Gene Regulation in Prokaryotes, Lac Operon, Practice Problems, FAQs

Have you ever thought how disastrous it would be if your eyes started producing the same hydrochloric acid that your stomach produces? Your stomach cells produce HCl to activate the gastric enzymes needed for digestion. But you definitely don’t need HCl in your eyes. But it's surprising that in spite of having the same chromosomes and same DNA sequence in both the stomach cells and the cells of the eye, the genes expressed by these cells vary. This indicates that there must be some sort of a regulation mechanism through which cells regulate which genes to express and when to express them.

For proper functioning of a cell, all necessary proteins must be synthesised at the right time. The information needed to synthesise every protein needed by a cell is encoded in its DNA. But do you think a cell requires all the proteins encoded by its DNA at all times? It definitely does not. Cells control the synthesis of proteins by turning on specific genes at specific times and this is known as gene expression. Be it a multicellular organism like a human or a single-celled bacteria, regulation of gene expression occurs to control which protein is synthesised when, in what quantities and when should its synthesis be stopped. But why is this needed?

Regulation of gene expression is required because it saves energy and space. Expressing every gene all the time can be energy consuming and would also need the entire DNA to be transcribed into mRNA at all times. This would require the entire DNA to be unwound which would take up a lot of space in the cell and the cells would have to be enormous. Expressing only a few genes at a time allows only segments of DNA to be transcribed at a time and this helps to save up a lot of cell space.

Do you think the mechanism for regulation is the same for all cells? Will it be the same for a bacterial cell and a plant cell? No, it won’t because prokaryotes like bacteria and eukaryotes like plants, animals, fungi and protists have very different genetic compositions. In this article we will learn about the regulation of a group of genes, known as the lac operon in prokaryotes.

Table of contents

• Regulation of gene expression
• Gene regulation in prokaryotes
• Lac operon 
• Practice problems
• FAQs

Regulation of Gene Expression

Gene expression is the process by which genetic information stored in the DNA is converted into proteins within the cell. Therefore, we can say that the expression of the gene can be quantified in terms of the amount of protein synthesised by the genes.

There are two types of genes in a particular genome, housekeeping genes and regulated genes. The housekeeping genes or constitutive genes code for proteins that are needed at all times, e.g, enzymes of the respiratory pathway. Thus, these genes are expressed at all times. The regulated genes or non-constitutive genes do not express at all times and their expression is regulated as per the needs of the cell. 

Gene regulation is the mechanism of turning gene expression on or off. Environmental, metabolic and physiological conditions regulate the expression of genes.

We can infer that the gene regulation can take place at various steps of gene expression which includes the following:

• Replication level – Any error in copying the DNA may result in an altered expression.
• Transcriptional level – During transcription, any error in the polymerization may again lead to a change in expression of the gene.
• Post-transcriptional level – During the post-transcriptional modification i.e., RNA splicing, there may be some changes.
• Translational level – During translation, if there is an error in the attachment of mRNA to the tRNA molecules, some changes may arise.

Gene regulation can be of two types - Positive and negative. Positive gene regulation occurs with the help of activators which keep the gene expression on until they need to be suppressed. Negative gene regulation occurs when the expression of a gene is suppressed with the help of repressor proteins and is induced only when needed. An inducer of a negatively regulated gene can be a metabolite, hormone, substrate, etc.

Gene Regulation in Prokaryotes

Prokaryotes are organisms such as bacteria, cyanobacteria and archaebacteria that lack a well defined nucleus. Their genetic material lies naked in the cytoplasm and is known as a nucleoid. The genetic material in most prokaryotes is composed of a single chromosome. 

Gene regulation in prokaryotes is most extensively observed at the initiation of transcription. The regulation of the non-constitutive genes is carried out with the help of regulatory proteins or accessory proteins which affect the ability of RNA polymerase to bind to the transcription initiation site. Both positive and negative gene regulation is observed in prokaryotes. 

From their study in bacterial genetics, Jacob and Monod (1961) proposed that genetic material has a number of functional units known as operons which are made up of a few genes that are regulated and operated simultaneously. The lac operon is a negatively regulated operon in prokaryotes which codes for proteins needed for lactose breakdown. The trp operon is a positively regulated operon which is involved with synthesis of the tryptophan amino acid.

What is an operon?

An operon is a functional unit consisting of a few genes that are transcribed together to form a single mRNA that can code for multiple proteins. All the genes in an operon are regulated simultaneously. It consists of four types of nucleotide sequences - regulator, operator, promoter and structural.

• Regulator - It codes for a regulatory protein which binds to the operator gene to repress its expression.

• Operator - It promotes the functioning of the operon, as long as it is not bound by the repressor gene.

• Promoter region - This is the gene which serves as the transcription initiation point and provides a recognition site for the binding of RNA polymerase. 

• Structural genes - These are the genes that code form the mRNA for the synthesis of the polypeptide..

Lac Operon

lac operon is a group of genes with a single promoter that regulates the transcription of three genes that code for proteins responsible for the transport and metabolism of lactose in prokaryotes. The mRNA coded by the structural genes of this operon is a polycistronic mRNA that is translated to produce three enzymes.

Gene regulation in prokaryotes can be explained with the help of the lac operon model. It is a negatively regulated operon system in which the expression of the structural genes remains repressed until an inducer is present in the medium. The inducer in lac operon is lactose, which is a substrate for one of the enzymes coded by the operon. This operon model was studied and observed by Jacob and Monod. 

Fig: lac operon

The lac operon consists of :

• Promoter of inhibitor or Pi - This is the promoter region for the inhibitor or repressor gene, to which RNA polymerase binds and initiates transcription of mRNA that can be translated into the repressor protein.
• Repressor/Inhibitor gene or lacI gene – This gene produces mRNA that codes for a repressor protein. The repressor is a tetrameric protein having two sites of attachment, one for the operator gene and the other for the inducer. The i gene is expressed constitutively.
• Promoter - The promoter gene possesses two sites, one for cyclic AMP (cAMP) and the other for RNA polymerase. The promoter gene is activated only after cAMP and a cAMP induced catabolite activator protein (CAP) binds to it. It then functions as a recognition site for the RNA polymerase enzyme to initiate transcription.
• Operator - This gene controls the expression of the operon. In the absence of lactose (inducer) in the system, the repressor gene binds with it and prevents the RNA polymerase from passing over from the promoter to the structural genes. Thus, the expression of the operon remains turned off.
• Structural genes - The lac operon consists of three structural genes which transcribe into a polycistronic mRNA which produces three enzymes. These genes are expressed non-constitutively.
• lacZ gene – It codes for beta-galactosidase which catalyses the hydrolysis of lactose into glucose and galactose.
• lacY gene – It codes for a membrane bound enzyme named permease which regulates the entry of lactose into the cell.
• lacA gene – It codes for transacetylase which helps in acetylation reaction by transferring the acetyl group from acetyl coA to galactoside.

Regulation of lac operon

In the absence of lactose

When both glucose and lactose are present in the growing medium, E.coli does not feed on lactose and lactose is not found within the cell. In the absence of the inducer lactose, the RNA polymerase binds to the promoter of the inhibitor gene (lac I) and transcribes repressor mRNA which is translated into active repressor protein. Active repressor protein has affinity for the operator region, so it binds to the operator region. Since the repressor is present at the operator region, RNA polymerase cannot move forward hence falls off. Thus transcription of the structural genes is prevented and their expression remains turned off.

Fig: Regulation of lac operon in absence of lactose

Once bound to the operator region, the repressor does not get fixed over there permanently. Every protein has a fixed life span. So, the repressor protein bound to the operator region is degraded after some time and is replaced by another repressor protein. This is the reason that the repressor protein should be constitutively produced.

In the presence of lactose

If the growth medium has lactose only, then the E. coli feeds on lactose. The small quantity of permease enzyme present in the cell allows lactose to enter the cell in the first place. Even in the presence of lactose, RNA polymerase binds to the promoter of the inhibitory gene and transcribes repressor mRNA which forms the active repressor. But in this scenario, lactose binds to the active repressor and changes its conformation, such that it can no longer bind to the operator gene. The inactive repressor cannot bind to the operator now because it has lost its affinity. Hence the operator region is free and RNA pol can move further to express structural genes. Thus, the polycistronic mRNA is transcribed and the three enzymes , β-galactoside, permease and transacetylase are translated from it. The three enzymes help in further metabolism of lactose. Thus, lac gene expression is switched ON.

Fig: Regulation of lac operon in presence of lactose

Thus, the regulation of the lac operon can be summarised as follows -

Fig: Summary of regulation of lac operon

Practice problems

Q1. In which of the given conditions will the lac operon be expressed?

A. When lactose and glucose concentrations are equal
B. When glucose concentration is higher than lactose concentration
C. As long as lactose concentration is higher than glucose concentration
D. It is turned on all the time

Solution: The lac operon is a negatively regulated operon whose expression is turned off as long as the proteins coded by it are not required by the cell. Lactose acts as an inducer of the system and turns it on. E. coli cells prefer glucose compared to that of lactose and when both are present in equal concentrations or when glucose concentration is higher, the bacterial cells will take up glucose rather than taking up lactose and hence the operon will be turned off. Only when the lactose concentration in the growth medium is higher than that of the glucose concentration, the cells of E. coli use the little amount of permease that is always present in the cell to take up the lactose molecules. These lactose molecules act as inducers and bind to the repressor proteins to disable them from binding to the operator region of the operon. Hence RNA polymerase can easily pass on from the promoter region to the structural genes and transcribe them. This turns on the gene expression.

Thus, the correct option is c.

Fig: Regulation of lac operon in presence of lactose

Q2. Which of the following genes is responsible for the synthesis of the repressor protein?

A. lac Z
B. lac Y
C. lac I
D. lac A

Solution: The repressor protein is synthesised by the lac I gene of the operon. In the absence of the inducer lactose, the RNA polymerase binds to the promoter of the inhibitor gene (lac I) and transcribes repressor mRNA which is translated into active repressor protein. Active repressor protein has affinity for the operator region, so it binds to the operator region. Since the repressor is present at the operator region, RNA polymerase cannot move forward hence falls off. Thus transcription of the structural genes is prevented and their expression remains turned off.

Thus, the correct option is c.

Q3.  Lactose turns on the lac operon by

A. binding to the repressor
B. binding to the promoter gene
C. binding to the operator
D. Binding to the structural genes

Solution: Lactose binds to the active repressor and changes its conformation, such that it can no longer bind to the operator gene. The inactive repressor cannot bind to the operator now because it has lost its affinity. Hence the operator region is free and RNA pol can move further to express structural genes. Thus, the polycistronic mRNA is transcribed and the three enzymes , β-galactoside, permease and transacetylase are translated from it. The three enzymes help in further metabolism of lactose. Thus, lac gene expression is switched ON.

Thus, the correct option is a.

Fig: Regulation of lac operon in presence of lactose

Q4. Which of the following is the correct statement?

A. lac operon expresses itself in the absence of lactose
B. The repressor protein coded by lac I gene binds to the promoter region of the structural genes.
C. The lac operon produces a single enzyme, that is, β-galactosidase when it is turned on
D. The messenger RNA produced by the lac operon is a polycistronic mRNA

Solution:  lac operon is a group of genes with a single promoter that regulates the transcription of three genes that code for proteins responsible for the transport and metabolism of lactose in prokaryotes. The mRNA coded by the structural genes of this operon is a polycistronic mRNA that is translated to produce three enzymes.

Fig: lac operon

The lac operon is a negatively regulated operon whose expression is turned off as long as the proteins coded by it are not required by the cell. Lactose acts as an inducer of the system and it turns on only in the presence of lactose.

In the absence of lactose, the repressor protein binds to the operator region of the operon and prevents the RNA polymerase enzymes from passing over to the structural genes. Hence, their transcription is turned off and the genes are unable to express themselves.

The lac operon consists of three structural genes which transcribe into a polycistronic mRNA which produces three enzymes. These genes are expressed non-constitutively.

• lacZ gene – It codes for beta-galactosidase which catalyses the hydrolysis of lactose into glucose and galactose.
• lacY gene – It codes for a membrane bound enzyme named permease which regulates the entry of lactose into the cell.
• lacA gene – It codes for transacetylase which helps in acetylation reaction by transferring the acetyl group from acetyl coA to galactoside.

Thus, the correct option is d.

FAQs

Q1. Do eukaryotes have operons?
Answer: An operon is a set of genes transcribed from the same promoter to give a polycistronic mRNA. Eukaryotes, however, translate only the first coding sequence on an mRNA which is why they cannot use a polycistronic mRNA to translate multiple genes. Thus, for a long time it was assumed that eukaryotic genomes do not contain operons.

However, proper operons are rarely found in a few eukaryotic organisms. Most organisms have a few operons; however some eukaryotes possess high numbers of operons. For example the worm, Caenorhabditis elegans, has around 2600 genes of roughly 20,000 genes arranged into 1000 operons.

Q2. What is the significance of lac operon to humans?
Answer: The lac operon of the E. coli cells that are present in our gut as commensals helps in digestion of lactose when we consume dairy products.

Q3. How long is the lac operon?
Answer: The lac operon of E. coli is around 5300 base pairs long.

Q4. Does the lac operon always remain turned off in the absence of lactose?
Answer: Sometimes RNA polymerase sneaks in and crosses the operator region before the new repressor binds to the operator, thus, allowing transcription to occur. But this happens very rarely and due to this, low level of transcription and subsequent translation of transcribed mRNA keeps occurring even in the presence of a repressor. This helps in maintaining the basal level of proteins in the cell at any given time. 

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