Riboregulator
Encyclopedia
In molecular biology
, a riboregulator is a ribonucleic acid (RNA
) that responds to a signal nucleic acid
molecule by Watson-Crick base pairing. A riboregulator may respond to a signal molecule in any number of manners including, translation
(or repression of translation) of the RNA into a protein
, activation of a ribozyme
, release of silencing RNA (siRNA
), conformational change, and/or binding other nucleic acids.
Riboregulators contain two canonical domains, a sensor domain and an effector domain. These domains are also found on riboswitches, but unlike riboswitches, the sensor domain only binds complementary RNA or DNA
strands as opposed to small molecule
s. Because binding is based on base-pairing, a riboregulator can be tailored to differentiate and respond to individual genetic sequences and combinations thereof.
complex to scan, assemble, and/or translate an RNA molecule into a protein. In translational riboregulators, the RNA molecule is repressed or de-repressed depending on the secondary structure
of the RNA molecule. Signal-responsive structures are usually introduced into the 5' untranslated region (5' UTR) of the RNA molecules using standard molecular biological techniques.
As discovered by Marilyn Kozak, the small (40S
) ribosome complex scans an RNA molecule from 5' untranslated region to the start codon. When the complex encounters secondary structure, it must melt the structure to reach the start codon or it will fall off the molecule. The complex moves along through the untranslated region until it stalls just prior to reaching the start codon
because it encounters a highly conserved sequence (a Kozak consensus sequence
in eukaryotes, or Shine-Dalgarno sequence
in prokaryotes). The stalled complex then combines with the large ribosome (60S
) to begin translating the RNA into protein.
Lechner invented the first riboregulator in 1991. Lechner used a self pairing stem-loop
which inhibited translation of a prokaryotic RNA, unless a complementary RNA sequence (anti-inhibitor) was present. In 1997, Black engineered the first eukaryotic riboregulator using antisense molecules to prevent translation (Nucline RNA). In the Nucline system, antisense molecules block translation unless removed via competitive hybridization and strand-displacement by specific signal RNA sequences such as HIV RNA and oncogene RNA. In 2003, Black demonstrated that Nucline RNA could perform Boolean
and arithmetic operations (If-then-else, AND gate
, OR gate
and "Molar concentration of x > molar concentration of y" calculations ) in HIV
infected monocytes and breast cancer
cells.
In 2004, Isaacs et al. modified Lechner’s original system to demonstrate both trans and cis repression in a prokaryotic cells. Bayer & Smolke further developed a translational riboregulator in 2005 that could respond to small molecules creating a hybrid riboswitch
/riboregulator molecule, termed an anti-switch. In an anti-switch, the presence of a small organic molecule binds an aptamer
sequence in the RNA molecule which unmasks an otherwise sequestered antisense sequence, which can bind and block target RNA translation.
, NOR
, and OR gates.
in response to pH
changes. The only known pH riboregulator upregulates the alx gene in E. coli when in alkaline conditions, particularly above pH 8. The pH riboregulator was found, through experimentation, to reside in the 5' UTR of the alx gene. When in pH 7 or below, the pH riboregulator element (PRE) exists in an inactive 'N' state; in alkali conditions, RNA polymerase
controls the folding of this ncRNA to its 'H' active form. The ribosome binding site of the alx gene is now exposed and the 30s subunit is allowed to bind.
and personalized medicine
. In synthetic biology, riboregulators can be used to regulate bacterial responses and probe gene regulatory networks. Undergraduate systems biology students use standardized riboregulators to compete in an annual bio-engineering competition (The international Genetically Engineered Machine competition, iGEM
). In addition, because the various riboregulators can be tailored to respond to complex biological signals, riboregulators offer the promise of individually tailored RNA based drugs which respond to genes, small molecules and proteins in individuals cells.
Molecular biology
Molecular biology is the branch of biology that deals with the molecular basis of biological activity. This field overlaps with other areas of biology and chemistry, particularly genetics and biochemistry...
, a riboregulator is a ribonucleic acid (RNA
RNA
Ribonucleic acid , or RNA, is one of the three major macromolecules that are essential for all known forms of life....
) that responds to a signal nucleic acid
Nucleic acid
Nucleic acids are biological molecules essential for life, and include DNA and RNA . Together with proteins, nucleic acids make up the most important macromolecules; each is found in abundance in all living things, where they function in encoding, transmitting and expressing genetic information...
molecule by Watson-Crick base pairing. A riboregulator may respond to a signal molecule in any number of manners including, translation
Translation
Translation is the communication of the meaning of a source-language text by means of an equivalent target-language text. Whereas interpreting undoubtedly antedates writing, translation began only after the appearance of written literature; there exist partial translations of the Sumerian Epic of...
(or repression of translation) of the RNA into a protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
, activation of a ribozyme
Ribozyme
A ribozyme is an RNA molecule with a well defined tertiary structure that enables it to catalyze a chemical reaction. Ribozyme means ribonucleic acid enzyme. It may also be called an RNA enzyme or catalytic RNA. Many natural ribozymes catalyze either the hydrolysis of one of their own...
, release of silencing RNA (siRNA
Sírna
Sírna Sáeglach , son of Dian mac Demal, son of Demal mac Rothechtaid, son of Rothechtaid mac Main, was, according to medieval Irish legend and historical tradition, a High King of Ireland...
), conformational change, and/or binding other nucleic acids.
Riboregulators contain two canonical domains, a sensor domain and an effector domain. These domains are also found on riboswitches, but unlike riboswitches, the sensor domain only binds complementary RNA or DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
strands as opposed to small molecule
Small molecule
In the fields of pharmacology and biochemistry, a small molecule is a low molecular weight organic compound which is by definition not a polymer...
s. Because binding is based on base-pairing, a riboregulator can be tailored to differentiate and respond to individual genetic sequences and combinations thereof.
Types of riboregulators
Translational riboregulator
Translational riboregulators regulate the ability of a ribosomeRibosome
A ribosome is a component of cells that assembles the twenty specific amino acid molecules to form the particular protein molecule determined by the nucleotide sequence of an RNA molecule....
complex to scan, assemble, and/or translate an RNA molecule into a protein. In translational riboregulators, the RNA molecule is repressed or de-repressed depending on the secondary structure
Secondary structure
In biochemistry and structural biology, secondary structure is the general three-dimensional form of local segments of biopolymers such as proteins and nucleic acids...
of the RNA molecule. Signal-responsive structures are usually introduced into the 5' untranslated region (5' UTR) of the RNA molecules using standard molecular biological techniques.
As discovered by Marilyn Kozak, the small (40S
40S
40S is the small subunit of eukaryotic ribosomes.It interacts with the internal ribosome entry site of the hepatitis C virus.The following is a list of proteins contained in the 40S ribosome:...
) ribosome complex scans an RNA molecule from 5' untranslated region to the start codon. When the complex encounters secondary structure, it must melt the structure to reach the start codon or it will fall off the molecule. The complex moves along through the untranslated region until it stalls just prior to reaching the start codon
Start codon
The start codon is generally defined as the point, sequence, at which a ribosome begins to translate a sequence of RNA into amino acids.When an RNA transcript is "read" from the 5' carbon to the 3' carbon by the ribosome the start codon is the first codon on which the tRNA bound to Met,...
because it encounters a highly conserved sequence (a Kozak consensus sequence
Kozak consensus sequence
The Kozak consensus sequence, Kozak consensus or Kozak sequence, is a sequence which occurs on eukaryotic mRNA and has the consensus gccRccAUGG, where R is a purine three bases upstream of the start codon , which is followed by another 'G'. The Kozak consensus sequence plays a major role in the...
in eukaryotes, or Shine-Dalgarno sequence
Shine-Dalgarno sequence
The Shine-Dalgarno sequence , proposed by Australian scientists John Shine and Lynn Dalgarno , is a ribosomal binding site in the mRNA, generally located 8 basepairs upstream of the start codon AUG. The Shine-Dalgarno sequence exists only in prokaryotes. The six-base consensus sequence is AGGAGG;...
in prokaryotes). The stalled complex then combines with the large ribosome (60S
60S
60S is the large ribosomal subunit in eukaryotes. It corresponds to 50S in prokaryotes.It consists of the following:* 5S* 28S* 5.8SThe following is a list of proteins contained within the 60S ribosome:...
) to begin translating the RNA into protein.
Lechner invented the first riboregulator in 1991. Lechner used a self pairing stem-loop
Stem-loop
Stem-loop intramolecular base pairing is a pattern that can occur in single-stranded DNA or, more commonly, in 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,...
which inhibited translation of a prokaryotic RNA, unless a complementary RNA sequence (anti-inhibitor) was present. In 1997, Black engineered the first eukaryotic riboregulator using antisense molecules to prevent translation (Nucline RNA). In the Nucline system, antisense molecules block translation unless removed via competitive hybridization and strand-displacement by specific signal RNA sequences such as HIV RNA and oncogene RNA. In 2003, Black demonstrated that Nucline RNA could perform Boolean
Boolean logic
Boolean algebra is a logical calculus of truth values, developed by George Boole in the 1840s. It resembles the algebra of real numbers, but with the numeric operations of multiplication xy, addition x + y, and negation −x replaced by the respective logical operations of...
and arithmetic operations (If-then-else, AND gate
AND gate
The AND gate is a basic digital logic gate that implements logical conjunction - it behaves according to the truth table to the right. A HIGH output results only if both the inputs to the AND gate are HIGH . If neither or only one input to the AND gate is HIGH, a LOW output results...
, OR gate
OR gate
The OR gate is a digital logic gate that implements logical disjunction - it behaves according to the truth table to the right. A HIGH output results if one or both the inputs to the gate are HIGH . If neither input is HIGH, a LOW output results...
and "Molar concentration of x > molar concentration of y" calculations ) in HIV
HIV
Human immunodeficiency virus is a lentivirus that causes acquired immunodeficiency syndrome , a condition in humans in which progressive failure of the immune system allows life-threatening opportunistic infections and cancers to thrive...
infected monocytes and breast cancer
Breast cancer
Breast cancer is cancer originating from breast tissue, most commonly from the inner lining of milk ducts or the lobules that supply the ducts with milk. Cancers originating from ducts are known as ductal carcinomas; those originating from lobules are known as lobular carcinomas...
cells.
In 2004, Isaacs et al. modified Lechner’s original system to demonstrate both trans and cis repression in a prokaryotic cells. Bayer & Smolke further developed a translational riboregulator in 2005 that could respond to small molecules creating a hybrid riboswitch
Riboswitch
In molecular biology, a riboswitch is a part of an mRNA molecule that can directly bind a small target molecule, and whose binding of the target affects the gene's activity. Thus, an mRNA that contains a riboswitch is directly involved in regulating its own activity, in response to the...
/riboregulator molecule, termed an anti-switch. In an anti-switch, the presence of a small organic molecule binds an aptamer
Aptamer
Aptamers are oligonucleic acid or peptide molecules that bind to a specific target molecule. Aptamers are usually created by selecting them from a large random sequence pool, but natural aptamers also exist in riboswitches. Aptamers can be used for both basic research and clinical purposes as...
sequence in the RNA molecule which unmasks an otherwise sequestered antisense sequence, which can bind and block target RNA translation.
Ribozyme riboregulator
Ribozyme riboregulators regulate the ability of a catalytic RNA molecule to cleave a target nucleic acid sequence. In ribozyme riboregulators, a hammerhead ribozyme RNA molecule is activated or inactivated depending on the change of the secondary structure induced by hybridizing a signal molecule such as a cognate DNA or RNA sequence. In 2008, Win & Smolke designed a ribozyme regulator that could function in yeast cells that carried out Boolean operations similar to the earlier translational riboregulators, including AND, NANDNand
NAND may stand for:*Nand , an Indian classical raga.*Logical NAND , a binary operation in logic.**NAND gate, an electronic gate that implements a logical NAND....
, NOR
Nor
Nor may refer to:*In grammar, nor is a coordinating conjunction*Nór, the eponymous founder-king of Norway in Norse mythology*Nor , a character in the book in Wicked...
, and OR gates.
RNAi-based riboregulator
RNAi riboregulators are small interfering RNAs which response to a signal input such as complementary hybridization with a DNA or RNA molecule. The presence or absence of a target molecule determines whether the siRNA downregulates gene expression. In 2007, Rinaudo et al. demonstrated that RNAi based riboregulators can also perform Boolean operations in cells.pH riboregulator
pH riboregulators regulate gene expressionGene expression
Gene expression is the process by which information from a gene is used in the synthesis of a functional gene product. These products are often proteins, but in non-protein coding genes such as ribosomal RNA , transfer RNA or small nuclear RNA genes, the product is a functional RNA...
in response to pH
PH
In chemistry, pH is a measure of the acidity or basicity of an aqueous solution. Pure water is said to be neutral, with a pH close to 7.0 at . Solutions with a pH less than 7 are said to be acidic and solutions with a pH greater than 7 are basic or alkaline...
changes. The only known pH riboregulator upregulates the alx gene in E. coli when in alkaline conditions, particularly above pH 8. The pH riboregulator was found, through experimentation, to reside in the 5' UTR of the alx gene. When in pH 7 or below, the pH riboregulator element (PRE) exists in an inactive 'N' state; in alkali conditions, RNA polymerase
RNA polymerase
RNA polymerase is an enzyme that produces RNA. In cells, RNAP is needed for constructing RNA chains from DNA genes as templates, a process called transcription. RNA polymerase enzymes are essential to life and are found in all organisms and many viruses...
controls the folding of this ncRNA to its 'H' active form. The ribosome binding site of the alx gene is now exposed and the 30s subunit is allowed to bind.
Uses of riboregulators
Currently, riboregulators have found utility in the area of synthetic biologySynthetic biology
Synthetic biology is a new area of biological research that combines science and engineering. It encompasses a variety of different approaches, methodologies, and disciplines with a variety of definitions...
and personalized medicine
Personalized medicine
Personalized medicine is a medical model emphasizing in general the customization of healthcare, with all decisions and practices being tailored to individual patients in whatever ways possible...
. In synthetic biology, riboregulators can be used to regulate bacterial responses and probe gene regulatory networks. Undergraduate systems biology students use standardized riboregulators to compete in an annual bio-engineering competition (The international Genetically Engineered Machine competition, iGEM
IGEM
The International Genetically Engineered Machine competition is a worldwide Synthetic Biology competition aimed at undergraduate university students.- Competition details :...
). In addition, because the various riboregulators can be tailored to respond to complex biological signals, riboregulators offer the promise of individually tailored RNA based drugs which respond to genes, small molecules and proteins in individuals cells.