Chapters 16.1-2 & 17 - Control of Gene Expression

Question 1

What are the 4 types of regulation parts and their functions?

Answer 1

Structural genes - encode proteins that are used in other pathways (metabolic, biochemical, etc.)

Regulatory genes - encode RNAs or proteins that interact with other DNA sequences and affect transcription or translation of those sequences

Regulatory elements - DNA sequences that are not transcribed, but they still play a role in gene expression

Constitutive genes - structural genes that are expressed continuously and are not regulated

Question 2

What type of control is stimulation of regulation?

Answer 2

Positive control

Question 3

What type of control in inhibition of regulation?

Answer 3

Negative control

Question 4

What are the 6 levels of gene regulation?

Answer 4

Alteration of DNA or chromatin structure
- Highest level of regulation
- Only in eukaryotes becase prokaryotes lack chromatin
- Determines what sequences become available for transcription and the rate at which they’re transcribed

Level of transcription
- Limit or inhibit the transcription of proteins early in the process (enhancers, repressors, insulators, response elements)

Level of mRNA processing
- Can determine the rate at which mRNAs are passed from nucleus to cytoplasm and inhibit the processing of mRNA sequences (splicing, degradation)

Level of mRNA stability
- Amount of protein produced is highly dependent on rate at which mRNA is degraded

Leve of translation
- How is it translated, how is it modified, active vs inactive

Level of posttranslational modifications
- Affects stability and activity of protein

Can have a combination of these

Question 5

What happens with DNA-binding proteins and what are the 3 classes?

Answer 5

Proteins have functional domains, which are responsible for binding to DNA to help regulatory proteins affect gene expression

Most amino acids on protein domains form hydrogen bonds with bases or phosphate-sugar backbone

DNA-binding proteins can also bind into DNA to allow binding protein itself to serve as a substrate to allow another DNA-binding protein to bind in

Classes:
- Helix-turn-helix
- Zinc fingers
- Leucine zipper

Question 6

What are operons?

Answer 6

Groups of bacterial genes that are clustered together and are under control of a single promoter

Regulated at the level of transcription

Question 7

What is the structure of an operon?

Answer 7

Set of structural genes downstream of promoter

Transcribed into single mRNA and translated to produce respective proteins
- Transcription controlled by one promoter

Within operon, there is a regulator gene, but it does not sit in same gene as the others, but is elsewhere that encodes for the regulatory protein - has its own promoter
- Protein that regulator gene produces binds to operon at the operator

Operator sits between the promoter and the structural genes
- Overlaps downstream end of promoter and upstream end of first structural gene

Question 8

What are the two types of transcriptional control within an operon?

Answer 8

Negative control - inhibit transcription through use of a repressor

Positive control - stimulate transcription through the use of an activator
- Activator protein binds to DNA sequence to stimulate transcription

Question 9

What is the idea of inducible operons?

Answer 9

Transcription is not taking place and must be activated/induced

Question 10

What happens with negative inducible operons and what is an example of one?

Answer 10

Example: lac operon

Regulatory gene encodes an active repressor protein that binds into operator to prevent transcription from occurring

Transcription turns on when inducer molecule becomes present
- Binds to repressor on operator, causes conformational change in repressor, which causes the repressor to be pulled off of operator, allowing transcription to occur
- Inducer is typically a precursor to the enzyme/protein that is going to be transcribed

Question 11

What is the idea of repressible operons?

Answer 11

Transcription is taking place and must be shut off/repressed

Question 12

What happens with negative repressible operons and what is an example of one?

Answer 12

Example: trp operon

Regulator protein is also a repressor protein, but it starts in inactive form, so it’s unable to bind to operator

Product of enzymes and biochemical pathways forms co-repressor, which binds to repressor, making the repressor active, so it can bind to operator and shut off transcription

Question 13

Who are the key players in lactose diffusion, breakdown, and metabolism?

Answer 13

Has to be actively transported into the cell by permease

Lactose can be converted to energy, but has to be broken down into galactose and glucose by β-Galactosidase
- Can also break down allolactose into galactose and glucose

Thiogalactoside transacetylase plays an active role in lactose metabolism

Question 14

Is the lac operon negatively repressible or inducible, and what does this mean for transcription?

Answer 14

Negatively inducible - transcription is normally turned off

Question 15

What are the 6 parts of the lac operon and what do they do?

Answer 15

Lac Z - encodes β-Galactosidase

Lac Y - encodes permease

Lac A - encodes thiogalactoside transacetylase

Lac P - promoter

Lac O - operator

Lac I - regulator gene (repressor bound to operon)
- Has its own promoter

Question 16

How does the presence/absence of lactose influence the production of the lac operon parts?

Answer 16

When lactose is absent - few molecules of the lac operon proteins are being produced

When lactose is present - rate of synthesis of proteins increases

Question 17

What is the inducer molecule of the lac operon?

Answer 17

Allolactose

Question 18

How does allolactose act as the inducer molecule for the lac operon?

Answer 18

Repressor inhibits RNA polymerase from binding to promoter

When lactose is present, some of it is converted into allolactose, which binds to repressor, allowing it to come off of promoter

This allows RNA polymerase to bind to promoter and transcribe operon

Question 19

How is permease transcribed if it needs to be present to transport lactose if lactose is not present, and how does β-Galactosidase metabolize lactose if lactose needs to be present first?

Answer 19

Expression is never fully repressed, so there is always a very small amount in the cell that can get the process started

There are also proteins that are similar to allolactose that can bind to repressor to start process

Question 20

What are the 4 types of mutations of the lac operon, where do each occur, and what is its effect?

Answer 20

Structural-gene mutations
- Location: lacZ and lacY
- Effect: alter amino acid sequence of protein encoded by gene in which mutation occurs

Regulator-gene mutations
- Location: lacI
- Effect: affect transcription of structural genes

Operator mutations
- Location: lacO
- Effect: affect transcription of structural genes

Promoter mutations
- Locatio: lacP
- Effect: affect transcription of structural genes

Question 21

What are the 4 other regulation mechanisms?

Answer 21

Enhancers

Anti-sense RNAs
- RNAs of other genes that are complementary of RNAs of regulatory genes that block transcription of RNA

Riboswitches
- Regulatory molecules that affect secondary structure of mRNA, which influences how genes are expressed

Ribozymes
- Induce degradation of mRNA when they’re bound to a certain regulatory molecule

Question 22

What are the 3 differences between eukaryotic and prokaryotic gene expression regulation?

Answer 22

Eukaryotic structural genes have individual promoters and are transcribed separately (no operons)

Chromatin structure affects gene expression

Transcription and translation take place in different parts of the cell owing to a greater diversity of mechanisms
- As sequence is shuttled, almost all levels of regulation can happen during that process

Question 23

What are the 3 similarities between eukaryotic and prokaryotic gene expression regulation?

Answer 23

Types of genes/elements that regulate gene expression

Regulation can occur at several levels along information pathway

Certain similar DNA binding proteins that help regulate gene expression

Question 24

What is epigenetics?

Answer 24

Alterations of DNA/chromatin structure that will affect inheritance of traits, not a change in base sequence

Question 25

What are the 8 types of gene expression regulation in eukaryotes?

Answer 25

Changes in chromatin structure

Transcription factor binding

Insulators

Response elements

Alternative splicing

Degradation of RNA

RNA interference

Translation or posttranslational gene regulation

Question 26

What happens with changes in chromatin structure?

Answer 26

DNA wrapping around histones in chromatin structure causes repression of genes because they are not open to be transcribed

Different things can help change the structure to allow for chromatin structure changes:
- DNase I hypersensitivity sites
- Chromatin remodeling complexes
- Histone modifications
- DNA methylation

Question 27

How do DNase I hypersensitivity sites influence chromatin structure?

Answer 27

As genes become transcriptionally active, there are areas of DNA around it that are sensitive to DNase I (chops up DNA)

These areas adopt a more open conformation

Question 28

How do chromatin remodeling complexes influence chromatin structure?

Answer 28

Complex that remodels chromatin structure without altering chemical structure of histones

Tend to bind into particular DNA sites and reposition nucleosome, which allows for different transcription factors or other transcription proteins to bind into the promoters that are now exposed

Causes nucleosome to slide down DNA, allowing the strand to open the area

Can cause conformational changes in the DNA to expose sites

Can work in tandem with other enzymes or transcription factors to target specific sequences

Can also work in tandem with other enzymes that can alter the histone chemical structure

Question 29

How do histone modifications influence chromatin structure?

Answer 29

Two different domains
Globular domain - associates with DNA and other histones

Positively charged tails - interact with negatively charged DNA wound around them
- Tails can be modified through addition of phophate groups, acetyl groups, or ubiquitin
- Modifications are called the histone code - codes for gene expression
- Alter gene expression by altering chromatin structure directly or by providing recgonition sites for proteins to bind into DNA, which will eventually regulate transcription

Most common modifications - methylation and acetylation
- Methylation can vary in whether it activates or suppresses gene expression - suppression depends on where in the tail the modification occurs
- Acetylation tends to activate/stimulate transcription
- Can also have phosphorylation or ubiquitination

Question 30

How does DNA methylation influence chromatin structure?

Answer 30

Cytosines are typically the only bases methylated in eukaryotic DNA

Typically associated with a repression of transcription

Cytosines that are methylated are often adjacent to guanines - adjacent units called CPGs
- Methylated Cs are diagonal to each other
- A lot of CPG sequences are called CPG islands - found near transcription start sites
- CPG islands are methylated when genes are repressed

CPG islands tend to be associated with deacetylation of histones
- Deacetylation of histones is seen with repression of genes. so CPG islands cause deacetylation

Question 31

What happens with transcription binding factors?

Answer 31

Bind to specific sites in DNA to regulate transcription by recruiting other proteins (co-factors)

Part of basal transcription apparatus and is required for initiation

General transcription factors recruit basal transcription apparatus and bind it to core promoter - will only initiate transcription at minimal levels

Other transcription factors that bind into regulatory promoter can affect whether transcription is activated or repressed
- Dependent on which transcription factors bind to the regulatory promoter

Includes enhancers and silencers - sites that transcription factors bind to that can affect gene expression
- Turn on different genes in different cell types to turn on/off certain genes - very often in development

Question 32

What are enhancers?

Answer 32

Regulatory elements that stimulate transcription

Done to distant genes

Enhance by affecting core promoter

Can loop out DNA near gene being transcribed to enhance transcription

Question 32

What is a silencer?

Answer 32

Regulatory element that represses transcription

Done to distant genes

Silence by affecting core promoter

Can loop out DNA near gene being transcribed to repress transcription

Question 33

What happens with response elements?

Answer 33

Coordinated expression can occur in eukaryotes, but genes are not clustered

Instead, they can respond to the same stimuli because they share short regulatory sequences within promoters or enhancers - known as response elements

At response elements, they recruit and bind transcription factors that activate transcription

If same response elements are at multiple genes, they will be activated in the same way

Single genes can also be regulated by several different response elements
- Allows for same gene to be activated or transcribed by different environmental stimuli
- If one stimulus is gone but another is there, the gene can still be expressed

Question 33

What happens with insulators?

Answer 33

Enhancers can affect different promoters of multiple genes

Insulators are DNA sequences that a protein bind to, which causes the enhancer’s activity to other promoters to be blocked

Question 34

What is coordinated expression?

Answer 34

When genes are grouped together and expressed together (operons in prokaryotes)

Question 35

What happens with alternative splicing?

Answer 35

Create different combinations of exons, creating different proteins downstream

Requires branch point, 5’ splice site, and 3’ splice site

Can have additional sequences called intronic or exonic splicing silencers or enhancers that can affect which sites get spliced in order to create specific proteins

Question 36

What happens with degradation of RNA?

Answer 36

Number of proteins synthesized is dependent on how much RNA template there is

Degradation of RNA prior to translation plays a role in gene expression

More stable RNAs = more proteins

Can have high levels of ribonucleases (eat up RNA) that can have differen paths
- Decapping - removes 5’ cap, causing degradation to occur
- Deadenylation - removes poly(A) tail to reduce stability
- Degradation - destroys RNA strand, resulting in no translation

Question 37

What happens with RNA interference?

Answer 37

Can have small interfering RNA (siRNAs) and microRNAs (miRNAs)
- Either singularly or together can trigger RNA interference

Will combine with different proteins to cleave RNA (causing degradation), inhibit translation, or change chromatin structure through methylation
- Won’t cause methylation, but will recruit demethylating proteins

Question 38

What happens with translation or posttranslational gene regulation?

Answer 38

Availability of the components of translation can play a role in rate of translation

Translational control can also target synthesis of specific proteins
- Are the components for translation going to be available in the cell?

Posttranslational - folding, cleavage, trimming, acetylation, and phosphorylation can all play a role in gene expression through transportation, function, stability, and activity of protein

Question 38

What are transposable elements (TEs)?

Answer 38

AKA: Transposon

Short DNA sequences that can jump around genome

Can insert themselves almost anywhere in the genome

Commonly occur in genome

Jump through genome through different mechanism than homologous recombination

Can cause mutations through movement depending on where they land in genome
- If they land in middle of gene, they’re going to disrupt the gene
- If they land in a promoter, they can disrupt gene expression

Question 39

What are the 2 general elements of a transposable element?

Answer 39

Flanking direct repeats
- Short directly repeated sequences that are found on both sides of most TEs
- Tend not to be part of the TE itself (doesn’t travel with TE) because it’s part of the DNA sequence that the TE has moved into
- Created by staggered cuts in the DNA - overhangs created are complements of each other, forming flanking direct repeats

Terminal inverted repeats
- At ends of most TEs
- Inverted complements of each other
- Regions that tend to be recognized by enzymes that catalyze transposition/movement of TEs

Question 40

What is transposition and its steps?

Answer 40

Movement of TEs from one location to another

Staggered strand breaks made in the target DNA region - created by transposase, which is encoded by TE

Steps:
- TE inserts into DNA that was cut
- Staggered cuts have short, single-stranded pieces of DNA - replication of this DNA creates the flanking direct repeats
- TEs are then fully incorporated into DNA

Question 41

What are the 2 classes of transposons?

Answer 41

Class II - DNA transposons (DNA to DNA)
- Replicative transposons - copy themselves and paste them somewhere else in the genome (old still exists while new is transposed)
- Nonreplicative transposons - cut itself out of genome and paste somewhere esle

Class I - Retrotransposons (DNA transcribed into mRNA intermediate and then reverse transcribed into DNA using reverse transcriptase)
- Only undergo replicative transposition

Question 42

How can transposable elements be controlled by methylation in DNA?

Answer 42

Alterations in chromatin structure can affect transcription of transposon

Can also be caused by translational effects that prevent transposon from being translated

Question 43

How can scientists use TEs?

Answer 43

Can use them to induce mutations in the genome

Allows them to understand function of genes and gene mapping

Question 44

What are TEs in bacteria like?

Answer 44

Only have DNA transposons

Groups
Simple TEs:
-AKA: insertion sequences - carry only genetic information for movement

Composite transposons - consist of two insertion sequences, flaning a segment of DNA
- 2 insertion sequences with a bacterial gene between them
- Each of the insertion sequences have two terminal inverted repeats and a single flanking direct repeat
- Everything travels together, including gene between them

Non-composite transposons
- Don’t have insertion sequences, only have inverted repeats
- WIll have gene between them

Question 45

What are TEs in eukaryotes?

Answer 45

Two groups:
Short inverted repeats

Retrotransposons:
- Mechanism of movement is different than other TEs, but they still generate flanking repeats at point of insertion

Question 46

What did McClintock discover and how does it affect pigmentation in corn?

Answer 46

Discovered that pigmentation in corn kernels was caused by a transposable element that moves between pigmentation gene to produce yellow, purple, or striped varieties

Described Ac and Ds elements
- When Ds elements were there, Ac had to be there also
- With more research, it was noticed that Ds and Ac were transposons
- Ac elements code for transposase, so they can transpose themselves
- Ds has deactivated form of transposase, so they require Ac to transpose

Pigmentation is controlled individually in every corn kernel
- For heterozygous individuals (Cc), Ds element sometimes can get transposed into C locus if near Ac element - when transposed it shuts off pigmentation gene
- Sometimes Ds element during development will transpose out of the gene, turning back on pigmentation
- Depending on where gene is jumping or when it leaves, there are different amounts of purple pigmentation in the corn - if excised early, there will be more purple pigmentation; if excised later, there will be less purple pigmentation