Prokaryotic Gene Expression

Prokaryotes are vital to our ecosystem (Fig. 1). They are found in the soil and air and contribute to both the carbon and nitrogen cycle. Prokaryotes are also found in our digestive systems as part of our microbiome and help to maintain our health. In fact, there are as many bacteria in our bodies as the number of cells that we have which suggests the importance of prokaryotes to our overall health. Importantly, bacteria must also adapt to their environment. Bacteria in the large intestine face different conditions than bacteria on the skin. Therefore, it is crucial that bacteria have tight gene regulation to adapt to their local environment.

Control of gene expression in prokaryotes

As a quick refresher, prokaryotes, which include Bacteria and Archaea, make up two out of three domains of life (with the last being Eukarya). Prokaryotes are single-celled organisms with no nuclei or organelles (Fig. 2). In addition, prokaryotes have circular chromosomes compared to linear chromosomes in eukaryotes. Since there are no boundaries set by organelles, this means that gene expression and its regulation in prokaryotes occur in the cytoplasm.

Gene expression is critical for adapting to environmental changes. Just like eukaryotes, prokaryotes must also adapt to environmental changes such as food availability, predation, temperature, salinity, and pH. This requires tight control of gene expression. Due to their circular chromosomes, prokaryotes can transcribe multiple genes into a single messenger RNA (mRNA) molecule. The group of genes that are synthesized into a single mRNA are called operons. An example of an operon is the tryptophan (trp) operon which is composed of five genes that are all regulated by a single promoter. The five genes in the operon are all related to tryptophan production and the gene products will work together to increase tryptophan synthesis.

Mechanism of gene expression in prokaryotes

To understand the mechanisms of gene expression in prokaryotes, it is important to describe all of the components of the prokaryotic transcription machinery and within the operon.

The promoter is the region of DNA adjacent to the genes that control the activity of transcription. Often there is a region of DNA called the operator that lies between the promoter and the genes. For both the promoter and operator, different proteins can bind to either region to activate or repress gene expression respectively (Fig. 3).

In general, the proteins that bind to the promoter are all called activators while proteins that bind to the operator are called repressors. Together, these two proteins are examples of a class of proteins called transcription factors that bind to regions of DNA and control its transcription. If we recall the mechanisms of transcription, RNA polymerase binds to the promoter to initiate transcription. However, if a repressor is bound to the operator, it will block the path of RNA polymerase thereby preventing transcription (Fig. 4).

Importantly, repressor and activator proteins are often made by the cells themselves so that the cell can regulate its own transcription depending on the current needs of the cell. For example, if there is an abundance of tryptophan present in the environment, the protein may overexpress a repressor which will then bind to the operator to prevent additional tryptophan from being made. This is useful so that energy is not wasted synthesizing molecules that are already abundant in the environment. In this case, tryptophan is an inducer that can interact with either the activator or repressor to control transcription.

In contrast, some genes always need to be expressed for the cell to survive. For example, proteins involved in DNA repair, transcription, translation, and metabolism need to be constantly available, otherwise, the cell will die. The genes that encode these proteins are called housekeeping genes which are not regulated by activators or repressors.

Regulation of gene expression in prokaryotes

Regulation of gene expression in prokaryotes can be classified as positive regulation or negative regulation. Generally, during negative regulation, there is competition between RNA polymerase and the repressor that prevents RNA polymerase from transcribing prokaryotic genes. In contrast, during positive regulation, the activator recruits RNA polymerase to the promoter region to begin transcription.

There are also other regulatory elements located away from the operon, either upstream or downstream, that can influence prokaryotic gene expression. How can some regulatory elements be so far away and still influence transcription? The answer is through DNA looping. Just like eukaryotes, prokaryotic DNA twists and curls as a way to conserve space and fit inside a cell. This looping allows regulatory elements that are located far away from the operon to still be able to interact with RNA polymerase (Fig. 5).

Altering patterns of gene expression in prokaryotes

In this section, we will dive into two examples of operons to solidify our understanding of prokaryotic gene regulation and further examine how regulation can change depending on the external environment. These two examples will be the tryptophan (trp) operon and the lactose (lac) operon.

Tryptophan (trp) operon

Tryptophan is one of the twenty amino acids that are used by many bacteria to build proteins. However, bacteria are very efficient and do not like to waste nutrients. When there is an abundance of tryptophan in the environment, bacteria will turn off the trp operon and instead use environmental tryptophan to build proteins. In contrast, when there is little tryptophan in the environment, bacteria will turn on the trp operon to encode enzymes that synthesize tryptophan. The altering pattern of gene expression of the tryptophan operon occurs within the promoter and operator region.

The trp operon is composed of a promoter region, an operator region, and five genes that encode enzymes that are responsible for tryptophan synthesis. At any given moment, there are two possible states of tryptophan regulation

1. The promoter is bound by the RNA polymerase and trp gene expression is turned on.

2. The operator is bound by repressor proteins and trp gene expression is turned off.

In the first state of the trp operon, when environmental tryptophan concentration is low, RNA polymerase binds to the promoter region which can then initiate transcription of the five trp genes into a single mRNA. This mRNA can be translated into proteins that will help synthesize tryptophan.

In contrast, when environmental tryptophan concentration is high, the cell will uptake tryptophan from the environment. Two tryptophans will then bind to the repressor causing the repressor to undergo a conformational change that allows the repressor protein to bind to the operator region. Due to the proximity of the promoter and the operator region with each other, when the repressor protein binds to the operator region, it will block RNA polymerase from binding to the promoter. Therefore, transcription of the five trp genes will not be possible.

The key point is that environmental tryptophan regulates the expression of the five trp genes. When environmental tryptophan is low, tryptophan will not be available to bind to the repressor. In contrast, when environmental tryptophan is high, the repressor will become activated to prevent transcription.

• TRYPTOPHAN AVAILABLE → BINDS TO REPRESSOR → REPRESSOR ON → GENES OFF → TRP NOT MADE
• TRYPTOPHAN NOT AVAILABLE → REPRESSOR NOT BOUND → REPRESSOR OFF → GENES ON → TRP MADE

Lactose (lac) operon

Similar to the trp operon, the lac operon is composed of three genes that are transcribed into a single mRNA. The lac operon is important for transporting lactose into the cell and breaking it down for energy. Importantly, lactose is only used as an energy source when glucose is not available. Therefore, only when glucose is low in the environment and lactose is high, will the lac operon be turned on.

The lactose repressor typically binds to the operator region when lactose is low and is only released when lactose is present. When the lactose repressor is bound to the operator, it blocks RNA polymerase and the operon is turned off to prevent transcription of the three lac genes. However, if lactose is high, small amounts of lactose will be broken down into allolactose in the cell. Allolactose will bind to the lactose repressor to induce a conformational change that will cause the lac repressor to be released from the operator. Once released, RNA polymerase can begin transcription of the lac operon.