Terminator (genetics)

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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.

Contents

In prokaryotes

Simplified schematics of the mechanisms of prokaryotic transcriptional termination. In Rho-independent termination, a terminating hairpin forms on the nascent mRNA interacting with the NusA protein to stimulate release of the transcript from the RNA polymerase complex (top). In Rho-dependent termination, the Rho protein binds at the upstream rut site, translocates down the mRNA, and interacts with the RNA polymerase complex to stimulate release of the transcript. Prokaryotic terminators-en.svg
Simplified schematics of the mechanisms of prokaryotic transcriptional termination. In Rho-independent termination, a terminating hairpin forms on the nascent mRNA interacting with the NusA protein to stimulate release of the transcript from the RNA polymerase complex (top). In Rho-dependent termination, the Rho protein binds at the upstream rut site, translocates down the mRNA, and interacts with the RNA polymerase complex to stimulate release of the transcript.

Two classes of transcription terminators, Rho-dependent and Rho-independent, have been identified throughout prokaryotic genomes. These widely distributed sequences are responsible for triggering the end of transcription upon normal completion of gene or operon transcription, mediating early termination of transcripts as a means of regulation such as that observed in transcriptional attenuation, and to ensure the termination of runaway transcriptional complexes that manage to escape earlier terminators by chance, which prevents unnecessary energy expenditure for the cell.

Rho-dependent terminators

Rho-dependent transcription terminators require a large protein called a Rho factor which exhibits RNA helicase activity to disrupt the mRNA-DNA-RNA polymerase transcriptional complex. Rho-dependent terminators are found in bacteria and phages. The Rho-dependent terminator occurs downstream of translational stop codons and consists of an unstructured, cytosine-rich sequence on the mRNA known as a Rho utilization site (rut), [1] and a downstream transcription stop point (tsp). The rut serves as a mRNA loading site and as an activator for Rho; activation enables Rho to efficiently hydrolyze ATP and translocate down the mRNA while it maintains contact with the rut site. Rho is able to catch up with the RNA polymerase because it is being stalled at the downstream tsp sites. Multiple different sequences can function as a tsp site. [2] Contact between Rho and the RNA polymerase complex stimulates dissociation of the transcriptional complex through a mechanism involving allosteric effects of Rho on RNA polymerase. [3] [4]

Rho-independent terminators

Intrinsic transcription terminators or Rho-independent terminators require the formation of a self-annealing hairpin structure on the elongating transcript, which results in the disruption of the mRNA-DNA-RNA polymerase ternary complex. The terminator sequence in DNA contains a 20 basepair GC-rich region of dyad symmetry followed by a short poly-A tract or "A stretch" which is transcribed to form the terminating hairpin and a 7–9 nucleotide "U tract" respectively. The mechanism of termination is hypothesized to occur through a combination of direct promotion of dissociation through allosteric effects of hairpin binding interactions with the RNA polymerase and "competitive kinetics". The hairpin formation causes RNA polymerase stalling and destabilization, leading to a greater likelihood that dissociation of the complex will occur at that location due to increased time spent paused at that site and reduced stability of the complex. [5] [6] Additionally, the elongation protein factor NusA interacts with the RNA polymerase and the hairpin structure to stimulate transcriptional termination. [7]

In eukaryotes

In eukaryotic transcription of mRNAs, terminator signals are recognized by protein factors that are associated with the RNA polymerase II and which trigger the termination process. The genome encodes one or more polyadenylation signals. Once the signals are transcribed into the mRNA, the proteins cleavage and polyadenylation specificity factor (CPSF) and cleavage stimulation factor (CstF) transfer from the carboxyl terminal domain of RNA polymerase II to the poly-A signal. These two factors then recruit other proteins to the site to cleave the transcript, freeing the mRNA from the transcription complex, and add a string of about 200 A-repeats to the 3' end of the mRNA in a process known as polyadenylation. During these processing steps, the RNA polymerase continues to transcribe for several hundred to a few thousand bases and eventually dissociates from the DNA and downstream transcript through an unclear mechanism; there are two basic models for this event known as the torpedo and allosteric models. [8] [9]

Torpedo model

After the mRNA is completed and cleaved off at the poly-A signal sequence, the left-over (residual) RNA strand remains bound to the DNA template and the RNA polymerase II unit, continuing to be transcribed. After this cleavage, a so-called exonuclease binds to the residual RNA strand and removes the freshly transcribed nucleotides one at a time (also called 'degrading' the RNA), moving towards the bound RNA polymerase II. This exonuclease is XRN2 (5'-3' Exoribonuclease 2) in humans. This model proposes that XRN2 proceeds to degrade the uncapped residual RNA from 5' to 3' until it reaches the RNA pol II unit. This causes the exonuclease to 'push off' the RNA pol II unit as it moves past it, terminating the transcription while also cleaning up the residual RNA strand.

Similar to Rho-dependent termination, XRN2 triggers the dissociation of RNA polymerase II by either pushing the polymerase off of the DNA template or pulling the template out of the RNA polymerase. [10] The mechanism by which this happens remains unclear, however, and has been challenged not to be the sole cause of the dissociation. [11]

In order to protect the transcribed mRNA from degradation by the exonuclease, a 5' cap is added to the strand. This is a modified guanine added to the front of mRNA, which prevents the exonuclease from binding and degrading the RNA strand. A 3' poly(A) tail is added to the end of a mRNA strand for protection from other exonucleases as well.

Allosteric model

The allosteric model suggests that termination occurs due to the structural change of the RNA polymerase unit after binding to or losing some of its associated proteins, making it detach from the DNA strand after the signal. [9] This would occur after the RNA pol II unit has transcribed the poly-A signal sequence, which acts as a terminator signal.

RNA polymerase is normally capable of transcribing DNA into single-stranded mRNA efficiently. However, upon transcribing over the poly-A signals on the DNA template, a conformational shift is induced in the RNA polymerase from the proposed loss of associated proteins from its carboxyl terminal domain. This change of conformation reduces RNA polymerase's processivity making the enzyme more prone to dissociating from its DNA-RNA substrate. In this case, termination is not completed by degradation of mRNA but instead is mediated by limiting the elongation efficiency of RNA polymerase and thus increasing the likelihood that the polymerase will dissociate and end its current cycle of transcription. [8]

Non-mRNAs

The several RNA polymerases in eukaryotes each have their own means of termination. Pol I is stopped by TTF1 (yeast Nsi1), which recognizes a downstream DNA sequence; the endonuclease is XRN2 (yeast Rat1). Pol III is able to terminate on its on on a stretch of As on the template strand. [12]

Finally, Pol II also have poly(A)-independent modes of termination, which is required when it transcribes snRNA and snoRNA genes in yeast. The yeast protein Nrd1 is responsible. [9] Some human mechanism, possibly PCF11, seems to cause premature termination when pol II transcribes HIV genes. [13]

See also

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.

In molecular biology and genetics, transcriptional regulation is the means by which a cell regulates the conversion of DNA to RNA (transcription), thereby orchestrating gene activity. A single gene can be regulated in a range of ways, from altering the number of copies of RNA that are transcribed, to the temporal control of when the gene is transcribed. This control allows the cell or organism to respond to a variety of intra- and extracellular signals and thus mount a response. Some examples of this include producing the mRNA that encode enzymes to adapt to a change in a food source, producing the gene products involved in cell cycle specific activities, and producing the gene products responsible for cellular differentiation in multicellular eukaryotes, as studied in evolutionary developmental biology.

<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.

<span class="mw-page-title-main">Exonuclease</span> Class of enzymes; type of nuclease

Exonucleases are enzymes that work by cleaving nucleotides one at a time from the end (exo) of a polynucleotide chain. A hydrolyzing reaction that breaks phosphodiester bonds at either the 3′ or the 5′ end occurs. Its close relative is the endonuclease, which cleaves phosphodiester bonds in the middle (endo) of a polynucleotide chain. Eukaryotes and prokaryotes have three types of exonucleases involved in the normal turnover of mRNA: 5′ to 3′ exonuclease (Xrn1), which is a dependent decapping protein; 3′ to 5′ exonuclease, an independent protein; and poly(A)-specific 3′ to 5′ exonuclease.

RNA polymerase 1 is, in higher eukaryotes, the polymerase that only transcribes ribosomal RNA, a type of RNA that accounts for over 50% of the total RNA synthesized in a cell.

<span class="mw-page-title-main">Stem-loop</span> Intramolecular base-pairing pattern in RNA and DNA

Stem-loop intramolecular base pairing is a pattern that can occur in single-stranded RNA. The structure is also known as a hairpin or hairpin loop. It occurs when two regions of the same strand, usually complementary in nucleotide sequence when read in opposite directions, base-pair to form a double helix that ends in an unpaired loop. The resulting structure is a key building block of many RNA secondary structures. As an important secondary structure of RNA, it can direct RNA folding, protect structural stability for messenger RNA (mRNA), provide recognition sites for RNA binding proteins, and serve as a substrate for enzymatic reactions.

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.

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<span class="mw-page-title-main">Termination signal</span>

A termination signal is a sequence that signals the end of transcription or translation. Termination signals are found at the end of the part of the chromosome being transcribed during transcription of mRNA. Termination signals bring a stop to transcription, ensuring that only gene-encoding parts of the chromosome are transcribed. Transcription begins at the promoter when RNA polymerase, an enzyme that facilitates transcription of DNA into mRNA, binds to a promoter, unwinds the helical structure of the DNA, and uses the single-stranded DNA as a template to synthesize RNA. Once RNA polymerase reaches the termination signal, transcription is terminated. In bacteria, there are two main types of termination signals: intrinsic and factor-dependent terminators. In the context of translation, a termination signal is the stop codon on the mRNA that elicits the release of the growing peptide from the ribosome.

<i>trp</i> operon Operon that codes for the components for production of tryptophan

The trp operon is a group of genes that are transcribed together, encoding the enzymes that produce the amino acid tryptophan in bacteria. The trp operon was first characterized in Escherichia coli, and it has since been discovered in many other bacteria. The operon is regulated so that, when tryptophan is present in the environment, the genes for tryptophan synthesis are repressed.

Antitermination is the prokaryotic cell's aid to fix premature termination of RNA synthesis during the transcription of RNA. It occurs when the RNA polymerase ignores the termination signal and continues elongating its transcript until a second signal is reached. Antitermination provides a mechanism whereby one or more genes at the end of an operon can be switched either on or off, depending on the polymerase either recognizing or not recognizing the termination signal.

<span class="mw-page-title-main">Bacterial transcription</span>

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.

<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">5'-3' exoribonuclease 2</span> Protein-coding gene in the species Homo sapiens

5'-3' Exoribonuclease 2 (XRN2) also known as Dhm1-like protein is an exoribonuclease enzyme that in humans is encoded by the XRN2 gene.

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

Intrinsic, or rho-independent termination, is a process to signal the end of transcription and release the newly constructed RNA molecule. In bacteria 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 (rU) and adenine (dA) 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.

Post-transcriptional regulation is the control of gene expression at the RNA level. It occurs once the RNA polymerase has been attached to the gene's promoter and is synthesizing the nucleotide sequence. Therefore, as the name indicates, it occurs between the transcription phase and the translation phase of gene expression. These controls are critical for the regulation of many genes across human tissues. It also plays a big role in cell physiology, being implicated in pathologies such as cancer and neurodegenerative diseases.

<span class="mw-page-title-main">Abortive initiation</span>

Abortive initiation, also known as abortive transcription, is an early process of genetic transcription in which RNA polymerase binds to a DNA promoter and enters into cycles of synthesis of short mRNA transcripts which are released before the transcription complex leaves the promoter. This process occurs in both eukaryotes and prokaryotes. Abortive initiation is typically studied in the T3 and T7 RNA polymerases in bacteriophages and in E. coli.

Transcription-translation coupling is a mechanism of gene expression regulation in which synthesis of an mRNA (transcription) is affected by its concurrent decoding (translation). In prokaryotes, mRNAs are translated while they are transcribed. This allows communication between RNA polymerase, the multisubunit enzyme that catalyzes transcription, and the ribosome, which catalyzes translation. Coupling involves both direct physical interactions between RNA polymerase and the ribosome, as well as ribosome-induced changes to the structure and accessibility of the intervening mRNA that affect transcription.

References

  1. ↑ Di Salvo, Marco; Puccio, Simone; Peano, Clelia; Lacour, Stephan; Alifano, Pietro (7 March 2019). "RhoTermPredict: an algorithm for predicting Rho-dependent transcription terminators based on Escherichia coli, Bacillus subtilis and Salmonella enterica databases". BMC Bioinformatics. 20 (1): 117. doi: 10.1186/s12859-019-2704-x . PMC   6407284 . PMID   30845912.
  2. ↑ Richardson, J. P. (1996). "Rho-dependent Termination of Transcription Is Governed Primarily by the Upstream Rho Utilization (rut) Sequences of a Terminator". Journal of Biological Chemistry. 271 (35): 21597–21603. doi: 10.1074/jbc.271.35.21597 . ISSN   0021-9258. PMID   8702947.
  3. ↑ Ciampi, MS. (Sep 2006). "Rho-dependent terminators and transcription termination". Microbiology. 152 (Pt 9): 2515–28. doi: 10.1099/mic.0.28982-0 . PMID   16946247.
  4. ↑ Epshtein, V; Dutta, D; Wade, J; Nudler, E (Jan 14, 2010). "An allosteric mechanism of Rho-dependent transcription termination". Nature. 463 (7278): 245–9. Bibcode:2010Natur.463..245E. doi:10.1038/nature08669. PMC   2929367 . PMID   20075920.
  5. ↑ von Hippel, P. H. (1998). "An Integrated Model of the Transcription Complex in Elongation, Termination, and Editing". Science. 281 (5377): 660–665. Bibcode:1998Sci...281..660.. doi:10.1126/science.281.5377.660. PMID   9685251. S2CID   11046390.
  6. ↑ Gusarov, Ivan; Nudler, Evgeny (1999). "The Mechanism of Intrinsic Transcription Termination". Molecular Cell. 3 (4): 495–504. doi: 10.1016/S1097-2765(00)80477-3 . ISSN   1097-2765. PMID   10230402.
  7. ↑ Santangelo, TJ.; Artsimovitch, I. (May 2011). "Termination and antitermination: RNA polymerase runs a stop sign". Nat Rev Microbiol. 9 (5): 319–29. doi:10.1038/nrmicro2560. PMC   3125153 . PMID   21478900.
  8. 1 2 Watson, J. (2008). Molecular Biology of the Gene. Cold Spring Harbor Laboratory Press. pp. 410–411. ISBN   978-0-8053-9592-1.
  9. 1 2 3 Rosonina, Emanuel; Kaneko, Syuzo; Manley, James L. (2006-05-01). "Terminating the transcript: breaking up is hard to do". Genes & Development. 20 (9): 1050–1056. doi: 10.1101/gad.1431606 . ISSN   0890-9369. PMID   16651651.
  10. ↑ Luo, W.; Bartley D. (2004). "A ribonucleolytic rat torpedoes RNA polymerase II". Cell. 119 (7): 911–914. doi: 10.1016/j.cell.2004.11.041 . PMID   15620350.
  11. ↑ Luo, Weifei; Johnson, Arlen W.; Bentley, David L. (2006-04-15). "The role of Rat1 in coupling mRNA 3′-end processing to transcription termination: implications for a unified allosteric–torpedo model". Genes & Development. 20 (8): 954–965. doi:10.1101/gad.1409106. ISSN   0890-9369. PMC   1472303 . PMID   16598041.
  12. ↑ Arimbasseri, AG; Rijal, K; Maraia, RJ (March 2013). "Transcription termination by the eukaryotic RNA polymerase III". Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms. 1829 (3–4): 318–30. doi:10.1016/j.bbagrm.2012.10.006. PMC   3568203 . PMID   23099421.
  13. ↑ Gilmour, David S.; Fan, Ruopeng (January 2008). "Derailing the Locomotive: Transcription Termination". Journal of Biological Chemistry. 283 (2): 661–664. doi: 10.1074/jbc.R700032200 . PMID   17998201.
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