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US 8,301,387 B2Granted
Combinatorial transcription control for programmed genetic response
AssigneeThe Regents of the University of California
InventorsTerence Tai-Li Hwa, Ulrich Gerland, Nicolas Buchler
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Abstract
A scheme is provided for selecting the regulatory DNA sequences to exert combinatorial control of gene transcription in vivo by multiple regulatory proteins, i.e., transcription factors (TFs). The method provides the ability to implement a wide range of complex logic functions through the manipulation of the regulatory DNA sequences, typically several hundred bases in length. The method includes procedures for selecting the strengths of the binding sites of different TFs and their relative positioning in order to implement a plurality of different logic functions.
Claims36
A method for combinatorially controlling the transcription of a target gene in a cell for producing a programmed response in the cell, the method comprising:
- identifying a plurality of logic functions, wherein each logic function combines two or more different inputs to generate an output comprising a unique gene expression in the cell;
- implementing at least one selected logic function that produces a desired genetic response when a combination of specified conditions are present, wherein the two or more different inputs comprise two or more regulatory proteins and the at least one selected logic function is implemented by producing interactions between the two or more regulatory proteins and interactive binding of the two or more regulatory proteins at DNA binding sites of the target gene to control a probability of promoter occupancy, wherein the target gene comprises one or more cis-regulatory regions having individual DNA binding sites, and wherein each DNA binding site has a binding strength and a binding location which are adjustable by varying composition of the one or more cis-regulatory regions to select one of a cooperative binding and competitive binding of the two or more regulatory proteins at the binding sites and one of a cooperative binding and competitive binding between the two or more regulatory proteins and polymerase;
- wherein the interactions comprise contact interactions and long-distance interactions that avoid interference between the DNA binding sites;
- wherein the desired genetic response in the cell comprises reporting or treatment of a disease or condition in which the two or more regulatory proteins are expressed;
- wherein each step of the method is executed on a transcription apparatus which corresponds to a computing machine;
- wherein the probability of promoter occupancy is determined according to the relationship Ppo=ZON/(ZON+ZOFF), where ZON and ZOFF are the total statistical weight of all states in which the promoter is occupied and unoccupied, respectively;
- wherein when the selected logic function is AND, ZON=qo(1+ωqA+qB+2ω2qAqB) and ZOFF=1+qA+qB+ωqAqB;
- wherein when the selected logic function is OR, ZON=qo(1+ωqA+ωqB+2ωqAqB) and ZOFF=1+qA+qB+qAqB;
- wherein when the selected logic function is XOR, ZON=qo(1+ωqA+ωqB+2ωqAqB) and ZOFF=(1+qA+qB+qAqB)×(1+qA+qB+ωqAqB);
- and
- wherein when the selected logic function is NAND, ZON=qo and ZOFF=1+qA+qB+ωqAqB;
- where qo is the Boltzmann weight for promoter occupancy, qA and qB are ratios of the regulatory protein concentrations to the binding thresholds for input A and input B, respectively, and w is the mutual cooperativity factor.
9 more elements
A method for combinatorial transcription control for controlling gene expression in a cell for producing a programmed response in the cell, the method comprising:
- identifying a plurality of logic functions, each logic function having two or more different inputs and an output comprising a unique and artificial genetic response;
- selecting at least one logic function that produces a desired genetic response;
- and
- implementing the at least one logic function by producing interactions among different inputs comprising two or more transcription factors and controlling interactive binding of the two or more transcription factors at corresponding binding sites of one or more target genes, wherein the target genes each comprise one or more cis-regulatory regions having individual DNA binding sites, and wherein each binding site has a binding strength and a binding location which are adjustable by varying composition of the one or more cis-regulatory regions to select one of a cooperative interaction and competitive interaction between the two or more transcription factors and to select cooperative or competitive binding of between the two or more transcription factors and polymerase, all to control a probability of promoter occupancy;
- generating the desired genetic response in the cell as the output, wherein the desired genetic response in the cell comprises reporting or treatment of a disease or condition in which the two or more transcription factors are
- expressed;
- wherein each step of the method is executed on a transcription apparatus which corresponds to a computing machine;
- wherein the probability of promoter occupancy is determined according to the relationship Ppo=ZON/(ZON+ZOFF), where ZON and ZOFF are the total statistical weight of all states in which the promoter is occupied and unoccupied, respectively;
- wherein when the selected logic function is AND, ZON=qo(1+ωqA+qB+2ω2qAqB) and ZOFF=1+qA+qB+ωqAqB;
- wherein when the selected logic function is OR, ZON=qo(1+ωqA+ωqB+2ωqAqB) and ZOFF=1+qA+qB+qAqB;
- wherein when the selected logic function is XOR, ZON=qo(1+ωqA+ωqB+2ωqAqB) and ZOFF=(1+qA+qB+qAqB)×(1+qA+qB+ωqAqB);
- and
- wherein when the selected logic function is NAND, ZON=qo and ZOFF=1+qA+qB+ωqAqB;
- where qo is the Boltzmann weight for promoter occupancy, qA and qB are ratios of the regulatory protein concentrations to the binding thresholds for input A and input B, respectively, and w is the mutual cooperativity factor.
11 more elements
A method of combinatorially controlling gene expression in a cell by encoding control functions in regulatory DNA sequences for producing a programmed response in the cell, the method comprising:
- selecting a relative binding strength and a relative binding position of individual DNA binding sites within a cis-regulatory region of one or more regulatory DNA sequences to exert combinatorial control of gene expression to operate as at least one logic function selected from a plurality of different logic functions for combining two or more inputs, each input comprising a different regulatory protein, to generate an output comprising a programmed gene expression upon DNA binding of two or more different regulatory proteins as determined by the selected relative binding strength and relative binding position of the DNA binding sites to control probability of promoter occupancy, wherein each different logic function produces a different programmed gene expression in the cell;
- and
- wherein the programmed gene expression in the cell comprises reporting or treatment of a disease or condition in which the two or more different regulatory proteins are expressed.
A method for combinatorial control of gene expression to produce a programmed genetic response in a cell, the method comprising:
- producing a plurality of different glue-like contact interactions between two or more transcription factors at DNA binding sites of genes for selectively activating or repressing expression in a target gene under specified conditions that comprise to inputs into one or more logic functions, wherein the one or more logic functions are selected according to a relationship between the two or more transcription factors and a disease or condition, wherein each DNA binding site has a protein-DNA binding affinity and a binding location that are variable by varying a cis-regulatory sequence at the DNA binding site to select a predetermined combination of degrees of freedom selected from the group consisting of DNA binding thresholds for each transcription factor, Boltzmann weight for promoter occupancy, and mutual cooperativity factors between the transcription factors and RNA polymerase, wherein the predetermined combination of degrees of freedom controls a probability of promoter occupancy to produce a desired genetic response in the cell;
- and
- wherein the desired genetic response in the cell comprises reporting or treatment of the disease or condition in which the two or more transcription factors are
- expressed;
- wherein each step of the method is executed on a transcription apparatus which corresponds to a computing machine;
- wherein the probability of promoter occupancy is determined according to the relationship Ppo=ZON/(ZON+ZOFF), where ZON and ZOFF are the total statistical weight of all states in which the promoter is occupied and unoccupied, respectively;
- wherein when the selected logic function is AND, ZON=qo(1+ωqA+qB+2ω2qAqB) and ZOFF=1+qA+qB+ωqAqB;
- wherein when the selected logic function is OR, ZON=qo(1+ωqA+ωqB+2ωqAqB) and ZOFF=1+qA+qB+qAqB;
- wherein when the selected logic function is XOR, ZON=qo(1+ωqA+ωqB+2ωqAqB) and ZOFF=(1+qA+qB+qAqB)×(1+qA+qB+ωqAqB);
- and
- wherein when the selected logic function is NAND, ZON=qo and ZOFF=1+qA+qB+ωqAqB;
- where qo is the Boltzmann weight for promoter occupancy, qA and qB are ratios of the regulatory protein concentrations to the binding thresholds for input A and input B, respectively, and w is the mutual cooperativity factor.
9 more elements
Dependencies are inferred from the claim text.