Bacterial transcription

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Transcription is the process of copying DNA into RNA, usually mRNA. Process of transcription (13080846733).jpg
Transcription is the process of copying DNA into RNA, usually mRNA.

Bacterial transcription is the process in which a segment of bacterial DNA is copied into a newly synthesized strand of messenger RNA (mRNA) with use of the enzyme RNA polymerase.

Contents

The process occurs in three main steps: initiation, elongation, and termination; and the end result is a strand of mRNA that is complementary to a single strand of DNA. Generally, the transcribed region accounts for more than one gene. [1] In fact, many prokaryotic genes occur in operons, which are a series of genes that work together to code for the same protein or gene product and are controlled by a single promoter. [2] Bacterial RNA polymerase is made up of four subunits and when a fifth subunit attaches, called the sigma factor (σ-factor), the polymerase can recognize specific binding sequences in the DNA, called promoters. [3] The binding of the σ-factor to the promoter is the first step in initiation. Once the σ-factor releases from the polymerase, elongation proceeds. [4] The polymerase continues down the double stranded DNA, unwinding it and synthesizing the new mRNA strand until it reaches a termination site. There are two termination mechanisms that are discussed in further detail below. Termination is required at specific sites for proper gene expression to occur. [5] Gene expression determines how much gene product, such as protein, is made by the gene. [2] Transcription is carried out by RNA polymerase but its specificity is controlled by sequence-specific DNA binding proteins called transcription factors. Transcription factors work to recognize specific DNA sequences and based on the cells needs, promote or inhibit additional transcription. [6] Similar to other taxa, bacteria experience bursts of transcription. [7] : 125  [8] [9] [10] [11] [12] [13] The work of the Jones team in Jones et al 2014 explains some of the underlying causes of bursts and other variability, including stability of the resulting mRNA, [7] : 125  the strength of promotion encoded in the relevant promoter [9] and the duration of transcription due to strength of the TF binding site. [9] [10] [11] [12] [13] They also found that bacterial TFs linger too briefly for TFs' binding characteristics to explain the sustained transcription of bursts. [8]

Bacterial transcription differs from eukaryotic transcription in several ways. In bacteria, transcription and translation can occur simultaneously in the cytoplasm of the cell, whereas in eukaryotes transcription occurs in the nucleus and translation occurs in the cytoplasm. [14] There is only one type of bacterial RNA polymerase whereas eukaryotes have 3 types. [2] Bacteria have a σ-factor that detects and binds to promoter sites but eukaryotes do not need a σ-factor. Instead, eukaryotes have transcription factors that allow the recognition and binding of promoter sites. [2]

Overall, transcription within bacteria is a highly regulated process that is controlled by the integration of many signals at a given time. Bacteria heavily rely on transcription and translation to generate proteins that help them respond specifically to their environment. [4]

RNA polymerase

RNA polymerase is composed of a core and a holoenzyme structure. The core enzymes contains the catalytic properties of RNA polymerase and is made up of ββ′α2ω subunits. This sequence is conserved across all bacterial species. The holoenzyme is composed of a specific component known as the sigma factor (σ-factor). The sigma factor functions in aiding in promoter recognition, correct placement of RNA polymerase, and beginning unwinding at the start site. After the sigma factor performs its required function, it dissociates, while the catalytic portion remains on the DNA and continues transcription. [4] Additionally, RNA polymerase contains a core Mg+ ion that assists the enzyme with its catalytic properties. RNA polymerase works by catalyzing the nucleophilic attack of 3’ OH of RNA to the alpha phosphate of a complementary NTP molecule to create a growing strand of RNA from the template strand of DNA. Furthermore, RNA polymerase also displays exonuclease activities, meaning that if improper base pairing is detected, it can cut out the incorrect bases and replace them with the proper, correct one. [15]

Initiation

Initiation of transcription requires promoter regions, which are specific nucleotide consensus sequences that tell the σ-factor on RNA polymerase where to bind to the DNA. [1] The promoters are usually located 15 to 19 bases apart and are most commonly found upstream of the genes they control. [2] [1] RNA polymerase is made up of 4 subunits, which include two alphas, a beta, and a beta prime (α, α, β, and β'). A fifth subunit, sigma (called the σ-factor), is only present during initiation and detaches prior to elongation. Each subunit plays a role in the initiation of transcription, and the σ-factor must be present for initiation to occur. When all σ-factor is present, RNA polymerase is in its active form and is referred to as the holoenzyme. When the σ-factor detaches, it is in core polymerase form. [4] [1] The σ-factor recognizes promoter sequences at -35 and -10 regions and transcription begins at the start site (+1). The sequence of the -10 region is TATAAT and the sequence of the -35 region is TTGACA. [1]

Abortive cycling occurs prior to sigma factor release Abortive cycling.png
Abortive cycling occurs prior to sigma factor release

The promoter region is a prime regulator of transcription. Promoter regions regulate transcription of all genes within bacteria. As a result of their involvement, the sequence of base pairs within the promoter region is significant; the more similar the promoter region is to the consensus sequence, the tighter RNA polymerase will be able to bind. This binding contributes to the stability of elongation stage of transcription and overall results in more efficient functioning. Additionally, RNA polymerase and σ-factors are in limited supply within any given bacterial cell. Consequently, σ-factor binding to the promoter is affected by these limitations. All promoter regions contain sequences that are considered non-consensus and this helps to distribute σ-factors across the entirety of the genome. [17]

Elongation

During elongation, RNA polymerase slides down the double stranded DNA, unwinding it and transcribing (copying) its nucleotide sequence into newly synthesized RNA. The movement of the RNA-DNA complex is essential for the catalytic mechanism of RNA polymerase. Additionally, RNA polymerase increases the overall stability of this process by acting as a link between the RNA and DNA strands. [18] New nucleotides that are complementary to the DNA template strand are added to the 3' end of the RNA strand. [4] The newly formed RNA strand is practically identical to the DNA coding strand (sense strand or non-template strand), except it has uracil substituting thymine, and a ribose sugar backbone instead of a deoxyribose sugar backbone. Because nucleoside triphosphates (NTPs) need to attach to the OH- molecule on the 3' end of the RNA, transcription always occurs in the 5' to 3' direction. The four NTPs are adenosine-5'-triphosphate (ATP), guanoside-5'-triphosphate (GTP), uridine-5'-triphosphate (UTP), and cytidine-5'-triphosphate (CTP). [16] The attachment of NTPs onto the 3' end of the RNA transcript provides the energy required for this synthesis. [2] NTPs are also energy producing molecules that provide the fuel that drives chemical reactions in the cell. [4]

Multiple RNA polymerases can be active at once, meaning many strands of mRNA can be produced very quickly. [2] RNA polymerase moves down the DNA rapidly at approximately 40 bases per second. Due to the quick nature of this process, DNA is continually unwound ahead of RNA polymerase and then rewound once RNA polymerase moves along further. [18] [1] The polymerase has a proofreading mechanism that limits mistakes to about 1 in 10,000 nucleotides transcribed. [19] RNA polymerase has lower fidelity (accuracy) and speed than DNA polymerase. [2] DNA polymerase has a very different proofreading mechanism that includes exonuclease activity, which contributes to the higher fidelity. The consequence of an error during RNA synthesis is usually harmless, where as an error in DNA synthesis could be detrimental. [2]

The promoter sequence determines the frequency of transcription of its corresponding gene. [1]

Termination

In order for proper gene expression to occur, transcription must stop at specific sites. Two termination mechanisms are well known:

The termination of DNA transcription in bacteria may be stopped by certain mechanisms wherein the RNA polymerase will ignore the terminator sequence until the next one is reached. This phenomenon is known as antitermination and is utilized by certain bacteriophages. [20]

Related Research Articles

<span class="mw-page-title-main">Transcription (biology)</span> Process of copying a segment of DNA into RNA

Transcription is the process of copying a segment of DNA into RNA. The segments of DNA transcribed into RNA molecules that can encode proteins are said to produce messenger RNA (mRNA). Other segments of DNA are copied into RNA molecules called non-coding RNAs (ncRNAs). mRNA comprises only 1–3% of total RNA samples. Less than 2% of the human genome can be transcribed into mRNA, while at least 80% of mammalian genomic DNA can be actively transcribed, with the majority of this 80% considered to be ncRNA.

<span class="mw-page-title-main">RNA polymerase</span> Enzyme that synthesizes RNA from DNA

In molecular biology, RNA polymerase, or more specifically DNA-directed/dependent RNA polymerase (DdRP), is an enzyme that catalyzes the chemical reactions that synthesize RNA from a DNA template.

<span class="mw-page-title-main">Rho factor</span> Prokaryotic protein

A ρ factor is a bacterial protein involved in the termination of transcription. Rho factor binds to the transcription terminator pause site, an exposed region of single stranded RNA after the open reading frame at C-rich/G-poor sequences that lack obvious secondary structure.

A sigma factor is a protein needed for initiation of transcription in bacteria. It is a bacterial transcription initiation factor that enables specific binding of RNA polymerase (RNAP) to gene promoters. It is homologous to archaeal transcription factor B and to eukaryotic factor TFIIB. The specific sigma factor used to initiate transcription of a given gene will vary, depending on the gene and on the environmental signals needed to initiate transcription of that gene. Selection of promoters by RNA polymerase is dependent on the sigma factor that associates with it. They are also found in plant chloroplasts as a part of the bacteria-like plastid-encoded polymerase (PEP).

In genetics, a transcription terminator is a section of nucleic acid sequence that marks the end of a gene or operon in genomic DNA during transcription. This sequence mediates transcriptional termination by providing signals in the newly synthesized transcript RNA that trigger processes which release the transcript RNA from the transcriptional complex. These processes include the direct interaction of the mRNA secondary structure with the complex and/or the indirect activities of recruited termination factors. Release of the transcriptional complex frees RNA polymerase and related transcriptional machinery to begin transcription of new mRNAs.

DnaG is a bacterial DNA primase and is encoded by the dnaG gene. The enzyme DnaG, and any other DNA primase, synthesizes short strands of RNA known as oligonucleotides during DNA replication. These oligonucleotides are known as primers because they act as a starting point for DNA synthesis. DnaG catalyzes the synthesis of oligonucleotides that are 10 to 60 nucleotides long, however most of the oligonucleotides synthesized are 11 nucleotides. These RNA oligonucleotides serve as primers, or starting points, for DNA synthesis by bacterial DNA polymerase III. DnaG is important in bacterial DNA replication because DNA polymerase cannot initiate the synthesis of a DNA strand, but can only add nucleotides to a preexisting strand. DnaG synthesizes a single RNA primer at the origin of replication. This primer serves to prime leading strand DNA synthesis. For the other parental strand, the lagging strand, DnaG synthesizes an RNA primer every few kilobases (kb). These primers serve as substrates for the synthesis of Okazaki fragments.

The Pribnow box is a sequence of TATAAT of six nucleotides that is an essential part of a promoter site on DNA for transcription to occur in bacteria. It is an idealized or consensus sequence—that is, it shows the most frequently occurring base at each position in many promoters analyzed; individual promoters often vary from the consensus at one or more positions. It is also commonly called the -10 sequence or element, because it is centered roughly ten base pairs upstream from the site of initiation of transcription.

<span class="mw-page-title-main">Transcription preinitiation complex</span> Complex of proteins necessary for gene transcription in eukaryotes and archaea

The preinitiation complex is a complex of approximately 100 proteins that is necessary for the transcription of protein-coding genes in eukaryotes and archaea. The preinitiation complex positions RNA polymerase II at gene transcription start sites, denatures the DNA, and positions the DNA in the RNA polymerase II active site for transcription.

<span class="mw-page-title-main">RNA polymerase II</span> Protein complex that transcribes DNA

RNA polymerase II is a multiprotein complex that transcribes DNA into precursors of messenger RNA (mRNA) and most small nuclear RNA (snRNA) and microRNA. It is one of the three RNAP enzymes found in the nucleus of eukaryotic cells. A 550 kDa complex of 12 subunits, RNAP II is the most studied type of RNA polymerase. A wide range of transcription factors are required for it to bind to upstream gene promoters and begin transcription.

<span class="mw-page-title-main">General transcription factor</span> Class of protein transcription factors

General transcription factors (GTFs), also known as basal transcriptional factors, are a class of protein transcription factors that bind to specific sites (promoter) on DNA to activate transcription of genetic information from DNA to messenger RNA. GTFs, RNA polymerase, and the mediator constitute the basic transcriptional apparatus that first bind to the promoter, then start transcription. GTFs are also intimately involved in the process of gene regulation, and most are required for life.

In molecular biology, a termination factor is a protein that mediates the termination of RNA transcription by recognizing a transcription terminator and causing the release of the newly made mRNA. This is part of the process that regulates the transcription of RNA to preserve gene expression integrity and are present in both eukaryotes and prokaryotes, although the process in bacteria is more widely understood. The most extensively studied and detailed transcriptional termination factor is the Rho (ρ) protein of E. coli.

In eukaryote cells, RNA polymerase III is a protein that transcribes DNA to synthesize 5S ribosomal RNA, tRNA and other small RNAs.

Bacterial translation is the process by which messenger RNA is translated into proteins in bacteria.

<span class="mw-page-title-main">Transcription bubble</span>

A transcription bubble is a molecular structure formed during DNA transcription when a limited portion of the DNA double helix is unwound. The size of a transcription bubble ranges from 12 to 14 base pairs. A transcription bubble is formed when the RNA polymerase enzyme binds to a promoter and causes two DNA strands to detach. It presents a region of unpaired DNA, where a short stretch of nucleotides are exposed on each strand of the double helix.

<span class="mw-page-title-main">RNA-dependent RNA polymerase</span> Enzyme that synthesizes RNA from an RNA template

RNA-dependent RNA polymerase (RdRp) or RNA replicase is an enzyme that catalyzes the replication of RNA from an RNA template. Specifically, it catalyzes synthesis of the RNA strand complementary to a given RNA template. This is in contrast to typical DNA-dependent RNA polymerases, which all organisms use to catalyze the transcription of RNA from a DNA template.

<span class="mw-page-title-main">Prokaryotic DNA replication</span> DNA Replication in prokaryotes

Prokaryotic DNA Replication is the process by which a prokaryote duplicates its DNA into another copy that is passed on to daughter cells. Although it is often studied in the model organism E. coli, other bacteria show many similarities. Replication is bi-directional and originates at a single origin of replication (OriC). It consists of three steps: Initiation, elongation, and termination.

<span class="mw-page-title-main">Eukaryotic transcription</span> Transcription is heterocatalytic function of DNA

Eukaryotic transcription is the elaborate process that eukaryotic cells use to copy genetic information stored in DNA into units of transportable complementary RNA replica. Gene transcription occurs in both eukaryotic and prokaryotic cells. Unlike prokaryotic RNA polymerase that initiates the transcription of all different types of RNA, RNA polymerase in eukaryotes comes in three variations, each translating a different type of gene. A eukaryotic cell has a nucleus that separates the processes of transcription and translation. Eukaryotic transcription occurs within the nucleus where DNA is packaged into nucleosomes and higher order chromatin structures. The complexity of the eukaryotic genome necessitates a great variety and complexity of gene expression control.

<span class="mw-page-title-main">Intrinsic termination</span>

Intrinsic, or rho-independent termination, is a process in prokaryotes to signal the end of transcription and release the newly constructed RNA molecule. In prokaryotes such as E. coli, transcription is terminated either by a rho-dependent process or rho-independent process. In the Rho-dependent process, the rho-protein locates and binds the signal sequence in the mRNA and signals for cleavage. Contrarily, intrinsic termination does not require a special protein to signal for termination and is controlled by the specific sequences of RNA. When the termination process begins, the transcribed mRNA forms a stable secondary structure hairpin loop, also known as a Stem-loop. This RNA hairpin is followed by multiple uracil nucleotides. The bonds between uracil and adenine are very weak. A protein bound to RNA polymerase (nusA) binds to the stem-loop structure tightly enough to cause the polymerase to temporarily stall. This pausing of the polymerase coincides with transcription of the poly-uracil sequence. The weak adenine-uracil bonds lower the energy of destabilization for the RNA-DNA duplex, allowing it to unwind and dissociate from the RNA polymerase. Overall, the modified RNA structure is what terminates transcription.

<span class="mw-page-title-main">Transcription factor II B</span> Mammalian protein found in Homo sapiens

Transcription factor II B (TFIIB) is a general transcription factor that is involved in the formation of the RNA polymerase II preinitiation complex (PIC) and aids in stimulating transcription initiation. TFIIB is localised to the nucleus and provides a platform for PIC formation by binding and stabilising the DNA-TBP complex and by recruiting RNA polymerase II and other transcription factors. It is encoded by the TFIIB gene, and is homologous to archaeal transcription factor B and analogous to bacterial sigma factors.

RNA polymerase II holoenzyme is a form of eukaryotic RNA polymerase II that is recruited to the promoters of protein-coding genes in living cells. It consists of RNA polymerase II, a subset of general transcription factors, and regulatory proteins known as SRB proteins.

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