase evolved eons ago Signup and view all the answers

What is the function of RNA polymerase I in eukaryotes?

To synthesize precursors of most, but not all, ribosomal RNAs Signup and view all the answers

What is the structure of RNA polymerase II?

A complex of 10-12 subunits with a molecular weight of 500-700 kDa Signup and view all the answers

What is the function of the jaws formed by subunits RPB1, RPB5, and RPB9 in RNA polymerase II?

To grip the DNA downstream of the active site Signup and view all the answers

What is the characteristic of a positively controlled system?

The gene for the inducer is mutated, making it uninducible. Signup and view all the answers

What is the role of the co-repressor in a negatively controlled repressible system?

It binds to the repressor, preventing transcription. Signup and view all the answers

What is the function of the trp repressor protein?

It binds to the operator, preventing transcription of the trp operon. Signup and view all the answers

What is the mechanism of transcription attenuation in the trp operon?

The formation of secondary structures in the mRNA leader sequence terminates transcription. Signup and view all the answers

What is the role of the 1?2 pause structure in transcription attenuation?

It causes the RNA polymerase to pause, allowing the ribosome to translate the leader sequence. Signup and view all the answers

What is the consequence of the ribosome stopping over the UGA stop codon in the leader sequence?

The 3?4 terminator hairpin forms, terminating transcription. Signup and view all the answers

What is the effect of tryptophan levels on the trp operon?

Low tryptophan levels stimulate transcription of the trp operon. Signup and view all the answers

What is the role of autoregulation in the trpR operon?

It regulates the production of the trp repressor protein. Signup and view all the answers

What is the consequence of the 2?3 antiterminator hairpin forming in the mRNA leader sequence?

Transcription is continued. Signup and view all the answers

What is the role of the leader sequence in the trp operon?

It is involved in transcription attenuation. Signup and view all the answers

What is the primary function of protein kinase A?

To phosphorylate cyclic-AMP-response-element binding protein (CREB) Signup and view all the answers

What is the role of CREB-binding protein (CBP) in transcription activation?

To bridge the response element and the promoter region Signup and view all the answers

Which of the following hormones act on receptors in the nucleus to bind to CBP/p300?

Steroid and thyroid hormones Signup and view all the answers

What is the function of CBP/p300 in addition to being a mediator or coactivator?

To stimulate transcription via loosening of the nucleosomes Signup and view all the answers

What is the consequence of reduction of active levels of CBP?

Development of certain human diseases, including Huntington's disease Signup and view all the answers

What is the role of mitogen-activated protein kinase (MAPK) in transcription regulation?

To phosphorylate transcription factors AP-1 and Sap-1a Signup and view all the answers

What is the function of cyclic-AMP-response-element binding protein (CREB)?

To activate the associated genes Signup and view all the answers

What is the role of CBP in transcription regulation?

To act as a mediator or coactivator Signup and view all the answers

What is the primary goal of remodelers in transcription activation?

To create space between the nucleosomes and expose the DNA Signup and view all the answers

Which of the following histone modifications is associated with transcription repression?

Methylation of lysine 9 Signup and view all the answers

What is the term used to describe the complex set of variables that control transcription activation through histone modification and chromatin remodeling?

Histone code Signup and view all the answers

What is the role of response elements in transcription regulation?

To regulate gene expression in response to certain metabolic factors Signup and view all the answers

Which of the following is an example of a response element?

Heat-shock element (HSE) Signup and view all the answers

What is the role of histone acetyltransferases (HATs) in transcription activation?

To add acetyl groups to histone proteins Signup and view all the answers

What is the role of histone deacetylase (HDAC) in transcription regulation?

To remove acetyl groups from histone proteins Signup and view all the answers

What is the role of ubiquitin in transcription regulation?

To attach to histone proteins and affect transcription Signup and view all the answers

What is the role of cAMP in transcription regulation?

To activate protein kinase A Signup and view all the answers

What is the term used to describe the binding of transcription factors to response elements?

Enhancer binding Signup and view all the answers

Study Notes

Transcription in Prokaryotes

RNA polymerase is the enzyme that synthesizes RNA in prokaryotes.
The molecular weight of RNA polymerase in E. coli is approximately 470,000 Da.
The enzyme has a multisubunit structure, consisting of five different types of subunits: a, v, b, b', and s.
The s-subunit is responsible for recognizing specific promoters and is loosely bound to the rest of the enzyme.

RNA Polymerase Subunits

The b-, b'-, a-, and v-subunits combine to form the active site for polymerization.
The core enzyme (a2vbb') lacks specificity, but the holoenzyme (including the s-subunit) binds to specific DNA sequences.

Template Strand and Coding Strand

One of the two strands of DNA is the template for RNA synthesis, known as the template strand or antisense strand.
The other strand is the coding strand, which has the same sequence as the RNA that is produced (except for U replacing T).
The template strand is read from 3' to 5', and the coding strand is read from 5' to 3'.

Promoter Structure

Promoters are DNA sequences that direct RNA polymerase to the correct gene.
The promoter region is closer to the 3' end of the template strand than the actual gene.
The promoter sequence is usually given based on the coding strand, even though the RNA polymerase binds to the template strand.
Promoters are upstream, meaning they are located to the 5' side of the coding strand and to the 3' side of the template strand.

Components of Promoters

Most bacterial promoters have at least three components:
- The 210 region (also called the Pribnow box) is about 10 bases upstream of the transcription start site.
- The 235 region (also called the 235 element) is about 35 bases upstream of the transcription start site.
- The core promoter region is between the 235 region and the transcription start site.
- The UP element is an optional region that enhances the binding of RNA polymerase.

Chain Initiation

Chain initiation is the first phase of transcription, involving the binding of RNA polymerase to the promoter.
The s-subunit directs RNA polymerase to the promoter.
The formation of the open complex is a critical step in chain initiation.
A purine ribonucleoside triphosphate is the first base in RNA, and it binds to its complementary DNA base at position 11.

Chain Elongation

After the strands have separated, a transcription bubble of about 17 base pairs moves down the DNA sequence to be transcribed.
RNA polymerase catalyzes the formation of phosphodiester bonds between the incorporated ribonucleotides.
The s-subunit dissociates after about 10 nucleotides have been incorporated.
The transcription process supercoils DNA, which is relaxed by topoisomerases.

Chain Termination

Termination of RNA transcription involves specific sequences downstream of the actual gene.
There are two types of termination mechanisms:
- Intrinsic termination, which involves specific sequences called termination sites.
- Rho-dependent termination, which involves a protein called rho.

Transcription Regulation in Prokaryotes

Transcription is controlled in four principal ways:
- Alternative s factors
- Enhancers
- Operons
- Transcription attenuation

Alternative s Factors

Alternative s factors direct RNA polymerase to different genes.
Examples include the action of phage SPO1 and the response of E. coli to heat shock.

Enhancers

Enhancers are DNA sequences that can be bound by proteins called transcription factors.
Enhancers are usually upstream of the promoter region.
When binding the transcription factor increases the level of transcription, the element is said to be an enhancer.

Operons

Operons are groups of genes that are controlled as a unit.
Genes that encode enzymes of certain metabolic pathways are often controlled as a group.
Inducible proteins are produced only in the presence of a suitable substance called an inducer.

Lac Operon

The lac operon is a well-studied example of an inducible protein.
The lac operon includes three structural genes: lacZ, lacY, and lacA.
The expression of these structural genes is controlled by a regulatory gene (lacI) and the mode of operation of the regulatory gene.
The lac operon is induced when E. coli has lactose as its sole carbon source.
The presence of glucose represses the synthesis of the lac proteins.### Regulation of the Lac Operon
The lac operon is regulated by a combination of positive and negative regulation, requiring the presence of lactose and the absence of glucose for transcription.
The operon can be controlled by negative regulation mechanisms, involving a repressor protein that stops transcription when bound to the promoter.
The lac operon is an inducible system, where the presence of an inducer (lactose) releases the repression, allowing transcription to occur.

Types of Operons

Repressible operons: transcription is inhibited by the presence of a corepressor.
Inducible operons: transcription is stimulated by the presence of an inducer.
Constitutive operons: always expressed, even in the absence of an inducer or corepressor.

Trp Operon

The trp operon codes for the enzymes involved in tryptophan biosynthesis.
The operon is regulated by a repressor protein that binds to two molecules of tryptophan, preventing transcription when tryptophan is plentiful.
When tryptophan levels are low, the repression is lifted, allowing transcription to occur.

Transcription Attenuation

Transcription attenuation is a control mechanism that alters transcription after it has begun.
It involves the formation of secondary structures in the mRNA leader sequence, which can either terminate or continue transcription.
The trp operon is regulated by transcription attenuation, where the presence of tryptophan interrupts transcription.

Eukaryotic Transcription

Eukaryotes have three primary RNA polymerases (I, II, and III) that recognize different promoters and transcribe different types of RNA.
RNA polymerase II is responsible for transcribing mRNA precursors and is the most extensively studied.
The structure of RNA polymerase II consists of 12 subunits, with a core subunit that shares sequence homology with prokaryotic RNA polymerase.

RNA Polymerase II Promoters

Pol II promoters have four elements: upstream elements, the GC box, the CAAT box, and the TATA box.
The TATA box is a consensus sequence that is recognized by the TATA-binding protein (TBP).
The initiator element is a sequence that surrounds the transcription start site.

Transcription Initiation

The preinitiation complex is formed by the binding of RNA polymerase II and general transcription factors (GTFs) to the promoter.
The GTFs are required for all promoters and are added to the complex in a specific order.
The TBP protein binds to the TATA box, and the TAFIIs (TBP-associated factors) are also required for transcription initiation.

Elongation and Termination

Elongation is the process of RNA synthesis, and termination is the process of stopping RNA synthesis.
The phosphorylation of the Pol II CTD is required for elongation.
Elongation factors, such as TFIIF and TFIIS, are required for efficient elongation.

Transcription Regulation in Eukaryotes

Gene-specific transcription factors, or activators, are required for high levels of transcription.
Mediator is a protein complex that bridges the promoter region with remote enhancers and silencers, and is required for transcription activation and repression.
Mediator is essential for the transcription of almost every RNA polymerase II-dependent gene.

Transcription and Nucleosomes

Eukaryotic DNA is tightly wound around nucleosomes, making it inaccessible to RNA polymerase II.
Transcription requires the remodeling of chromatin structure to allow access to the promoter regions.### Transcription Repression and Activation
Nucleosomes repress transcription, and activation of eukaryotic transcription requires relief from this repression.
Two circumstances are necessary for transcription activation: interaction of RNA polymerase with the promoter and transcription machinery, and relief of repression caused by chromatin structure.

Chromatin Remodeling Complexes

Chromatin remodeling complexes are large assemblies containing ATP-dependent enzymes that loosen DNA:protein interactions in nucleosomes.
These complexes reorganize nucleosomes to create space and expose DNA, allowing RNA polymerase to be recruited to the promoter.
There are several families of remodeling complexes, including SNF/SWI and RSC families.

Covalent Modification of Histones

Histone modification involves covalent modifications of histone proteins to relax the tight binding between DNA and histones.
Acetylation of lysine residues on histone tails by histone acetyltransferases (HATs) removes positive charges and loosens DNA binding.
Other modifications include phosphorylation of serine residues, methylation of lysine and arginine residues, and attachment of proteins like ubiquitin.
The "histone code" describes the complex regulation of transcription by histone modification.

Response Elements

Response elements are enhancers that respond to specific metabolic factors, such as heat shock, glucocorticoids, metals, and cyclic-AMP.
These elements bind proteins (transcription factors) produced under certain cell conditions, activating multiple related genes.
Examples include the heat-shock element (HSE), glucocorticoid-response element (GRE), and cyclic-AMP-response element (CRE).

Cyclic-AMP-Response Element

Cyclic AMP is produced as a second messenger from hormones like epinephrine and glucagon.
Elevated cAMP levels activate cAMP-dependent protein kinase (protein kinase A), which phosphorylates and activates cyclic-AMP-response-element binding protein (CREB).
CREB binds to the CRE and activates associated genes with the help of CREB-binding protein (CBP) or p300.
CBP/p300 bridge the response element and promoter region, stimulating transcription above basal levels.

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Learn about the structure and function of RNA polymerase in E. coli, including its molecular weight, subunits, and composition.