Topic 5 and 6 Notes
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Think, Pair, Share
The cells in your body share the same DNA sequence, yet cells in different parts of your body look and function differently.
Similarly, bacteria sharing the same DNA, can show different phenotypes when cultured under different environmental conditions.
Question:
How can the same genotype produce different phenotypes?
Gene Expression
Gene expression: the process by which the information encoded in a gene is turned into a functional product (usually a protein, but also RNAs)
Gene Expression
Not all genes are expressed or turned on in a cell
The combination of genes expressed, and the level of expression determine the phenotype of a cell or organism (note: the environment can also impact phenotype, but for this section we will only focus on genotype)
Example: neuron vs muscle cell
Same DNA, but different gene expression
Expression of neuron-specific genes
DNA inside the nucleus of neuron
Expression of muscle-specific genes
DNA inside the nucleus of muscle cell
Example: butterfly wings
The color pattern is due to cells expressing different genes at different levels across the wing
Different genes expressed different colors
The amount of pigment/ protein produced intensity of color
Pattern of gene expression across wing different colors in different locations
Example: variable expressivity
Same genotype, but different phenotypes in different individuals
Seen in some genetic disorders
Different severity of symptoms
Same disease-causing genotype
Gene Expression
Some genes are constitutively expressed
Transcribed at all times
Ensures a continuous supply of essential products for basic cellular functions
Example: housekeeping genes in eukaryotes maintain basic cellular functions like glycolysis
Some genes are inducible
Transcription can be turned “on” or “off” based on environmental and internal cues
Example: Rice plants activate genes in response to pathogens for protection
Gene Expression
Prokaryotes and eukaryotes must be able to regulate gene expression (i.e., which genes are expressed and at what levels)
Regulation helps cells to:
Save energy and resources
Respond to changes in their environment
Develop and maintain different cell types (eukaryotes)
Prokaryotes: regulation occurs primarily at the level of transcription
Eukaryotes: regulation occurs at multiple stages, but many genes are regulated primarily at the level of transcription
Gene Regulation in Prokaryotes
Bacterial Gene Expression
Gene regulation in prokaryotes occurs primarily at the level of transcription
Bacterial Gene Expression
Operons: cluster of related genes that can be regulated and transcribed together under a single promoter
Operons have three parts:
Promoter: RNA polymerase binding site
Operator: the on/off switch
Structural Genes: code for related enzymes in pathway
Bacterial Gene Expression
Regulatory genes: produce regulatory proteins (like repressors and activators) that control the expression of other genes; usually separate from the operon
Repressors: regulatory proteins that reduce transcription when bound to operator (negative regulation)
Activators: regulatory proteins that increase transcription when bound to DNA (positive regulation)
Bacterial Gene Expression
Bacterial Gene Expression
Operons can be repressible or inducible
Repressible (on to off): transcription is usually on, but can be turned off (repressed)
Inducible (off to on): transcription is usually off, but can be turned on (induced)
Repressible Operons
Example: the trp operon in E. coli
Subject to negative regulation
The trp operon in bacteria controls the synthesis of tryptophan
Contains genes needed to make tryptophan
Promoter
Operator
trpE
trpD
trpC
trpB
trpA
Repressible Operons
The trp operon is normally “on” meaning the cell is synthesizing tryptophan
When tryptophan levels build up, tryptophan (which is a corepressor) binds to the repressor, changing its shape
Repressor can now bind to DNA to temporarily shut off transcription for tryptophan, so the cell does not waste energy
Overview: trp Operon
In the absence of tryptophan, the repressor dissociates from the operator, and RNA synthesis proceeds
When tryptophan is present, the trp repressor binds the operator, and RNA synthesis is blocked
Inducible Operon
Example: the lac operon in E. coli
Subject to both negative and positive regulation
Glucose is the preferred energy source, but if levels are low/absent, E. coli can switch to lactose
The lac operon includes 3 genes that encode enzymes for the use/breakdown of lactose
These genes are only expressed when lactose is present
Inducible Operon
Negative regulation
When there is no lactose present, the lac repressor is bound to the operator, preventing transcription
However, when lactose is present, the cell converts lactose to allolactose, which acts as an inducer for the lac repressor
Allolactose binds to the lac repressor repressor changes shape detaches from operator
RNA polymerase can now bind to the operator, but only loosely until it has help from a protein
Transcription is weak until there is positive regulation
CAP site
Promoter
Operator
LacZ
LacY
LacA
Lac repressor is active and bound to the operator
CAP site
Promoter
Operator
LacZ
LacY
LacA
Allolactose binds to lac repressor and inactivates it
Repressor
Glucose present; no lactose = no transcription
Glucose present; lactose present = low levels of transcription
Inducible Operon
Positive regulation
For strong transcription of the lac operon, RNA polymerase needs help binding to the DNA from a protein called CAP, BUT this protein needs to be activated first
When glucose is very low/absent, E. coli gets “hungry” and produces a small molecule called cAMP
cAMP binds to CAP, activating it
cAMP-CAP complex can now bind to the DNA, which enhances RNA polymerase binding, triggering high levels of transcription
Loose binding of RNA polymerase; low levels of transcription
Strong binding of RNA polymerase; high levels of transcription
CAP site
Promoter
Operator
LacZ
LacY
LacA
cAMP
Glucose present; lactose present = weak transcription
No glucose;
lactose present = strong transcription
Quick Check
Fill in the blanks: inducible operons are usually ____ but can be turned _____. In contrast, repressible operons are usually _____ but can be turned _____.
Inducible: Off, on
Represssible: on, off
What are the three parts of an operon?
Promoter: where RNA polymerase can attach. Operator: the on/off switch. Genes: code for related enzymes in pathway
In general, how do repressors differ from activators?
Repressors decrease/block transcription while activators increase transcription.
Gene Regulation in Eukaryotes
Early Gene Expression Patterns
Remember, cells within a multicellular organism have the same DNA sequence. So how does a fertilized egg develop into many different specialized cell types?
Early Gene Expression Patterns
Mitosis
Differential gene expression
Genetically identical daughter cells
Tissue-specific proteins
Specialized cells
During early embryonic development, cells receive internal and external cues that lead to differentiation
Cells become specialized in their structure and function through the expression of different genes for tissue-specific proteins
Early Gene Expression Patterns
Think of these cues as the first instructions about which genes need to be turned on/off in the cells
Establishes early gene expression patterns
Internal cues: cytoplasmic determinants
External cues: inductive signaling
Internal Cues
Cytoplasmic determinants: mRNA and proteins (including transcription factors) from the maternal egg cytoplasm that direct early animal development
Cytoplasmic determinants are unequally distributed in the cytoplasm of an egg cell
When a sperm fertilizes an egg, it forms a zygote
Two cytoplasmic determinants shown in the egg cell
How does this affect early development? As the zygote divides, daughter cells receive different cytoplasmic contents
Each nucleus is exposed to different cytoplasmic determinants
Leads to different patterns of gene expression
External Cues
Induction: cell to cell signals in early development that cause a change in gene expression in nearby cells
One group of cells (inducers) send inductive signals to another group of cells (responder cells) via cell-cell contact or paracrine signaling
Triggers signal transduction pathways in the responder cells (think back to unit 4) that activate transcription factors, changing gene expression
External Cues
Some inductive signals act as morphogens
Morphogens are a specific type of signaling molecule that diffuse through developing tissues, forming a concentration gradient
A cell’s response is dependent on the concentration of the morphogen it was exposed to
Leads to differential gene expression between cells
Different genes are activated based on the concentration the cells were exposed to
Morphogens
Early Gene Expression Patterns
The induction of transcription factors during early development (from both internal/external cues) will turn genes on/off
Some genes may code for other transcription factors Leads to sequential gene expression, ensuring the correct order and timing of development
TF1
Gene expression
TF2
Gene expression
TF2
TF3
Internal or external cue
Quick Check
In your own words, define differentiation.
Cells with the same DNA become specialized by turning some genes on/off
Fill in the blanks: cytoplasmic determinants are ____ cues that come from the ____ egg cytoplasm; inductive signals are _____ cues sent from nearby cells. Both are important for establishing early patterns of gene expression.
Internal; maternal; external
Cytoplasmic determinants are unequally distributed in the egg. What effect does this have on daughter cells during early cell divisions?
Daughter cells inherit different cytoplasmic determinants leading to different patterns of gene expression
Quick Check
Why do different cells respond differently to the same morphogen?
Because morphogens form a concentration gradient. The concentration the cell is exposed to determines the response.
Eukaryotic Gene Regulation
Now that we understand how early gene expression patterns are established, how is gene expression regulated?
Eukaryotic Gene Regulation
Gene regulation in eukaryotes can occur at several different stages:
Chromatin remodeling
Transcription (most genes are regulated at this level)
RNA processing
RNA transport
RNA stability
Translation
Post-translation
Chromatin Remodeling
If DNA is tightly wound it is less accessible for transcription
How can it be modified?
Histone acetylation adds acetyl groups to histones, which loosens the DNA
DNA methylation adds methyl groups to DNA, which causes the chromatin to condense
Chromatin Remodeling
Epigenetic inheritance:
Inheritance that involves changes in how a gene is expressed without any changes in that gene’s nucleotide sequence (i.e., methylation)
Can be passed on during cell division
Modifications can be reversed, unlike mutations
Explains why one identical twin may express a disease while the other does not
Transcription
Once chromatin modifications allow the DNA to be more accessible, specific transcription factors can bind to control elements
Sections of noncoding DNA that can be located near (proximal) or far (distal) from the promoter
Transcription
Distal control elements can be grouped together as enhancers
Can be either upstream or downstream of the transcription start site
The rate of gene expression can be regulated by binding of transcription factors
Transcription
Activators can bind to enhancers to increase the rate of transcription
Even though enhancers are located far from the promoter, DNA is flexible and can bend
Allows activators to interact with mediator proteins, which recruit general transcription factors and RNA polymerase to begin transcription
Transcription
Groups of genes with related functions may be regulated together if they share common control elements recognized by the same transcription factors (even if located on different chromosomes!)
Allows for coordinated regulation of related processes
RNA
RNA processing:
Adding poly-A tail and 5’ cap
Alternative splicing of pre-mRNA
RNA
RNA transport
Controlling access to nuclear pores regulates mRNA transport to cytosol
RNA stability:
Lifespan of mRNA in cytosol affects how many protein molecules can be translated from it
MicroRNAs and small interfering RNAs can bind to mRNA and degrade it or block translation
Translation
The initiation of translation can be blocked by:
Regulatory proteins that bind to sequences in the 5’ cap or poly-A tail, which prevent the ribosome from binding
Modification (activation/ inactivation) of proteins that help ribosomes attach to mRNA
Post-Translation
Protein modifications (i.e., addition/removal of phosphate groups for activation/inactivation or chemical tags for degradation)
