29. Regulation of gene expression
Learning objectives
- Describe the different gene “categories”
- What are promoters?
- What is the common structure of promoters?
- What are repressors?
- What are activators?
- Describe the DNA-protein binding domains
- What DNA-protein binding domain is found in HIF-1α?
- Describe the function of the lac operon
- Describe the SOS response in bacteria
- Describe in trans regulation of mRNA
- Describe in cis regulation of mRNA
- What are the differences in gene expression between prokaryotes and eukaryotes?
- What are epigenetics?
- Describe DNA methylation
- Describe histone modification
- What is the histone code?
- Describe nucleosome remodeling
Principles of gene regulation
Only a fraction of the genes of an organism are expressed at any given time. The amount of gene product transcriped also depends on the gene. Some proteins are present in very large amounts, like elongation factors used for protein synthesis, while others, like enzymes which repair rare DNA lesions, may only be present as a few molecules.
Some gene products are useless alone, and only useful when synthesised together with the products of other genes. Imagine a cell which increases its transcription of glucose 6-phosphatase, but not the transcription of the other enzymes of gluconeogenesis.
Different gene categories
The genome of an organism contains many genes. Some genes are always expressed, while some are only expressed at certain times. Accorrding to this we can distinguish between different gene “categories”.
Housekeeping genes, also called constitutive genes, are genes which are always expressed. These includes genes coding for proteins related to processes which are always active, like proteins involved in transcription and translation, and central metabolic pathways.
Cell type-specific genes are genes which are only turned on in certain cell types, which give each cell type its special properties and functions. For example, only β cells of the Langerhans islets express the gene for insulin.
Developmental regulatory genes are genes which are turned on during certain stages of growth and development of an organism. For example, the protein SRY, which is involved in determining the sex of the organism, is only expressed during a few days of foetal life.
Inducible genes are genes which are normally off, but which are turned on in response to external stimuli. This includes genes which are activated in response to hormones, or genes involved in DNA repair.
Repressible genes are genes which are normally on, but which are turned off in response to external stimuli. For example, when bacteria have good supply of tryptophan they repress the genes involved in tryptophan biosynthesis.
Promoters
RNA polymerase initiate transcription when binding to promoters in the DNA. Promoters are small segments of DNA upstream to (“before”) the gene. The structure of the promoter influence RNA polymerase‘s affinity to the gene in question, thereby regulating the rate of transcription. Promoters are usually AT-rich; common promotors include the TATA box and the CAAT box.
Regulatory proteins can enhance or interfere with the interactions between RNA polymerase and the promotor, thereby also influencing transcription. Repressors interfere with these interactions, while activators enhance them.
Repressors bind to binding sites on the DNA called operators, which are generally between the promoter and the gene. When repressors are bound to the operator, RNA polymerase can’t move along the DNA, which is how they prevent transcription.
Activators bind to regions called enhancers. Unlike operators, enhancers are often far away from the promoter. When activators bind to enhancers they somehow increase the binding of RNA polymerase to the promoter.
Repressors and activators aren’t always bound to their operators and enhancers – they’re only bound in response to a certain molecular signal. We’ll see an example of this later. This molecular signal, often a small molecule, changes the conformation of the regulatory protein, which causes it to bind to or dissociate from the DNA.
Recall the function of the specificity factors of RNA polymerase, which was discussed in topic 21. RNA polymerase can switch out its σ-subunit depending on what needs to be expressed, changing the affinity of RNA polymerase to different promoters. The σ70 subunit binds to the promoters of housekeeping genes, and so the σ70 subunit is the subunit which is active most of the time. During heat stress the σ70 subunit is switched out with σ32, which changes the affinity of RNA polymerase so that it binds to the promotors of heat-shock genes.
DNA-protein binding
For proteins like activators and repressors to bind to DNA they must have certain DNA-binding domains or motifs. The most important types of DNA-binding domains are helix-turn-helix, zinc fingers, homeodomain, leucine zipper, and helix-loop-helix.
DNA-binding proteins must not only bind DNA, but they must bind to very specific parts of DNA, i.e. a specific operator. For this to be possible the DNA-binding proteins must have amino acids side chains which bind to specific nucleotide sequences. These amino acids are often basic, which allows them to bind to the negatively charged DNA. For example, glutamine and asparagine side chains bind to adenine bases. Arginine side chains bind to guanine bases.
Helix-turn-helix is a 20 amino acid long domain consisting of some α-helical segments and a β turn. It’s mostly found in bacteria, most notably in the Lac repressor.
Zinc fingers is a 30 amino acid long domain containing zinc ions. It’s mostly found in eukaryotes. Multiple zing fingers are found in one DNA-binding protein.
Homeodomain is a 60 amino acid long domain. It’s mostly found in eukaryotes, but is similar to helix-turn-helix.
Leucine zipper is a 60 – 80 amino acid long domain. The name comes from how every seventh amino acid is leucine, and that the residues intertwine like a zipper. Leucine zipper is found in the CREB transcription factor.
Helix-loop-helix is comprised of two amphipatic α-helixes linked by a loop. This 50 amino acid long domain is found in many transcription factors, most importantly in HIF-1α, which we’ll discuss in detail later.
Gene regulation in bacteria
The lac operon
An operon is a cluster of genes that are controlled by the same promoters, and which are transcribed together. The best-known example of operons is the lac operon in E. coli. This operon codes for genes that are needed for lactose metabolism. However, to save resources, this operon should not be expressed when there is no lactose available for the bacteria anyway.
The lac repressor represses these genes when lactose is not available. When lactose is available, the lac repressor stops repressing these genes, so genes needed for lactose metabolism are expressed. However, when both glucose and lactose are available, lactose metabolism should be inhibited anyway, because glucose metabolism is more efficient than lactose metabolism.
The SOS response
The SOS response in bacteria is a cellular response to great DNA damage. During the response the cell cycle is arrested and DNA repair is induced.
E. coli contains genes for many proteins that try to fix this DNA damage. However, during normal conditions, these genes are inhibited by a repressor called LexA. When DNA damage occurs, a protein called RecA binds to the damaged DNA. RecA then breaks down LexA, so that it doesn’t repress the SOS response genes anymore. When this repression is broken, the genes are expressed, so the proteins can try to fix the DNA damage.
Regulation of bacterial mRNA function
Protein synthesis is not only regulated at the transcriptional level, but also the translational level. Bacterial mRNA can be regulated in two ways, in trans or in cis. Every mRNA contains a site where the ribosome binds to it. If this site is difficult to reach, then the mRNA has more trouble binding to the ribosome, so it’s translated less often. If the site is easy to reach, it’s translated more often.
In trans regulation refers to when small RNA molecules bind to mRNA and either block or expose the ribosome-binding site.
In cis regulation involves mRNA which contain ligand-binding structures called aptamers. These aptamers can bind certain ligands, like TPP, glycine or adoMet. When the aptamers bind these ligands, the mRNA coils, which makes the ribosome-binding site harder to reach, thereby inhibiting translation.
Gene expression in eukaryotes
Differences in regulation of gene expression
Gene expression in eukaryotes is different than in prokaryotes on several levels. In bacteria, transcription and translation happens at the same time and place (in the cytosol), while in eukaryotes, they’re separated in both time and space. This means that they don’t take place simultaneously, and not in the same cellular compartment.
In bacteria, most genes are active while only certain ones are repressed, while in eukaryotes, most genes are inactive, and only the ones that are needed are activated.
Bacteria contain no histones, and have different chromatin structure than eukaryotes.
Epigenetics
Epigenetics refers to reversible heritable chemical modifications of DNA and histones, which regulate gene expression in eukaryotes. There are multiple mechanisms of epigenetics, the most important being DNA methylation, histone modification, and nucleosome remodeling.
Epigenetics are influenced by environmental factors, like diet, exercise, chemicals, drugs, etc. They’re a testament to how your parents’ lifestyles can affect you; not by changing the genes themselves, but by altering their transcription pattern.
DNA methylation
DNA methylation refers to methylation of cytosine bases in promoter regions in DNA, converting them to 5-methylcytosine. This process typically decreases the affinity of RNA polymerase to the promoter, thereby inhibiting gene transcription. This only occurs on cytosine bases in promoters called CpG islands.
Recall from the DNA repair topic that when a DNA strand is duplicated the new strand is methylated just like the template strand.
Histone modification
Recall that a nucleosome is the complex of DNA around a histone. These histones are involved in regulating the structure of chromatin. By covalently modifying the “tails” of the histones, the structure of chromatin can be altered. Modifications which cause DNA to wrap around the histone tighter decrease gene transcription, and vice versa. We say that the DNA coiling is relaxed or tightened.
The histone code refers to the patterns of covalent modification of the histones, of which there are billions of possible combinations.
Acetylation of a histone decreases the number of positive charges on the histone, thereby relaxing the DNA coiling, increasing transcription. Deacetylation does the opposite. These processes are catalysed by histone acetyltransferase (HAT) and histone deacetylase (HDAC).
Methylation of a histone thightens the DNA coiling, decreasing transcription. This methylation occurs on the lysine residues of the histones, of which there are many. This process is catalysed by DNA methyltransferase (DNMT)
Nucleosome remodeling
Certain proteins can remodel the nucleosomes, thereby changing the coiling of DNA to influence gene expression. In eukaryotes the most important chromatin remodelers is the SWI/SNF protein complex.
Summary
- Describe the different gene “categories”
- Housekeeping genes, also called constitutive genes, are genes which are always expressed
- Cell type-specific genes are genes which are only turned on in certain cell types, which give each cell type its special properties and functions
- Developmental regulatory genes are genes which are turned on during certain stages of growth and development of an organism.
- Inducible genes are genes which are normally off, but which are turned on in response to external stimuli.
- Repressible genes are genes which are normally on, but which are turned off in response to external stimuli
- What are promoters?
- Promoters are small segments of DNA upstream to the gene
- This is the region recognized by RNA polymerase
- What is the common structure of promoters?
- Promoters are usually AT-rich; common promotors include the TATA box and the CAAT box.
- What are repressors?
- Repressors are regulatory proteins which bind to operators, thereby preventing transcription
- What are activators?
- Activators are regulatory proteins which bind to enhancers, thereby increasing transcription
- Describe the DNA-binding domains
- DNA-binding domains are domains on protein which bind to and recognize a specific DNA sequence
- Helix-turn-helix is found in bacteria, most notably in the Lac repressor
- Zinc fingers are mostly found in prokaryotes
- Homeobox is found in eukaryotes, but is similar to helix-turn-helix
- Leucine zipper has a leucine as every 7th amino acid residue
- Helix-loop-helix is found in many transcription factors
- What DNA binding domain is found in HIF-1α?
- Helix-loop-helix
- Describe the function of the lac operon
- The lac operon is a cluster of genes which codes for proteins involved in lactose metabolism
- When lactose is unavailable the lac repressor represses the lac operon
- When lactose is available the lac repressor stops repressing these genes
- Describe the SOS response in bacteria
- The SOS response is a response to great DNA damage, in which the cell cycle will be arrested and DNA repair will be induced
- During normal function a repressor called LexA represses the DNA repair genes
- When DNA damage occurs RecA breaks down LexA, allowing transcription of DNA repair genes
- Describe in trans regulation of mRNA
- In trans regulation refers to when small RNA molecules bind to mRNA and either block or expose the ribosome-binding site.
- Describe in cis regulation of mRNA
- In cis regulation refers to when the mRNA contains ligand-binding structures called aptamers, which coils the mRNA when certain ligands are present
- This coiling blocks the ribosome binding site
- What are the differences in gene expression between prokaryotes and eukaryotes?
- In bacteria, transcription and translation happens at the same time and place (in the cytosol), while in eukaryotes, they’re separated in both time and space.
- In bacteria, most genes are active while only certain ones are repressed, while in eukaryotes, most genes are inactive, and only the ones that are needed are activated.
- Bacteria contain no histones, and have different chromatin structure than eukaryotes.
- What are epigenetics?
- Epigenetics refers to reversible heritable chemical modifications of DNA and histones, which regulate gene expression in eukaryotes
- Describe DNA methylation
- Refers to methylation of cytosine bases in CpG island promoters
- DNA methylation inhibits gene transcription
- Describe histone modification
- Histone acetylation relaxes the DNA coiling, increasing transcription
- Histone methylation tightens the DNA coiling, decreasing transcription
- Describe nucleosome remodeling
- The SWI/SNF protein complex remodels the nucleosome, influencing gene expression
- What is the histone code?
- The histone code refers to the patterns of covalent modification of the histones, of which there are billions of possible combinations.