Cell death regulators
AssigneeWashington University
InventorStanley J. Korsmeyer
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Abstract
The invention provides a bcl-2 related protein, bcl-2 muteins, two-hybrid systems comprising interacting bcl-2-related polypeptide sequences, and uses thereof.
Claims12
A composition comprising
- (1) a first polynucleotide sequence encoding a first hybrid polypeptide comprising an activator domain of a transcriptional activator protein and a first bcl-2 family member having a BH1 domain and a BH2 domain,
- (2) a second polynucleotide sequence encoding a second hybrid polypeptide comprising a DNA-binding domain of said transcriptional activator protein and a second bcl-2 family member having a BH1 domain and a BH2 domain, and
- (3) a third polynucleotide sequence comprising a reporter gene linked to a transcriptional regulatory element whose transcriptional activity is dependent upon the presence or absence of a dimer comprised of the first and second hybrid polypeptides, wherein said first and second bcl-2 family members are independently selected from the group consisting of naturally occurring Bcl-2, Bcl-x.sub.L, Bax, Mcl-1, A1, fragments thereof, and mutants thereof having a mutation in the BH1 domain and/or the BH2 domain which alters intermolecular binding of said first and second bcl-2 family members.
A kit comprising a two-hybrid system, a host cell and an instruction manual, said two-hybrid system having
- (1) a first polynucleotide sequence encoding a first hybrid polypeptide comprising an activator domain of a transcriptional activator protein and a first bcl-2 family member having a BH1 domain and a BH2 domain, and
- (2) a second polynucleotide sequence encoding a second hybrid polypeptide comprising a DNA-binding domain of said transcriptional activator protein and a second bcl-2 family member having a BH1 domain and a BH2 domain, wherein said first and second bcl-2 family members are independently selected from the group consisting of naturally occurring Bcl-2, Bcl-x.sub.L, Bax, Mcl-1, A1, fragments thereof, and mutants thereof having a mutation in the BH1 domain and/or the BH2 domain which alters intermolecular binding of said first and second bcl-2 family members.
A yeast cell comprising
- (1) a first polynucleotide sequence encoding a first hybrid polypeptide comprising an activator domain of a transcriptional activator protein and a first bcl-2 family member having a BH1 domain and a BH2 domain,
- (2) a second polynucleotide sequence encoding a second hybrid polypeptide comprising a DNA-binding domain of said transcriptional activator protein and a second bcl-2 family member having a BH1 domain and a BH2 domain, and
- (3) a third polynucleotide sequence comprising a reporter gene linked to a transcriptional regulatory element whose transcriptional activity is dependent upon the presence or absence of a dimer comprised of the first and second hybrid polypeptides, wherein said first and second bcl-2 family members are independently selected from the group consisting of naturally occurring Bcl-2, Bcl-x.sub.L, Bax, Mcl-1, A1, fragments thereof, and mutants thereof having a mutation in the BH1 domain and/or the BH2 domain which alters intermolecular binding of said first and second bcl-2 family members.
A substantially pure composition comprising a bcl-2 family member polypeptide, a naturally occurring full-length Bax polypeptide, and an antibody which binds to said Bax polypeptide, wherein said bcl-2 family member polypeptide has a BH1 and a BH2 domain and is selected from the group consisting of naturally occurring Bcl-2, Bcl-x.sub.L, Bax, Mcl-1, A1, fragments thereof, and mutants thereof having a mutation in the BH1 domain and/or the BH2 domain which alters binding of said bcl-2 family member to said Bax polypeptide.
Dependencies are inferred from the claim text.