Receptors: their identification, characterization, preparation and use
AssigneeThe Salk Institute for Biological Studies
InventorsRonald M. Evans, Stanley M. Hollenberg
Life of this patent
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Abstract
Disclosed are novel hormone and hormone-like receptors Wherein the trans-activation transcription domain(s) is modified in terms of position and/or copy number or otherwise versus parent receptor, such novel receptors having increased trans-activation transcription activation properties surprisingly superior to the parent receptor. Also disclosed are the recombinant methods and means for preparing such receptors and assays based upon the use of such receptors for screening and identifying putative materials that can affect such receptors and/or for the expression via induced transcription of a reporter or other desired, preferably heterologous gene or DNA product.
Claims13
A DNA molecule encoding a member of the steroid hormone superfamily of receptors having trans-activation transcription domains in addition to those of the parental receptor,
- wherein said additional trans-activation transcription domains are selected from the group consisting of .tau..sub.1, .tau..sub.2, and functional fragment(s) thereof,
- wherein said additional trans-activation transcription domains are independent of both the DNA-binding and ligand-binding domains of parent receptor, and
- wherein said additional trans-activation transcription domains are located outside of said DNA-binding and ligand-binding domains, if present.
A process of preparing a human receptor which is a member of the steroid hormone superfamily of receptors having trans-activation transcription domains in addition to those of the parental receptor,
- wherein said additional trans-activation transcription domains are selected from the group consisting of .tau..sub.1, .tau..sub.2, and functional fragment(s) thereof,
- wherein said additional trans-activation transcription domains are independent of both the DNA-binding and ligand-binding domains of parent receptor, and
- wherein said additional trans-activation transcription domains are located outside of said DNA-binding and ligand-binding domains, if present,
- said process comprises expressing transfected DNA encoding said receptor in a host cell.
In a bioassay for determining the functionality of a hormone or hormone-like receptor, said bioassay comprising:
- (a) transfecting into receptor negative cells one or more expression vectors harboring an operative hormone responsive promoter/enhancer element functionally linked to an operative reporter or other desired DNA, and a construct containing an operative DNA sequence encoding said receptor,
- (b) culturing said transfected cells from step (a) in the presence or absence of extrinsic material having the putative ability to activate said hormone responsive promoter/enhancer element,
- (c) monitoring in said cells induction of the product of said reporter or desired DNA sequence, and
- (d) measuring in said cells the expression of reporter or other desired DNA, the improvement wherein said hormone or hormone-like receptor has increased trans-activation transcription activity compared with parent receptor, said increased activity due to additional transactivation transcription domains, relative to the parent receptor, wherein said domains are located said DNA-binding and ligand-binding domain of said receptor.
A DNA molecule encoding a member of the steroid hormone superfamily of receptors having trans-activation transcription activity, relative to trans-activation transcription activity of parental receptor,
- wherein said member of the steroid hormone superfamily of receptors contains a plurality of at least one parental receptor trans-activation transcription domain selected from the group consisting of .tau..sub.1, .tau..sub.2, and functional fragment(s) thereof, located outside of said receptor.
A process of preparing a human receptor which is a member of the steroid hormone superfamily of receptors having increased trans-activation transcription activity, relative to trans-activation transcription activity of parental receptor,
- wherein said member of the steroid hormone superfamily of receptors contains a plurality of at least one parental receptor tarns-activation transcription domain selected from the group consisting of .tau..sub.1, .tau..sub.2, and functional fragment(s) thereof, located outside of said DNA-binding and ligand-binding domains of said receptor,
- said process comprises expressing transfected DNA encoding said receptor in a host cell.
Dependencies are inferred from the claim text.