Transcription factors: definition of a concept, characteristic

In all organisms (with the exception of some viruses), the realization of genetic material occurs through the DNA-RNA-protein system. At the first stage, information is transcribed (transcribed) from one nucleic acid to another. The proteins that regulate this process are called transcription factors.

What is transcription?

Transcription is the biosynthesis of an RNA molecule based on a DNA template. This is possible due to the complementarity of certain nitrogenous bases that make up nucleic acids. The synthesis is carried out by specialized enzymes - RNA polymerases and is controlled by many regulatory proteins.

The entire genome is not transcribed immediately, but only a certain part of it, called a transcripton. The latter includes a promoter (the site of attachment of RNA polymerase) and a terminator (sequence that activates the completion of the synthesis).

A transkrypton prokaryote is an operon consisting of several structural genes (cistrons). On its basis, polycistronic RNA is synthesized containing information about the amino acid sequence of a group of functionally related proteins. Only one gene is included in the transcripton of eukaryotes.

The biological role of the transcription process is the formation of RNA template sequences, on the basis of which protein synthesis (translation) is carried out in ribosomes.

RNA synthesis in prokaryotes and eukaryotes

The RNA synthesis scheme is the same for all organisms and includes 3 stages:

• Initiation - addition of a polymerase to a promoter, activation of a process.
• Elongation - extension of the nucleotide chain in the direction from 3´ to 5´ end with the closure of phosphodiester bonds between nitrogenous bases, which are selected complementary to DNA monomers.
• Termination - completion of the synthesis process.

In prokaryotes, all types of RNA are transcribed by one RNA polymerase, consisting of five protomers (β, β ′, ω and two α subunits), which together form a core enzyme capable of increasing the chain of ribonucleotides. There is an additional unit σ, without which it is impossible to attach the polymerase to the promoter. The complex of core and sigma factor is called holoenzyme.

Despite the fact that the σ subunit is not always associated with the cortex, it is considered to be part of RNA polymerase. In a dissociated state, sigma is not able to bind to the promoter, only as part of a holoenzyme. After initiation is complete, this protomer is separated from the core, being replaced by an elongation factor.

A feature of prokaryotes is a combination of translation and transcription processes. Ribosomes immediately join the RNA that began to be synthesized and build the amino acid chain. Transcription is stopped due to the formation of a hairpin structure in the region of the terminator. At this stage, the DNA polymerase-RNA complex breaks down.

In eukaryotic cells, transcription is carried out by three enzymes:

• RNA polymerase l - synthesizes 28S and 18S ribosomal RNAs.
• RNA polymerase ll - transcribes genes encoding proteins and small nuclear RNAs.
• RNA polymerase lll - responsible for the synthesis of tRNA and 5S rRNA (a small subunit of ribosomes).

None of these enzymes is able to initiate transcription without the participation of specific proteins that provide interaction with the promoter. The essence of the process is the same as that of prokaryotes, but each stage is much more complicated with the participation of a greater number of functional and regulatory elements, including chromatin-modifying ones. Only at the initiation stage, about a hundred proteins are involved, including a number of transcription factors, while in bacteria, only one sigma subunit is sufficient for communication with the promoter and sometimes the help of an activator is needed.

The most important contribution of the biological role of transcription in the biosynthesis of various types of proteins determines the need for a strict gene reading control system.

Transcription regulation

In no cell is genetic material fully realized: only part of the genes is transcribed, and the rest are inactive. This is possible due to complex regulatory mechanisms that determine from which sections of DNA and in what quantity RNA sequences will be synthesized.

In unicellular organisms, the differential activity of genes is adaptive, while in multicellular organisms it also determines the processes of embryogenesis and ontogenesis, when different types of tissues are formed on the basis of one genome.

Gene expression is monitored at several levels. The most important step is the regulation of transcription. The biological meaning of this mechanism is to maintain the required amount of various proteins required by the cell or organism at a particular moment of existence.

There is an adjustment of biosynthesis at other levels, such as processing, translation, and transport of RNA from the nucleus to the cytoplasm (the latter is absent in prokaryotes). With positive regulation, these systems are responsible for the production of the protein based on the activated gene, which is the biological meaning of transcription. However, at any stage, the chain may be suspended. Some regulatory features in eukaryotes (alternative promoters, splicing, modification of polyadenylation sites) lead to the appearance of various variants of protein molecules based on a single DNA sequence.

Since RNA formation is the first step in deciphering genetic information on the path to protein biosynthesis , the biological role of the transcription process in modifying the cellular phenotype is much more significant than the regulation of processing or translation.

The determination of the activity of specific genes in both prokaryotes and eukaryotes occurs at the initiation stage using specific switches, which include regulatory regions of DNA and transcription factors (TF). The operation of such switches is not autonomous, but is under the strict control of other cellular systems. There are also mechanisms for nonspecific regulation of RNA synthesis, ensuring the normal passage of initiation, elongation and termination.

The concept of transcription factors

Unlike regulatory elements of the genome, transcription factors are proteins by chemical nature. By binding to certain sections of DNA, they can activate, inhibit, accelerate or slow down the transcription process.

Depending on the effect produced, the transcription factors of prokaryotes and eukaryotes can be divided into two groups: activators (initiate or increase the intensity of RNA synthesis) and repressors (inhibit or inhibit the process). Currently, over 2000 TFs have been detected in various organisms in total.

Regulation of transcription in prokaryotes

In prokaryotes, the control of RNA synthesis occurs mainly at the initiation stage due to the interaction of TF with a specific region of the transcripton - an operator that is located next to the promoter (sometimes intersecting with it) and, in fact, is a landing site for a regulatory protein (activator or repressor). For bacteria, another way of differential control of genes is characteristic - the synthesis of alternative σ-subunits intended for different groups of promoters.

Partially, the expression of the operon can be regulated at the stages of elongation and termination, but not due to TF binding to DNA, but due to proteins interacting with RNA polymerase. These include Gre-proteins and antiterminative factors Nus and RfaH.

Elongation and termination of transcription in prokaryotes are influenced in a certain way by protein synthesis occurring in parallel. In eukaryotes, both these processes themselves and the transcription and translation factors are spatially separated, which means that they are not functionally related.

Activators and repressors

Prokaryotes have two mechanisms of transcription regulation at the initiation stage:

• positive - carried out by activator proteins;
• negative - controlled by repressors.

With positive regulation, the addition of the factor to the operator activates the gene, and with negative regulation, on the contrary, it turns it off. The ability of a regulatory protein to bind to DNA depends on the attachment of the ligand. The role of the latter is usually played by low molecular weight cellular metabolites, which in this case play the role of coactivators and corepressors.

The mechanism of action of the repressor is based on overlapping areas of the promoter and operator. In operons with this structure, the attachment of the protein factor to DNA covers part of the landing site for RNA polymerase, preventing the latter from initiating transcription.

Activators work on weak promoters with low functionality that are poorly recognized by RNA polymerases or difficult to melt (separation of the DNA helix chains necessary for transcription to begin). Joining the operator, the protein factor interacts with the polymerase, significantly increasing the likelihood of initiation. Activators are able to increase the transcription intensity by 1000 times.

Some TF prokaryotes can act both as activators and as repressors, depending on the location of the operator in relation to the promoter: if these regions overlap, the factor inhibits transcription, and otherwise triggers.

Negative regulation can be considered by the example of the tryptophan operon of the bacterium E. coli, which is characterized by the location of the operator within the promoter sequence. The repressor protein is activated by the addition of two tryptophan molecules that change the angle of inclination of the DNA-binding domain so that it can enter the large groove of the double helix. At low tryptophan concentrations, the repressor loses the ligand and becomes inactive again. In other words, the frequency of transcription initiation is inversely proportional to the amount of metabolite.

Some bacterium operons (for example, lactose) combine positive and negative regulatory mechanisms. Such a system is necessary when a single signal for rational control of expression is not enough. So, the lactose operon encodes enzymes that transport into the cells and then cleave lactose - an alternative energy source that is less beneficial than glucose. Therefore, only at a low concentration of the latter does the CAP protein bind to DNA and initiate transcription. However, this is advisable only in the presence of lactose, the absence of which leads to the activation of a Lac repressor that blocks the access of the polymerase to the promoter even in the presence of a functional form of the activator protein.

Due to the operon structure in bacteria, several genes are controlled by one regulatory region and 1-2 TFs, while in eukaryotes, a single gene has a large number of regulatory elements, each of which is dependent on many other factors. Such complexity corresponds to a high level of organization of eukaryotes, and especially to multicellular organisms.

Regulation of mRNA synthesis in eukaryotes

The control of the expression of eukaryotic genes is determined by the combined action of two elements: protein transcription facts (TF) and regulatory DNA sequences that can be located near the promoter, much higher than it, in the introns or after the gene (this refers to the coding region, and not the gene in full value )

Some sections act as switches, while others do not interact directly with TF, but give the DNA molecule the flexibility necessary to form a loop-like structure that accompanies the process of transcriptional activation. Such sections are called spacers. All regulatory sequences, together with the promoter, constitute the control region of the gene (gene conrol region).

It is worth noting that the effect of transcription factors themselves is only part of a complex multilevel regulation of genetic expression, in which a huge number of elements are added to the resulting vector, which determines whether RNA will be synthesized from a particular part of the genome.

An additional factor in the control of transcription in a nuclear cell is a change in chromatin structure. Here, there is both total regulation (provided by the distribution of heterochromatin and euchromatin sites) and local, associated with a specific gene. For polymerase to work, all levels of DNA compaction, including nucleosomal, must be eliminated.

A variety of transcription factors in eukaryotes is associated with a large number of regulators, which include amplifiers, silencers (enhancers and silencers), as well as adapter elements and insulators. These sites can be located either close to or at a considerable distance from the gene (up to 50 thousand bp).

Enhancers, silencers and adapter elements

Enhancers are short, sequential DNAs capable of triggering transcription when interacting with a regulatory protein. The approach of the amplifier to the promoter region of the gene is due to the formation of a loop-like structure of DNA. The binding of the activator to the enhancer either stimulates the assembly of the initiation complex or helps the polymerase to proceed to elongation.

An enhancer has a complex structure and consists of several module sites, each of which has its own regulatory protein.

Silencers are sections of DNA that inhibit or completely eliminate the possibility of transcription. The mechanism of operation of such a switch is still unknown. One of the proposed methods is the occupation of large areas of DNA with special proteins of the SIR group, which block access to initiation factors. In this case, all genes located within a few thousand nucleotide pairs from the silencer are turned off.

The adapter elements in combination with TFs that bind to them form a separate class of genetic switches that selectively respond to steroid hormones, cyclic AMP, and glucocorticoids. This regulatory unit is responsible for the reaction of the cell to heat shock, the effects of metals and some chemical compounds.

Among the control sections of DNA, another type of element is isolated - insulators. These are specific sequences that inhibit the influence of transcription factors on distant genes. The mechanism of action of insulators has not yet been clarified.

Eukaryotic transcription factors

If bacteria have transcription factors only have a regulatory function, then in nuclear cells there is a whole group of TFs providing background initiation, but at the same time directly dependent on regulatory proteins binding to DNA. The number and variety of the latter in eukaryotes is huge. So, in the human body, the proportion of sequences encoding protein transcription factors is about 10% of the genome.

At present, eukaryotic TFs have not been sufficiently studied, as are the mechanisms of genetic switches, the arrangement of which is much more complicated than the models of positive and negative regulation in bacteria. Unlike the latter, the activity of nuclear cell transcription factors is influenced not by one or two, but by tens and even hundreds of signals that can mutually amplify, weaken or exclude each other.

On the one hand, the activation of a specific gene requires a whole group of transcription factors, but on the other hand, one regulatory protein may be enough to trigger the expression of several genes by the cascade mechanism. This whole system is a sophisticated computer that processes signals from different sources (both external and internal) and adds their effects to the final result with a plus or minus sign.

Regulatory transcription factors in eukaryotes (activators and repressors) do not interact with the operator, like bacteria, but with DNA scattered in control regions and affect initiation through intermediaries, which can be mediator proteins, initiation complex factors, and enzymes that change chromatin structure .

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Operating principle

The functioning of basal factors is a cascade assembly of various subunits with the formation of an initiation complex and transcription activation. In fact, this process is the final stage of exposure to the activator protein.

Specific factors can regulate transcription in two stages:

• assembly of the initiation complex;
• transition to productive elongation.

In the first case, the work of specific TFs is reduced to the primary chromatin rearrangement, as well as the involvement, orientation, and modification of the mediator, polymerase, and basal factors on the promoter, which leads to transcription activation. The main element of signal transmission is the mediator - a complex of 24 subunits, acting as an intermediary between the regulatory protein and RNA polymerase. The sequence of interactions is individual for each gene and its corresponding factor.

Elongation is regulated by the interaction of the factor with the P-Tef-b protein, which helps RNA polymerase overcome the pause associated with the promoter.

Functional structures of TF

Transcription factors have a modular structure and perform their work due to three functional domains:

1. DNA-binding (DBD) - needed for recognition and interaction with the regulatory region of the gene.
2. Trans-activating (TAD) - allows you to interact with other regulatory proteins, including transcription factors.
3. Signal Recognition (SSD) - necessary for the perception and transmission of regulatory signals.

In turn, the DNA-binding domain has many types. The main motives in its structure include:

• "zinc fingers";
• homeodomain;
• "β" layers;
• loops;
• leucine zipper;
• spiral loop spiral;
• spiral-turn-spiral.

Due to this domain, the transcription factor “reads” the nucleotide sequence of DNA in the form of a pattern on the surface of a double helix. Due to this, specific recognition of certain regulatory elements is possible.

The interaction of motifs with the DNA helix is ​​based on the exact correspondence between the surfaces of these molecules.

Regulation and synthesis of TF

There are several ways to regulate the effect of transcription factors on transcription. These include:

• activation - a change in the functionality of a factor with respect to DNA due to phosphorylation, ligand attachment, or interaction with other regulatory proteins (including TF);
• translocation - transportation of a factor from the cytoplasm to the nucleus;
• accessibility of the binding site - depends on the degree of chromatin condensation (in the state of heterochromatin, DNA is not available for TF);
• a complex of mechanisms characteristic of other proteins (regulation of all processes from transcription to post-translational modification and intracellular localization).

The latter method determines the quantitative and qualitative composition of transcription factors in each cell. Some TFs are able to regulate their synthesis according to the type of classical feedback when its own product becomes an inhibitor of the reaction. In this case, a certain concentration of the factor stops the transcription of the gene encoding it.

Common Transcription Factors

These factors are necessary for the start of transcription of any genes and are designated in the nomenclature as TFl, TFll and TFlll depending on the type of RNA polymerase with which they interact. Each factor consists of several subunits.

Basal TFs have three main functions:

• the correct location of RNA polymerase on the promoter;
• unwinding DNA strands at the start of transcription;
• release of the polymerase from the promoter at the time of transition to elongation;

Certain subunits of basal transcription factors bind to regulatory elements of the promoter. The most important is the TATA box (not typical for all genes), located at a distance of "-35" nucleotides from the point of initiation. Other binding sites include the INR, BRE, and DPE sequences. Some TFs do not directly contact DNA.

The group of major transcription factors for RNA polymerase ll includes TFllD, TFllB, TFllF, TFllE, and TFllH. The Latin letter at the end of the designation indicates the sequence of detection of these proteins. So, the TFlllA factor related to lll RNA polymerase was the first to be isolated.

The assembly of basal TF occurs only with the assistance of an activator, mediator and chromatin-modifying proteins.

Specific TF

Through the control of genetic expression, these transcription factors regulate the biosynthetic processes of both individual cells and the whole organism from embryogenesis to fine phenotypic adaptation to changing environmental conditions. The scope of TF includes 3 main blocks:

• development (embryo and ontogenesis);
• cell cycle;
• response to external signals.

A special group of transcription factors regulates the morphological differentiation of the embryo. This protein set is encoded by a special consensus sequence of 180 pairs of nucleotides in length called homeobox.

In order to determine which gene should be transcribed, the regulatory protein must “find” and bind to a specific region of DNA that acts as a genetic switch (enhancer, silencer, etc.). Each such sequence corresponds to one or several related transcription factors that recognize the desired site due to the coincidence of the chemical conformations of a particular external segment of the helix and the DNA-binding domain (key-lock principle). For recognition, a portion of the primary DNA structure, called the large groove, is used.

After binding to DNA, the actions of the activator protein trigger a series of sequential steps leading to the assembly of the pre-initiator complex. A generalized diagram of this process is as follows:

1. The binding of the activator to chromatin in the promoter region, the involvement of ATP-dependent rearrangement complexes.
2. Chromatin rearrangement, activation of histone-modifying proteins.
3. Covalent modification of histones, the involvement of other activator proteins.
4. The binding of additional activating proteins to the regulatory region of the gene.
5. Attraction of a mediator and common TF.
6. Assembly of the pre-initiator complex on the promoter.
7. Exposure to other activator proteins, rearrangement of subunits of the preinitiator complex.
8. The beginning of transcription.

The order of these events from gene to gene may vary.

Such a large number of activation mechanisms corresponds to an equally wide range of methods of repression. That is, by inhibiting one of the steps towards initiation, a regulatory protein can reduce its effectiveness or completely block it. Most often, the repressor uses several mechanisms at once, guaranteeing the absence of transcription.

Coordinated Gene Control

Despite the fact that each transcripton has its own regulatory system, eukaryotes have a mechanism that allows bacteria to start or stop groups of genes aimed at a specific task, like bacteria. This is achieved with the help of a transcription determining factor that completes the combinations of other regulatory elements necessary for maximum activation or suppression of the gene.

In transcriptons subject to such regulation, the interaction of different components leads to the same protein, which plays the role of the resulting vector. Therefore, the activation of such a factor affects several genes at once. The system works on the principle of a cascade.

The coordinated control scheme can be considered by the example of ontogenetic differentiation of skeletal muscle cells, the precursors of which are myoblasts.

Transcription of genes encoding the synthesis of proteins characteristic of a mature muscle cell is triggered by any of four myogenic factors: MyoD, Myf5, MyoG and Mrf4. These proteins activate the synthesis of themselves and each other, and also include the genes of the additional transcription factor Mef2 and structural muscle proteins. Mef2 is involved in the regulation of further differentiation of myoblasts, while maintaining the concentration of myogenic proteins by the mechanism of positive feedback.