Biological circuit chemotactic converters

Patent metadata

Field	Value
Publication number	US-9284562-B2
Application number	US-201013512453-A
Country	US
Kind code	B2
Filing date	Nov 30, 2010
Priority date	Nov 30, 2009
Publication date	Mar 15, 2016
Grant date	Mar 15, 2016

Abstract

Described herein are novel biological circuit chemotactic converter that utilize modular components, such as genetic toggle switches and single invertase memory modules (SIMMs), for detecting and converting external inputs, such as chemoattractants, into outputs that allow for autonomous chemotaxis in cellular systems. Flexibility in these biological circuit chemotactic converter is provided by combining individual modular components, i.e., SIMMs and genetic toggle switches, together. These biological converter switches can be combined in a variety of network topologies to create network systems that regulate chemotactic responses based on the combination and nature of input signals received.

First claim

We claim:

1. A biological circuit chemotactic converter composition comprising:
at least one input module (IM) n nucleic acid comprising an inducible promoter nucleic acid sequence (iP A ) operably linked to a repressor nucleic acid sequence encoding a repressor protein (R A );
at least one genetic toggle switch (T S ) n nucleic acid comprising a first repressible promoter nucleic acid sequence (rP 1 ) that drives expression of a second repressor nucleic acid sequence (R 2 ) encoding a second repressor protein, and a second repressible promoter nucleic acid sequence (rP 2 ) that drives expression of a first repressor nucleic acid sequence (R 1 ) encoding a first repressor protein, (rP 1 -R 2 and rP 2 -R 1 ), wherein the rP 1 is inhibited by the first repressor protein and the rP 2 is inhibited by the second repressor protein and wherein R A of at least one input module acts as a repressor for rP 1 of at least one genetic toggle switch;
at least one logic module (L M ) n nucleic acid consisting essentially of a repressible promoter nucleic acid sequence (rP B ) operably linked to a repressor nucleic acid sequence (R B ), wherein rP B is inhibited by the second repressor protein of at least one genetic toggle switch; and
at least two sensor module (SM A and SM B ) n+1 nucleic acids, wherein each of the two sensor module nucleic acids comprises a repressible promoter nucleic acid sequence (rP C and rP D , respectively) operably linked to a nucleic acid sequence encoding a sensor molecule (sensor A and sensor B) and wherein n ≧1; wherein the repressible promoter nucleic acid sequence of at least one sensor module is repressed by the repressor protein encoded by at least one logic module, and the repressible promoter nucleic acid sequence of another of the at least two sensor modules is directly repressed by the second repressor protein encoded by at least one genetic toggle switch; and wherein if no input signal is received by the at least one input module nucleic acids, only sensor A is expressed, and wherein if an input signal is received by at least one input module nucleic acid, then sensor A is not expressed; and
wherein the repressor proteins encoded by any of the logic module nucleic acid sequences are different from the repressor proteins encoded by any of the at least one input module nucleic acid sequences and are different from the repressor proteins encoded by any of the at least one genetic toggle switch nucleic acid sequences.

2. The biological circuit chemotactic converter composition of claim 1 , wherein the inducible promoter nucleic acid sequence of the input module is induced by a biological agent, a chemical agent, a metal ion, a toxin, light or a pollutant.

3. The biological circuit chemotactic converter composition of claim 1 , wherein the repressor protein encoded by the first repressor nucleic acid sequence of the toggle switch (R 1 ) and the repressor protein encoded by the input module (R A ) nucleic acid sequence are the same repressor protein.

4. The biological circuit chemotactic converter composition of claim 1 , wherein the second repressible promoter nucleic acid sequence of the genetic toggle switch (rP 2 ), the repressible promoter nucleic acid sequence of the logic module (rP B ), and the repressible promoter nucleic acid sequence of one sensor module (rP D ) are repressed by the same repressor protein.

5. The biological circuit chemotactic converter composition of claim 1 , wherein the second repressible promoter nucleic acid sequence of the genetic toggle switch (rP 2 ), the repressible promoter nucleic acid sequence of the logic module (rP B ), and the repressible promoter nucleic acid sequence of one sensor module (rP D ) comprise the same repressible promoter nucleic acid sequence.

6. The biological circuit chemotactic converter composition of claim 1 , wherein the repressible promoter nucleic acid sequence of one sensor module (rP C ) is repressed by the repressor encoded by the logic module (R B ) nucleic acid sequence.

7. The biological circuit chemotactic converter composition of claim 1 , wherein the at least two sensor module nucleic acids encode for different sensor molecules.

8. The biological circuit chemotactic converter composition of claim 1 , wherein n is an integer value between and including 1 and 100.

9. The biological circuit chemotactic converter composition of claim 1 , further comprising at least one ribosome binding nucleic acid sequence.

10. The biological circuit chemotactic converter of claim 1 , further comprising at least one terminator nucleic acid sequence.

11. The biological circuit chemotactic converter composition of claim 1 , further comprising at least one degradation nucleic acid tag sequence.

12. The biological circuit chemotactic converter of composition claim 1 , further comprising at least one nucleic acid sequence encoding an iRNA molecule specific for at least one protein encoded by the biological circuit chemotactic converter.

13. The biological circuit chemotactic converter composition of claim 1 , wherein at least one repressor protein is an engineered zinc-finger protein.

14. The biological circuit chemotactic converter composition of claim 1 , where the promoter nucleic acid sequence of the inducible and repressible promoters is selected from any of the promoter sequences of SEQ ID NOs: 1-7, SEQ ID NOs: 167-843, and SEQ ID NOs: 1009-1010.

15. The biological circuit chemotactic converter composition of claim 1 , where the sensor molecule encoded by the nucleic acid sequence of the sensor domain is a chemotaxis receptor or chemotaxis sensor.

16. An isolated cell comprising the biological circuit chemotactic converter nucleic acid composition of claim 1 .

17. The cell of claim 16 , wherein the cell is an artificial cell, a prokaryotic cell, or a eukaryotic cell.

18. The cell of claim 16 comprising two or more biological circuit chemotactic converter nucleic acid compositions of claim 1 acting together.

19. A vector comprising the biological circuit chemotactic converter nucleic acid composition of claim 1 .

20. A biological circuit chemotactic converter composition comprising the circuit depicted in FIG. 1B , the content of which is herein incorporated by reference in its entirety.

Assignees

• Collins James J
• Lu Timothy Kuan-Ta
• Univ Boston
• Massachusetts Inst Technology

Inventors

• Collins James J
• Lu Timothy Kuan-Ta

Classifications

• C12N15/63 Primary

  Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression · CPC title

• C12N15/635

  Externally inducible repressor mediated regulation of gene expression, e.g. tetR inducible by tetracyline · CPC title