Compounds and methods for regulating bacterial growth and pathogenesis

1. A method for identifying a compound that regulates the activity of a non-homoserine lactone autoinducer-2 comprising:

(a) comparing the measured activity of non-homoserine lactone autoinducer-2 in the presence of the compound to the measured activity of non-homoserine lactone autoinducer-2 in the absence of the compound; and

(b) identifying the compound that regulates the activity of non-homoserine lactone autoinducer-2, wherein non-homoserine lactone autoinducer-2 is selected from the group consisting of 4-hydroxy-5-methyl-2H-furan-3- one and 2,3,4-trihydroxy-5-penten-l-one.

2. The method of claim 1 , wherein the autoinducer-2 is 4-hydroxy-5-methyl-2H-furan-3-one.

3. The method of claim 1 , wherein the autoinducer 2 is contacted with the compound in vivo.

4. The method of claim 1 , wherein the autoinducer 2 is contacted with the compound in vitro.

5. The method of claim 1 , wherein the regulation is by increasing the activity of the automducer-2.

6. The method of claim 1 , wherein the regulation is by decreasing the activity of the autoinducer-2.

7. The method of claim 1 , wherein the compound is a polypeptide.

8. The method of claim 1 , wherein the compound is a small molecule.

9. The method of claim 1 , wherein the compound is a nucleic acid.

10. A method for identifying an analog that regulates the activity of a non-homoserine lactone autoinducer-2, comprising:

(a) providing a bacterial cell that is capable of producing a detectable amount of light in response to the non-homoserine lactone autoinducer-2;

(b) contacting the bacterial cell with an analog of the non-homoserine lactone autoinducer-2; and

(c) comparing the amount of light produced by the bacterial cell in the presence and absence of the analog, wherein a change in the production of light is indicative of an analog that regulates the activity of the non-homoserine lactone autoinducer-2, wherein the non-homoserine lactone autoinducer-2 is selected from the group consisting of 4-hydroxy-5-methyl-2H-furan-3-one and 2,3,4-trihydroxy -5-penten-l-one.

11. The method of claim 10 , wherein the bacterial cell contains non-homoserine lactone autoinducer-2 that is endogenous non-homoserine lactone auto inducer-2.

12. The method of claim 10 , wherein the bacterial cell is also contacted with non-homoserine lactone autoinducer-2 that is synthesized in a bacterial cell.

13. The method of claim 10 , wherein the bacterial cell is also contacted with non-homoserine lactone autoinducer-2 that is exogenous autoinducer-2.

14. The method of claim 10 , wherein the contacting is in vitro.

15. The method of claim 10 , wherein the contacting is in vivo.

16. The method of claim 10 , further comprising contacting the bacterial cell with the non-homoserine lactone autoinducer-2.

17. The method of claim 10 , wherein the regulation is by inhibition of non-homoserine lactone autoinducer-2 activity.

18. The method of claim 10 , wherein the regulation is by enhancement of non-homoserine lactone autoinducer-2 activity.

19. The method of claim 10 , wherein the bacterial cell further comprises at least one alteration in a gene locus that participates in an autoinducer pathway, wherein the alteration inhibits the production or detection of an autoinducer.

20. The method of claim 19 , wherein the alteration in a gene locus comprises an alteration in the LuxS gene.

21. The method of claim 19 , wherein the alteration in a gene locus comprises an alteration in the LuxN gene.

22. The method of claim 19 , wherein the alteration in a gene locus inhibits detection of autoinducer-1.

23. The method of claim 19 , wherein the alteration is in the LuxN and LuxS loci.

24. The method of claim 19 , wherein the bacterial cell is V. harveyi strain MM32 (ATCC access No. BAA-1121).

25. A method for identifying a compound that regulates the production or activity of non-homoserine lactone autoinducer-2, comprising:

contacting a bacterial cell that produces non-homoserine lactone autoinducer-2 with the compound, and

determining whether non-homoserine lactone autoinducer-2 activity is present in the bacterial cell, wherein non-homoserine lactone autoinducer-2 is selected from the group consisting of 4-hydroxy-5-methyl-2H-furan-3-one and 2,3,4-trihydroxy-5-penten-l-one.

26. The method of claim 25 , wherein non-homoserine lactone autoinducer-2 activity is determined by detecting the inhibition of non-homoserine lactone autoinducer-2 production.

27. The method of claim 25 , wherein non-homoserine lactone autoinducer-2 activity is determined by detecting a signal produced in the presence of non-homoserine lactone autoinducer-2.

28. The method of claim 27 , wherein the method detects an antagonist of non-homoserine lactone autoinducer-2.

29. The method of claim 28 , wherein the method detects a change in luminescence from a reporter bacterial strain.

30. The method of claim 29 , wherein the bacterial strain is of the genus Vibrio.

31. The method of claim 30 , wherein the bacterial strain is of the species Vibric harveyi.

32. The method of claim 31 , wherein the bacterial strain is Vibrio harveyi BB 170 (ATCC access No. BAA-1117).

33. The method of claim 31 , wherein the bacterial strain is Vibrio harveyi MM32 (ATCC access No. BAA-1121).

34. A method for detecting a non-homoserine lactone autoinducer-2-associated bacterial biomarker comprising;

(a) providing at least one bacterial cell that responds to non-homoserine lactone autoinducer-2 by generating a bacterial biomarker;

(b) contacting said at least one bacterial cell with a non-homoserine lactone autoinducer-2 molecule under conditions and for such time as to promote induction of a bacterial biomarker; and

(c) detecting the bacterial biomarker, wherein the non-homoserine lactone autoinducer-2 is selected from the group consisting of 4-hydroxy-5-methyl-2H-furan-3-one and 2,3,4-trihydroxy-5-penten-l-one.

35. A method for detecting an autoinducer-associated biomarker comprising:

(a) providing at least one cell that responds to an autoinducer by a change in a biomarker of the cell,

(b) contacting the at least one cell with an autoinducer molecule under conditions and for such time as to promote induction of a biomarker; and

(c) detecting the biomarker, wherein the autoinducer is not a homoserine lactone, wherein the autoinducer is non-homoserine lactone autoinducer-2, and wherein the non-homoserine lactone autoinducer-2 is selected from the group consisting of 4-hydroxy-5-methyl-2H-furan-3-one and 2,3,4-trihydroxy-5-penten-1-one.

36. The method of claim 35 , wherein the autoinducer-2 is 4-hydoroxy-5-methyl-2H-furan-3-one.

37. A method for identifying a compound that regulates non-homoserine lactone autoinducer-2 binding to a non-homoserine lactone autoinducer-2 receptor, comprising:

(a) contacting non-homoserine lactone autoinducer-2 and the non-homoserine lactone autoinducer-2 receptor with the compound to allow non-homoserine lactone autoinducer-2 binding to the receptor;

(b) contacting the product of (a) with a cell capable of producing light in response to non-homoserine lactone autoinducer-2 binding to the receptor; and

(c) measuring the effect of the compound on light production, wherein a change in light production in the presence of the compound, compared to light production in the absence of the compound, identifies the compound as one that regulates binding of non-homoserine lactone autoinducer-2 to receptor, wherein the non-homoserine lactone autoinducer-2 is selected from the group consisting of 4-hydroxy-5-methyl-2H-furan-3-one and 2,3,4-trihydroxy-5-penten-l-one.

38. The method of claim 37 , wherein the compound is selected from the group consisting of competitive inhibitors and suicide inhibitors.

39. The method of claim 37 , wherein the receptor is selected from the group consisting of luxP and luxN.

40. The method of claim 37 , wherein the non-homoserine lactone autoinducer-2 is allowed to form a complex with the receptor in the absence of the compound.

41. The method of claim 37 , wherein the non-homoserine lactone autoinducer-2/receptor complex is bound to a solid support medium.

42. The method of claim 41 wherein the solid support medium is selected from the group consisting of a column matrix and a microtiter dish well.

43. The method of claim 42 , wherein the non-homoserine lactone automducer-2/receptor complex is bound to a solid support medium through a linkage selected from the group consisting of amide, ester, and ether.

44. A method for identifying a compound that regulates the activity of autoinducer-2 comprising:

(a) comparing the measured activity of autoinducer-2 in the presence of the compound to the measured activity of autoinducer-2 in the absence of the compound; and

(b) identifying the compound that regulates the activity of autoinducer-2, wherein the autoinducer-2 is 4-hydroxy-5-methyl-2H-furan-3-one.

45. A method for detecting an autoinducer-associated biomarker comprising:

(a) providing at least one cell that responds to an autoinducer-2 by a change in a biomarker of the cell,

(b) contacting the at least one cell with an autoinducer-2 molecule under conditions and for such time as to promote induction of a biomarker; and

(c) detecting the biomarker, wherein the autoinducer-2 is 4-hydroxy-5-methyl-2H-furan-3-one.