In this section, you will explore the following question:
- What are operons and what are the roles of activators, inducers, and repressors in regulating operons and gene expression?
Connection for AP® Courses
The regulation of gene expression in prokaryotic cells occurs at the transcriptional level. Simply stated, if a cell does not transcribe the DNA’s message into mRNA, translation (protein synthesis), does not occur. Bacterial genes are often organized into common pathways or processes called operons for more coordinated regulation of expression. For example, in E. coli, genes responsible for lactose metabolism are located together on the bacterial chromosome. (The operon model includes several components, so when studying how the operon works, it is helpful to refer to a diagram of the model. See Figure 16.3 and Figure 16.4.) The operon includes a regulatory gene that codes for a repressor protein that binds to the operator, which prevents RNA polymerase from transcribing the gene(s) of interest. An example of this is seen in the structural genes for lactose metabolism. However, if the repressor is inactivated, RNA polymerase binds to the promoter, and transcription of the structural genes occurs.
There are three ways to control the transcription of an operon: inducible control, repressible control, and activator control. The lac operon is an example of inducible control because the presence of lactose “turns on” transcription of the genes for its own metabolism. The trp operon is an example of repressible control because it uses proteins bound to the operator sequence to physically prevent the binding of RNA polymerase. If tryptophan is not needed by the cell, the genes necessary to produce it are turned off. Activator control (typified by the action of Catabolite Activator Protein) increases the binding ability of RNA polymerase to the promoter. Certain genes are continually expressed via this regulatory mechanism.
Information presented and the examples highlighted in the section support concepts outlined in Big Idea 3 of the AP® Biology Curriculum Framework. The learning objectives listed in the Curriculum Framework provide a transparent foundation for the AP® Biology course, an inquiry-based laboratory experience, instructional activities, and AP® exam questions. A learning objective merges required content with one or more of the seven science practices.
|Big Idea 3||Living systems store, retrieve, transmit and respond to information essential to life processes.|
|Enduring Understanding 3.B||Expression of genetic information involves cellular and molecular mechanisms.|
|Essential Knowledge||3.B.1 Gene regulation results in differential gene expression, leading to cell specialization|
|Science Practice||1.4 The student can use representations and models to analyze situations or solve problems qualitatively and quantitatively|
|Learning Objective||3.21 The student can use representations to describe how gene regulation influences cell products and function.|
|Essential Knowledge||3.B.2 A variety of intercellular and intracellular signal transmissions mediate gene expression.|
|Science Practice||1.4 The student can use representations and models to analyze situations or solve problems qualitatively and quantitatively.|
|Learning Objective||3.23 The student can use representations to describe mechanisms of the regulation of gene expression.|
When discussing the operons with students, challenge them to think about what would happen if there were a gene mutation that disrupted the function of one of the proteins that controls transcription of the operon. For example, if the repressor protein in the lac operon has a mutation that prevents it from binding to lactose, then the repressor will remain bound to the operator and will prevent transcription of the operon even in the presence of lactose. This video describes two other examples of mutations in the lac operon.
Introduce the regulation of transcription in the lac operon using visuals such as this video.
The DNA of prokaryotes is organized into a circular chromosome supercoiled in the nucleoid region of the cell cytoplasm. Proteins that are needed for a specific function, or that are involved in the same biochemical pathway, are encoded together in blocks called operons. For example, all of the genes needed to use lactose as an energy source are coded next to each other in the lactose (or lac) operon.
In prokaryotic cells, there are three types of regulatory molecules that can affect the expression of operons: repressors, activators, and inducers. Repressors are proteins that suppress transcription of a gene in response to an external stimulus, whereas activators are proteins that increase the transcription of a gene in response to an external stimulus. Finally, inducers are small molecules that either activate or repress transcription depending on the needs of the cell and the availability of substrate.
The trp Operon: A Repressor Operon
Bacteria such as E. coli need amino acids to survive. Tryptophan is one such amino acid that E. coli can ingest from the environment. E. coli can also synthesize tryptophan using enzymes that are encoded by five genes. These five genes are next to each other in what is called the tryptophan (trp) operon (Figure 16.3). If tryptophan is present in the environment, then E. coli does not need to synthesize it and the switch controlling the activation of the genes in the trp operon is switched off. However, when tryptophan availability is low, the switch controlling the operon is turned on, transcription is initiated, the genes are expressed, and tryptophan is synthesized.
A DNA sequence that codes for proteins is referred to as the coding region. The five coding regions for the tryptophan biosynthesis enzymes are arranged sequentially on the chromosome in the operon. Just before the coding region is the transcriptional start site. This is the region of DNA to which RNA polymerase binds to initiate transcription. The promoter sequence is upstream of the transcriptional start site; each operon has a sequence within or near the promoter to which proteins (activators or repressors) can bind and regulate transcription.
A DNA sequence called the operator sequence is encoded between the promoter region and the first trp coding gene. This operator contains the DNA code to which the repressor protein can bind. When tryptophan is present in the cell, two tryptophan molecules bind to the trp repressor, which changes shape to bind to the trp operator. Binding of the tryptophan–repressor complex at the operator physically prevents the RNA polymerase from binding, and transcribing the downstream genes.
When tryptophan is not present in the cell, the repressor by itself does not bind to the operator; therefore, the operon is active and tryptophan is synthesized. Because the repressor protein actively binds to the operator to keep the genes turned off, the trp operon is negatively regulated and the proteins that bind to the operator to silence trp expression are negative regulators.
Watch this video to learn more about the trp operon.
The cell would not be able to break down tryptophan.
The cell will gradually produce more tryptophan over time.
The cell would not be able to make tryptophan.
The cell would make tryptophan when it was not needed.
Catabolite Activator Protein (CAP): An Activator Regulator
Just as the trp operon is negatively regulated by tryptophan molecules, there are proteins that bind to the operator sequences that act as a positive regulator to turn genes on and activate them. For example, when glucose is scarce, E. coli bacteria can turn to other sugar sources for fuel. To do this, new genes to process these alternate genes must be transcribed. When glucose levels drop, cyclic AMP (cAMP) begins to accumulate in the cell. The cAMP molecule is a signaling molecule that is involved in glucose and energy metabolism in E. coli. When glucose levels decline in the cell, accumulating cAMP binds to the positive regulator catabolite activator protein (CAP), a protein that binds to the promoters of operons that control the processing of alternative sugars. When cAMP binds to CAP, the complex binds to the promoter region of the genes that are needed to use the alternate sugar sources (Figure 16.4). In these operons, a CAP binding site is located upstream of the RNA polymerase binding site in the promoter. This increases the binding ability of RNA polymerase to the promoter region and the transcription of the genes.
The lac Operon: An Inducer Operon
The third type of gene regulation in prokaryotic cells occurs through inducible operons, which have proteins that bind to activate or repress transcription depending on the local environment and the needs of the cell. The lac operon is a typical inducible operon. As mentioned previously, E. coli is able to use other sugars as energy sources when glucose concentrations are low. To do so, the cAMP–CAP protein complex serves as a positive regulator to induce transcription. One such sugar source is lactose. The lac operon encodes the genes necessary to acquire and process the lactose from the local environment. CAP binds to the operator sequence upstream of the promoter that initiates transcription of the lac operon. However, for the lac operon to be activated, two conditions must be met. First, the level of glucose must be very low or non-existent. Second, lactose must be present. Only when glucose is absent and lactose is present will the lac operon be transcribed (Figure 16.5). This makes sense for the cell, because it would be energetically wasteful to create the proteins to process lactose if glucose was plentiful or lactose was not available.
The trp operon is inducible and is positively regulated. Therefore it is ON by default whereas the lac operon is repressible and is OFF by default.
The trp operon synthesizes tryptophan which is essential for the cell and therefore remains ON, whereas the lac operon synthesizes enzymes for the breakdown of a sugar that is not always available and remains OFF by default.
The trp operon is constitutive and remains ON by default, whereas the lac operon is repressible and therefore is OFF by default.
The lac operon undergoes transcriptional attenuation and therefore is OFF by default, whereas the trp operon is not regulated by any such mechanism and is ON by default.
If glucose is absent, then CAP can bind to the operator sequence to activate transcription. If lactose is absent, then the repressor binds to the operator to prevent transcription. If either of these requirements is met, then transcription remains off. Only when both conditions are satisfied is the lac operon transcribed (Table 16.2).
|Glucose||CAP binds||Lactose||Repressor binds||Transcription|
Watch an animated tutorial about the workings of lac operon here.
The cell would make more lactose.
There would be no lactose outside of the cell.
The cell would not be able to process tryptophan.
The cell would not be able to process lactose.
Modeling the Operon. Use construction paper or more elaborate materials, such as Styrofoam noodles, electrical tape, and Velcro tabs, to create a model of the lac and trp operons that include a regulator, promoter, operator, and structural genes. Then use the model to show how the presence of substrate, e.g., allolactose or tryptophan, can change the activity of the operons. As an extension of the activity, use the model to make predictions about the effects of mutations in any of the regions on gene expression.
In E. coli, the trp operon is on by default, while the lac operon is off by default. Why do you think this is the case?
The activity is an application of Learning Objectives 3.21 and 3.23 and Science Practice 1.4 because students are using a representation to describe how operons regulate gene expression in prokaryotes. In addition, students are applying Science Practice 6.4 because they will use the model to make predictions about gene regulation and expression.
The question is an application of Learning Objectives 3.2 and 3.23 and Science Practice 1.4 because students are using the operon model of gene regulation in prokaryotes to describe an observed phenomenon.
Tryptophan is an amino acid necessary for making proteins, so the cell always needs to have some on hand. However, if plenty of tryptophan is present, it is wasteful to make more, and the expression of the trp genes is repressed. Lactose, a sugar found in milk, is not always available. Cells need not make the enzymes necessary to digest an energy source that is not available, so the lac operon is only turned on when lactose is present.