CONSTITUTIVE CHIMERIC RECEPTORS, AND METHODS OF USE THEREOF

Abstract: The present invention is directed to novel chimeric constitutively active recptor proteins and DNA sequences encoding these proteins. The chimeric receptors comprise a receptor, or functional fragments thereof, operatively fused to a ligand for the receptor, or functional fragments thereof. The receptor and ligand are not normally found associated together and only associate to induce activity of the receptor. By fusing the ligand to the receptor, however, the receptor is capable of constitutive transduction of a signal and activation of a signaling pathway in the cell, whereby the cell may be induced to carry out various functions relating to the signalling pathway. A wide variety of cell-surface receptors and ligands therefor, may be employed in the present invention, wherein the receptors and ligands may be naturally occuring or synthetic. There are also provided nucleic acid constructs, vectors, cells, and transgenic animals which are capable of expressing the chimeric receptors. In addition, there are provided methods for transforming an inducible receptor into a chimeric constitutively active receptor by fusing a ligand therefor to the receptor. In addition, there are provided methods for producing such chimeric receptors, as well as methods for the use thereof, including assays to identify additional ligands for receptors and therapeutic and diagnostic applications.

Description: CONSTITUTINE CHIMERIC RECEPTORS. AND METHODS OF USE THEREOF FIELD OF INVENTION The present invention relates to novel chimeric proteins for use in modulating signal transduction. BACKGROUND Regulation of cell activities is frequently achieved through inducible receptors, which are activated (or deactivated) by the binding of a ligand to the receptor. The formation of the complex with the receptor results in a change in conformation with the receptor undergoing a change which results in a signal being transduced. In some instances, the conformational change in the receptor results in binding to other proteins, where the other proteins are activated and may carry out various functions. In some situations, the receptor is autophosphorylated or phosphorylated, resulting in a change in its activity. These events are frequently coupled with secondary messengers, such as calcium, cyclic adenosine monophosphate, cyclic guanadine monophosphate, inositol phosphate, diacylglycerol, and the like. The binding of the ligand results in a particular signal being induced. Constitutively active receptors are of theoretical, practical and physiological importance. From a theoretical viewpoint, knowledge of residues in the receptor that account for constitutive activity will further our understanding of structural determinants that determine the mechanism of action of the receptor. From a practical point of view, constitutively active receptors would prove extremely useful in screening for libraries of compounds that inactivate the receptors or that act as inverse agonists. Finally, the roles of receptor mutations that result in constitutively active receptors in disease states are of increasing interest and importance. Therefore, there is a need in the art for methods to render inducible receptors constitutively active. The resulting receptor proteins will have broad and general applicability. BRIEF DESCRIPTION OF THE INVENTION The present invention is directed to novel chimeric constitutively active receptor proteins and DNA sequences encoding these proteins. The chimeric receptors comprise a receptor, or functional fragments thereof, operatively fused to a ligand therefor, or functional fragments thereof. The receptor and ligand are not normally found associated together and only associate to induce activity of the receptor. By fusing the ligand to the receptor, however, the receptor is rendered capable of constitutive transduction of a signal and activation of signaling pathway(s), whereby the cell may be turned on to carry out various functions relating to the signaling pathway. A wide variety of receptors and ligands therefor may be employed in the practice of the present invention, wherein the receptors and ligands may be naturally occurring or synthetic. In accordance with the present invention, there are also provided nucleic acid constructs, vectors, cells, and transgenic animals which are capable of expressing the invention chimeric receptors. In addition, there are provided methods for transforming an inducible receptor into a chimeric constitutively active receptor by fusing a ligand therefor to the receptor. Also provided are methods for producing such chimeric receptors, as well as methods for the use thereof, including assays to identify additional ligands for receptors and therapeutic and diagnostic applications. BRIEF DESCRIPTION OF THE FIGURES Figure 1 provides a schematic describing a chimeric construct according to the invention in which r/h CRF(1-41) replaces the N-terminal domain of hCRF-Rl. Figure 2 presents a schematic describing a chimeric construct according to the present invention, wherein r/h CRF(l-43) is fused to hCRF-Rl via an engineered putative eighth transmembrane helix fused to the intracellular C-terminal domain of the receptor. In this chimera, the peptide portion has a free C-terminus. Figure 3 is a schematic outline of Rl and related constructs. Based on alignment and structural modeling of the transmembrane region of secretin-like receptors, transmembrane segment 1 starts at position 124 in Rl. Constructs in the lower left of the figure all have the N-terminal domain corresponding to residues 1-111 of the receptor replaced with the indicated portion of CRF and are expressed with the HA-signal peptide placed upstream of the peptide portion. The construct in the lower right has residues 1-16 of CRF inserted into the N-terminal domain of Rl between residue 28 and 29. The gray shaded dots in transmembrane segments 3 and 5, respectively, indicate residues important for binding of a Rl -specific non-peptide antagonist. Figures 4A and 4B graphically show the constitutive activation and stimulation of Rl and peptide/Rl chimeras. Figure 4A shows the level of cAMP produced by host cells in the absence (white bars) and in the presence (gray bars) of 10 μM antalarmin. Figure 4B shows the level of c AMP produced by host cells in the absence (white bars) and in the presence (black bars) of 10 μM urocortin. cAMP level is normalized in each experiment to that observed in the presence of 10 μM antalarmin. Data are presented as mean ± S.E.M. from 4-8 independent experiments each performed in triplicate. The absolute level of cAMP in presence of 10 μM antalarmin is similar for all constructs. Figures 5 A and 5B depict the dose response with urocortin and antalarmin on CRF(1-16)/R1 ΔN (Figure 5A) and CRF(1-16[L8A])/R1 ΔN (Figure 5B). Data are presented as mean ± SD from triplicate determinations and are representative of 3 independent assays performed in parallel. For CRF(1-16)/R1 Δ N, the IC50 for inhibition of constitutive activation by antalarmin is 14 ± 1 nM. For CRF(1-16[L8A])/R1 ΔN , the EC 50 for stimulation by urocortin is 140 ± 20 nM. DETAILED DESCRIPTION OF THE INVENTION In accordance with the present invention, there are provided chimeric proteins comprising receptors, or functional derivatives thereof, operatively fused to a ligand therefor, or functional derivatives thereof. By operatively fusing a receptor to its respective ligand, preferably the agonist, the receptor is thereby locked into a constitutively active state, which is distinct from basal activity. Independent of endogenous receptor agonist, constitutively activated chimeric receptor(s) of the present invention can transduce signal indefinitely. This kinetic trapping mechanism represents a novel, potent, long-lasting positive regulatory mechanism with potentially profound and wide-ranging physiological implications. Any receptor undergoing this type of constitutive activation would be expected to contribute dominantly to the overall basal tone in the body, thereby modifying tonicity caused by continuously released hormones and neuro transmitters. As contemplated herein, a number of inducible receptors may be employed in the practice of the present invnetion. Receptors contemplated for use in the practice of the present invention are readily recognized by those skilled in the art, and include those associated with signal transduction, transcription, the uptake of nutrients, cell adhesion, cell aggregation, endocytosis, and the like. In addition, receptors contemplated for use in the practice of the present invention can be cell surface associated receptors, membrane associated receptors, cytoplasmic receptors, nuclear receptors, combinations thereof, and the like. Nuclear receptors include members of the steroid/thyroid hormone receptor superfamily, and the like. Cell surface associated receptors are of particular interest including those which may be involved with one or more second messenger pathways, particularly pathways involved with a protein kinase. Cell surface receptors include ion channel receptors, G-protein coupled receptors, receptors with single transmembrane segments or tyrosine kinase- containing receptor, and the like, as well as functional derivatives thereof. Ion channel receptors contemplated for use in the practice of the present invention include nicotinic, NMDA and non-NMDA, GABA, 5-HT, and the like. G protein-coupled receptors contemplated for use in the practice of the present invention include all subtypes of the opioid, muscarinic, dopamine, adrenergic, cAMP, opsins, angiotensin, serotonin, thyrotropin, gonadotropin, substance-K, substance-P and substance-R receptors, melanocortin, metabotropic glutamate, corticotropin-releasing factor receptors (CRF-R), or any other GPCR receptors known to couple via G proteins (e.g., the secretin-like family of GPCRs, also designated the class 2 or class B receptor family, including receptors for secretin, calcitonin, gastric inhibitory peptide, growth hormone-releasing hormone, glucagon, glucagon-like peptide I, parathyroid hormone (PTH), pituitary adenylate cyclase-activating polypeptide, vasoactive intestinal polypeptide, and the like). Receptors with single transmembrane segments contemplated for use in the practice of the present invention include growth factor receptors, insulin, cytokine, natriureticm, and the like. Other cell surface receptors contemplated for use in the practice of the present invention will be readily recognized by those of skill in the art (see, e.g., Lauffenburger and Linderman, Receptors : Models for Binding, Trafficking, and Signaling (1996), Williams et al., Receptor Pharmacology and Function (1998)). Ligands contemplated for use in the practice of the present invention will depend on the specific receptor employed and can be readily identified by those of skill in the art. Ligands contemplated include naturally-occurring, semi-synthetic and synthetic agonists and antagonists, and modifiers of the activity of agonists and antagonists which are capable of inducing their respective receptor to transduce signals within the cell. For example, ligands contemplated for use in the practice of the present invention, include natural ligands (e.g., neurotransmitters, growth factors, hormones, steroids, autoacoids, chemotactic factors, exogenous stimulants such as odorants, cytokines, modifications thereof, and the like), recombinant peptides, peptidomimetics, antibodies or fragments thereof, synthetic molecules (e.g., drug, or any other agent which is capable of inducing a signal), and the like. Additional ligands which can be employed in the practice of the present invention will be readily apparent to those skilled in the art. As employed herein, the phrase "functional derivative" of either protein or nucleic acid, is a molecule that possesses a biological activity (either functional or structural) that is substantially similar to a biological activity of the non-derivatized (i.e., parental) protein or nucleic acid sequence. Thus, for example, a functional derivative of a receptor will possess the ability to transduce signals, whereas a functional derivative of a ligand will be able to either inhibit or induce signal transduction upon association with the receptor. A functional derivative of a protein can contain post-translational modifications such as covalently linked carbohydrate, or the like, depending on the necessity of such modifications for the performance of a specific function. The term "functional derivative" is intended to include mutants, fragments, segments, variants, analogs, or chemical derivatives of a molecule. The term "functional derivative" also is intended to include chimeric combinations of one or more receptors, i.e., recombinant receptors comprising domains from different receptors (e.g., transmembrane domains from different receptors, DNA binding domain swaps, and the like). In a preferred embodiment of the present invention, the chimeric receptor will comprise only those portions of a cell surface receptor and ligand therefor necessary for signal transduction. In a presently most preferred embodiment of the present invention, when the chimeric is derived from a cell surface receptor, the chimeric receptor will comprise at least one transmembrane domain and the cytoplasmic domain of a cell surface receptor operatively fused to a portion of an agonist therefor. As used herein, a molecule is said to be a "chemical derivative" of another molecule when it contains additional chemical moieties not normally a part of the molecule. Such moieties can improve the molecule's solubility, absorption, biological half life, and the like. The moieties can alternatively decrease the toxicity of the molecule, eliminate or attenuate any undesirable side effect(s) of the molecule, and the like. Procedures for coupling such moieties to a molecule are well known in the art. The term "operatively fused" in the context of the present invention refers to any type of covalent or non-covalent binding, linkage, fusion or association, including chemical and biological fusions, wherein the components so described are in a relationship permitting them to function in their intended manner. Such fusion, performed to join together components of desired functions to generate a desired combination of functions, i.e., constitutive activity, includes covalent bonding, hydrophobic/hydrophilic interaction, Van der Walls forces, ion pairing, ligand- receptor interaction, epitope-antibody binding site interaction, enzyme-substrate interaction, liposome-hydrophobic interaction, nucleotide base pairing, membrane- hydrophobic interaction, and the like. Preferably, the fusion does not contemplate the endogenous (i.e., normal) interaction of receptor and ligand, but instead contemplates a permanent interaction typically associated with pre- and post-translational fusions, such as recombinant constructs, and the like. In a presently preferred embodiment of the present invention, ligand, or functional fragment thereof, is fused to transmembrane or extracellular domain(s) of its cognate receptor. As readily recognized by those of skill in the art, ligand(s) can be introduced into receptor(s) in a variety of locations, e.g., within and/or attached to at least one intracellular domain (i.e., hinge domain, DNA binding domain, cytoplasmic domain, and the like), within and/or attached to at least one transmembrane domain of the receptor, within and/or attached to an exogenous transmembrane domain introduced to a chimeric receptor, within and/or attached to at least one extracellular domain at the amino terminus and/or carboxyl terminus of the domain, and/or inserted therebetween, and the like. As employed herein, the term "introduced" refers to the addition, insertion, replacement, substitution, and the like, of the ligand, or derivatives thereof, or alternatively, the nucleic acid encoding the ligand, preferably the open reading frame, to the receptor, or functional derivatives thereof. The term "recombinant" polynucleotide or nucleic acid refers to one which is not naturally occurring, or is made by the artificial combination of two otherwise non- contiguous segments of sequence. This artificial combination is often accomplished by either chemical synthesis means, or by the artificial manipulation of isolated segments of nucleic acids, e.g., by genetic engineering techniques. Generally, DNA sequences encoding the structural coding sequence of a gene product can be assembled from cDNA fragments and short oligonucleotide linkers, or from a series of oligonucleotides, to provide a synthetic gene which is capable of being expressed in a recombinant transcriptional unit. Such sequences are preferably provided in the form of an open reading frame uninterrupted by internal nontranslated sequences, or introns, which are typically present in eukaryotic genes. Genomic DNA containing the relevant sequences may also be used. Sequences of non-translated DNA may be present 5' or 3' from the open reading frame, where such sequences do not interfere with manipulation or expression of the coding regions. In yet another presently preferred embodiment of the present invention, a chimeric receptor can be further modified to facilitate signal transduction and/or protein orientation. For example, normally, the signal sequence at the 5' terminus of the open reading frame (ORF) which directs the chimeric protein to the surface membrane will be the signal sequence of the extracellular domain. However, in some instances, one may wish to exchange this sequence for a different signal sequence. In most cases, associated with the signal sequence will be a naturally occurring cleavage site, which will also typically be the naturally occurring cleavage site associated with the signal sequence or the extracellular domain. Such modifications to facilitate signal transduction can be introduced either pre- or post-translational, and may be placed upstream or downstream of the ligand in order to insure insertion of the chimeric receptor in the plasma membrane of the cell (e.g., HA-signal peptide, and the like). Additional modifications which will facilitate chimeric receptor orientation, function, stability, and the like, are known to those skilled in the art. For example, the constructs may be designed so as to avoid their interaction with other surface membrane proteins native to the target host. Thus, for the most part, one will avoid the chimeric receptor binding to other proteins present in the surface membrane. In order to achieve this, one may select a transmembrane domain which is known not to bind to other transmembrane domains, one may modify specific amino acids, e.g. substitute for a cysteine residue, or the like. The present invention also provides DNA molecules encoding the chimeric proteins of the present invention. Invention DNA molecules generally comprise a DNA sequence encoding the chimeric protein, and associated regulatory sequences such as transcription promoter, a transcription terminator, and the like. In general, prokaryotic expression vectors such as plasmid vectors containing replication and control sequences which are compatible with the host cells are used as cloning vectors for the DNA molecules of the present invention. Other vectors, such as λ-phage, cosmids, or yeast artificial chromosomes may also be employed in the practice of the present invention. The vector ordinarily carries a replication site, as well as sequences which encode proteins that are capable of providing phenotypic selection of transformed cells. In addition, promoters have been discovered and utilized, and details concerning their nucleotide sequences have been published, enabling a skilled worker to ligate them functionally into the chosen vector. The promoters are operably linked to a nucleic acid sequence encoding the chimeric protein. The promoters may be inducible or constitutive and provide a means to express the encoded chimeric protein in the host. Following expression, the polypeptide may be purified by standard methods such as described below. Alternatively, a DNA sequence encoding the chimeric proteins of the present invention may be inserted into a suitable eukaryotic expression vector, which in turn is used to transfect eukaryotic cells. A eukaryotic expression vector, as used herein, is meant to indicate a DNA construct containing regulatory elements which direct the transcription and translation of DNA sequences encoding invention chimeric proteins. Such regulatory elements include promoters, enhancers, transcription terminators and polyadenylation signals. By virtue of the inclusion of these elements operably linked within the DNA constructs, the resulting eukaryotic expression vectors contain the information necessary for expression of the polypeptides of interest. In a preferred embodiment of the present invention, the vector comprises nucleic acid encoding a chimeric receptor operatively linked to an inducible promoter, thereby providing for inducible expression of the chimeric receptor in host cells. Suitable vectors will generally include a selectable marker, which may be one of any number of genes that exhibit a dominant phenotype for which a phenotypic assay exists to enable transformants to be selected. Preferred selectable markers are those that complement host cell auxotrophy, provide antibiotic resistance or enable a cell to utilize specific carbon sources. Choice of a particular host and selectable marker is well within the level of ordinary skill in the art. Any cell line can be used as a suitable "host" in the practice of the present invention. Host cells contemplated for use in expressing recombinant chimeric proteins of interest include mammalian cells, avian cells, insect cells, fungal cells, and the like. Thus, cells contemplated for use in the practice of the present invention include transformed cells, non-transformed cells, neoplastic cells, primary cultures of different cell types, and the like. Examples of fungal cells contemplated for use in the practice of the present invention include species of yeast (a.g., Saccharomyces spp., Schizosaccharomyces spp.), filamentous fungi (e.g., Aspergillus spp., Neurospora spp.), and the like. Cultured mammalian cells may be used as host cells in the practice of the present invention. Cultured mammalian cells contemplated for use herein include human monocytoid, lymphocytoid, fibroblastoid cell lines, and the like. A useful mammalian cell line is the HeLa-tat cells that are HeLa derived cells. Mammalian expression vectors contemplated for use in carrying out the present invention include a promoter capable of directing the transcription of a cloned gene or cDNA. Preferred promoters include viral promoters and cellular promoters. Other promoters will be readily recognized by those skilled in the art. Such mammalian expression vectors may also contain a set of RNA splice sites located downstream from the promoter and upstream from the DNA sequence encoding the polypeptide or protein of interest. Preferred RNA splice sites may be obtained from adenovirus and/or immunoglobulin genes. Also contained in the expression vectors may be a polyadenylation signal located downstream of (i.e., 3' to) the coding sequence of interest. Cloned DNA sequences may be introduced into cultured mammalian cells by methods well known in the art, including, for example, calcium phosphate-mediated transfection, electroporation, protoplast fusion, biolistics, using DNA-coated particles, transfection, infection (where the chimeric construct is introduced into an appropriate virus, particularly a non-replicative form of the virus), and the like. In order to identify cells that have integrated the cloned DNA, a selectable marker is generally introduced into the cells along with the gene or cDNA of interest. Preferred selectable markers for use in cultured mammalian cells include genes that confer resistance to drugs, such as neomycin, hygromycin, methotrexate, and the like. The selectable marker may be an amplifiable selectable marker such as the DHFR gene, or the like. Selectable markers are reviewed by Thilly (Mammalian Cell Technology, Butterworth Publishers, Stoneham, Mass., which is incorporated herein by reference). The choice of selectable markers is well within the level of ordinary skill in the art. In accordance with another aspect of the present invention, there are provided methods for producing chimeric receptors according to the present invention, i.e., converting inducible receptors into constitutively active receptors. Methods for making chimeric receptors include chemical synthesis, recombinant nucleic acid technology (including viral and non-viral vectors), purification from natural sources (and thereafter manipulation), or any other methods which may be used to make chimeric polypeptides. Standard reference works setting forth the general principles of recombinant DNA technology include Watson, J. D. et al., Molecular Biology of the Gene, Volumes I and II, The Benjamin/Cummings Publishing Company, Inc., publisher, Menlo Park, Calif. (1987); Darnell, J. E. et al., Molecular Cell Biology, Scientific American Books, Inc., publisher, New York, NY. (1986); Lewin, B. M., Genes II, John Wiley & Sons, publishers, New York, NY. (1985); Old, R. W., et al., Principles of Gene Manipulation: An Introduction to Genetic Engineering, 2d edition, University of California Press, publisher, Berkeley, Calif. (1981); Sambrook, J. et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Second Edition, Cold Spring Harbor, NY. (1989); and Ausubel et al Current Protocols in Molecular Biology, Wiley Interscience, NY., (1987, 1993). These references are incorporated herein by reference in their entirety. In accordance with yet another embodiment of the present invention, there are provided methods for treating and diagnosing diseases and disorders associated with non-normal, i.e., constitutive or insufficient, signal transduction, including neurological disorders such as Parkinsonism, Alzheimers, depression, and the like, diabetes, growth abnormalities, cancer therapy, surgery adjuvants, and the like. In a particular aspect of the present invention, therapeutic methods are provided wherein tissue or cells from a subject are transformed with nucleic acid capable of expressing at least one chimeric receptor. Other means for modulating signal transduction employing chimeric receptors will be readily recognized by those skilled in the art. In a presently preferred embodiment of the present invention, chimeric receptors are produced by transformation of host cells from a given individual with retroviral vector constructs directing the synthesis of the chimeric construct. By transformation of such cells and reintroduction of the cells into the patient one may achieve autologous gene therapy applications. In accordance with another embodiment of the present invention, there are provided screening assays for identifying modulators of inducible receptors. In a preferred embodiment of the present invention, use of screening assays permits the identification of agonists, neutral antagonists, negative antagonists, and receptor inhibitors capable of reducing the constitutively active state of invention chimeric receptors. Such methods comprise determining the effect of test compounds on chimeric receptor activity upon exposure of chimeric receptors to a library of compounds. Thus, compounds which increase the level of signal transduction are identified as agonists and compounds which decrease the level of signal transduction are identified as antagonists. Two classes of ligands have been identified, particularly with respect to antagonists, i.e., neutral antagonists which block only agonist induced effects without changing basal activity, and inverse agonists, or negative antagonists, which also block basal receptor activity. Agonists which also increase the signal transduction activity irrespective of whether a ligand is pre-associated with the receptor can also be identified, and are contemplated as within the scope of the present invention. Any synthetic, semi-synthetic or naturally-occurring compound can be evaluated, including further evaluating known ligands for cell-surface receptors. Small molecules (e.g., Antalarmin, Astressin, and the like) are included among the compounds which can be evaluated and/or identified by employing invention receptors in assays described herein. Any reporter can be employed which will facilitate identification of the level of signal transduction, e.g., by monitoring the level of secondary messengers. Additional parameters which can be monitored will be readily recognized by those of skill in the art. In addition, other means for identifying ligands for receptors with constitutive activity will be readily apparent to those employing the present invention (see, e.g., Sadee et al., U.S. Patent No. 5,882,944, the entire contents which are hereby incorporated by reference). Accordingly, the practice of the present invention is useful to determine or to screen for new pharmaceuticals useful for treating disease states mediated by receptors capable of displaying constitutive activity, to enhance the clinical utility of existing pharmaceuticals targeted to receptors, to devise therapeutic treatments from agents identified by the screening methods of the invention, and the like. Particularly, compounds identified according to the present invention are useful in the treatment of diseases and disorders associated with prolonged agonist exposure, such as narcotic addiction, drug resistance, and the like. In accordance with another embodiment of the present invention, there are provided methods for studying the interaction between receptors and their respective ligands employing chimeric receptors of the present invention. Any method, including three dimensional modeling, two dimensional modeling, protein-protein interaction studies, and any other method can be employed to study the interaction between the ligand component and the ligand binding domain component of a chimeric receptor. Chimeric receptors of the present invention are suitable for such studies because of the binding stability associated with the ligand and the ligand binding domain. The present invention is also directed to a transgenic non-human eukaryotic animal (preferably a rodent, such as, but not limited to, a mouse), the germ cells and/or somatic cells of which contain nucleic acid according to the present invention which codes for chimeric receptor. The nucleic acid encoding chimeric receptor according to the present invention is introduced into the animal to be made transgenic, or an ancestor of the animal, at an embryonic stage, preferably at the one-cell, or fertilized oocyte stage, and generally not later than about the 8-cell stage. The term "transgene," as used herein, means a gene which is incorporated into the genome of the animal and is expressed in the animal, resulting in the presence of protein in the transgenic animal. There are several means known to those of skill in the art by which such a gene can be introduced into the genome of the animal embryo so as to be chromosomally incorporated and expressed. Chimeric non-human mammals in which fewer than all of the somatic and germ cells contain nucleic acid encoding chimeric receptor are also provided by the present invention. Such animals are produced when fewer than all of the cells of the morula are transfected in the process of producing the transgenic mammal. Chimeric non-human mammals having human cells or tissue engrafted therein are also encompassed by the present invention. Such chimeras can be used for testing expression of chimeric receptors in human tissue and/or for testing the effectiveness of therapeutic and/or diagnostic agents associated with delivery vectors which preferentially bind to chimeric receptors of the present invention. Methods for providing chimeric non-human mammals will be readily apparent to those skilled in the art employing the present invention. The techniques described in Leder, U.S. Pat. No. 4,736,866 (hereby incorporated by reference in its entirety) for producing transgenic non-human mammals may be used for the production of transgenic non-human mammals of the present invention. The various techniques described in Palmiter, R. et al., Ann. Rev. Genet. 20:465-99 (1986), the entire contents of which are hereby incorporated by reference, may also be used. Animals carrying nucleic acid encoding invention chimeric receptor(s) can be used to test compounds or other treatment modalities which may prevent, suppress or cure a human disease associated with non-normal, i.e., constitutive or insufficient, signal transduction. Such animals can also serve as a model for testing of diagnostic methods for the same human pathologies or diseases. Such pathologies or diseases include cancer, neurological diseases, and the like. Transgenic or chimeric animals according to the present invention can also be used as a source of cells for cell culture. The invention will now be described in greater detail by reference to the following non-limiting examples. EXAMPLE 1 CONSTITUTIVELY ACTIVE CRF RECEPTOR CHIMERAS In a first approach, a CRF receptor (see, e.g., Perrin et al., U.S. Patent No. 5,728,545) is made constitutively active by constructing, for example, a chimera in which all or a fragment of r/h CRF replaces the first extracellular domain (i.e., N- terminus) of the CRF receptor. An optimized signal peptide (e.g. the HA-signal peptide) which is cleaved post-translationally, is placed upstream of the entire construct in order to insure insertion of the chimeric protein in the cell's plasma membrane. This approach ensures a free N-terminus for the peptide (see Figure 1). A cDNA encoding human CRF-R 1 (hereinafter "Rl")(Chen, R., Lewis, K. A., Perrin, M. H., and Vale, W. W. (1993) Proc. Natl. Acad. Sci. USA 90, 8967-8971) was subcloned into the pCI vector (Promega), positioning an optimized Kozak sequence upstream of the initiation codon (Kozak, M. (1987) J. Mol. Biol. 196(4), 947-50). Silent Mlul- and BspEI restriction sites were created at position 285 and position 450, respectively, and the endogenous BspEI site at position 1121 was removed. All constructs were made by a modified overlap-extension PCR protocol (Horton, R. M., Hunt, H. D., Ho, S. N., Pullen, J. K., and Pease, L. R. (1989) Gene 77(1), 61-8) using flanking primers with a 5' add-on sequence (Stappert, j., Wirsching, J., and Kemler, R. (1992) Nucleic Acids Res 20(3), 624). All PCR products were cloned as EcoRI/BspEI fragments into the pCI vector containing Rl except for the CRF(1-16)/R1 chimera which was cloned as a EcoRI/MluI fragment. For R1 Δ N, residues 1-111 were replaced by the HA-signal peptide (Guan, X., Kobilka, TS. and Kobilka, BK. (1992) J. Biol. Chem. 267, 21995-21998) followed by the FLAG epitope. The chimeras CRF(l-16)/RlΛN and CRF( 17-41)/R1 ΔN have residues 1-111 replaced by the HA-signal peptide followed by the indicated portions of rat/human CRF (Vaughan, J., Donaldson, C, Bittencourt, J., Perrin, M. H., Lewis, K., Sutton, S., Chan, R., Turnbull, A. V., Lovejoy, D., Rivier, C, Rivier, j., Sawchenko, P. E., and Vale, W. (1995) Nature 378, 287-292). Chimera CRF(1-16)/R1 has the c-myc epitope and a glycine residue separating it from CRF(1- 16) (EQKLISEEDLGSEEPPISLDLTFHLLR) (SEQ ID NO: 8) inserted between residue 28 and 29 of Rl. All chimeras were identified by restriction enzyme digestion and verified by automatic sequencing. To conduct functional assays, one day prior to transfection, 2 x 10 6 COS M6 cells were plated in a 10 cm dish (Falcon) and transfected using the DEAE-dextran method (Perrin, M. H., Sutton, S., Bain, D. L., Berggren, W. T., and Vale, W. W. (1998) Endocrinology 139(2), 566-570). Approximately 20 hours later, the cells were trypsinized (Sigma) and seeded into 48 well (Costar) plates at a density of 2 x 10 4 cells/well. The cells were assayed approximately 40-48 hours after transfection. Cells were washed twice with DMEM containing 0.1 % fetal calf serum and incubated for 2 hours in 200 ul of this media in a 37 °C humidified atmosphere containing 5 % CO2. Cells were subsequently preincubated for 15 minutes with antagonist before IB MX was added (final concentration 0.2 mM). 10 minutes later agonists were added and the assay was stopped 15 minutes after that by aspiration of media and addition of 0.5 ml of ice-cold ethanol containing 0.1 N HC1. The cAMP content was determined by RIA (Biomed. Tech.). The results of this assay are presented in Table 1 and discussed below. Unstimulated Stimulated EC50 Fold (n) Fold (n) nM (n) Rl 1.3 ± 0.2 (8) 91.4 ± 11.1 (8) 0.21 ± 0.04 (4) CRF(1-16)/R1ΔN 22.9 ± 2.7 (8) 27.9 ± 3.0 (8) ~ (5) CRF(17-41)/R1 ΔN 1.2 ± 0.1 (7) 50.6 ± 6.6 (7) 110 ± 25 (5) CRF(1-16)/R1 1.1 ± 0.2 (7) 75.6 ± 4.5 (7) 0.20 ± 0.06 (5) Table 1. Summarized basal activity and potency of urocortin on Rl and peptide/Rl chimeras. Data are presented as mean ± S.E.M from (n) independent experiments each performed in triplicate. The cAMP level is normalized in each experiment to that observed in the presence of 10 μM antalarmin. Because of the low level of urocortin stimulation of CRF(1 -16)/R1 Δ N, the EC50 could not be determined. Chimeras were designed between Rl and the amino-terminal residues (1 to 16) or the carboxy-terminal residues (17 to 41) of CRF, and the CRF peptide portion positioned in place of the N-terminal domain of the receptor (Fig 3). Because these chimeras lack the signal peptide of the receptor, they are expressed using the HA- signal peptide derived from influenza hemaglutinin and introduced at the amino- termini of the chimeric receptors. The introduction of the HA-signal peptide ensures proper membrane targeting of the expressed constructs. Furthermore, the HA-signal peptide is cleaved by the expressing cells, leaving the CRF peptide with a free amino- terminus, and tethered at its carboxy end to the transmembrane region of the receptor. Transient transfection of COS-M6 cells with the peptide/receptor chimera in which the first 16 residues of CRF replace the receptor's N-terminal domain (CRF(1- 16)/R1 ΔN ) results in high levels of receptor activity (Fig. 4A). The activity of this chimera is -20 fold (Fig. 4A and Table 1) higher than the activity observed in the presence of 10 μM antalarmin (a Rl-specific non-peptide antagonist (Webster, W. L., Lewis, D. B., Torpy, D. J., Zachman, E. K., Rice, K. C, and Chrousos, G. P. (1996) Endocrinology 137, 5747-5750)) and is blocked in a dose-dependent manner by antalarmin (Fig 5 A). By contrast, the constitutive activation is not inhibited by 10 μM of the peptide antagonist, astressin (Gulyas, J., Rivier, C, Perrin, M., Koerber, S. C, Sutton, S., Corrigan, A., Lahrichi, S. L., Craig, A. G., Vale, W., and Rivier, J. (1995) Proc. Natl. Acad. Sci. USA 92, 10575-10579 ). The transmembrane region of this chimera is not responsible for the constitutive activation because the N-terminally truncated receptor, R1 Δ N > (Fig 3) does not display constitutive activity. This observation indicates that the N-terminal domain of Rl does not constrain the transmembrane region of the receptor in an inactive conformation as has been proposed for the TSH receptor. Urocortin, another mammalian member of the CRF- like family of peptides, activates R1 Δ N wi h an EC50 = 0.1 μM, which is a ~500-fold reduction in potency compared to that of the native receptor. This reduction in potency probably reflects the distinct, yet overlapping, receptor regions that contribute to the urocortin-receptor interaction. The N-terminal receptor domain is involved in relatively high affinity binding, whereas the body of the receptor appears to display a lower affinity interaction. Urocortin treatment of cells expressing the chimera CRF(1-16)/R1 ΔN results in a minor, but not statistically significant, further stimulation of cAMP production (Figs. 4B, 5B and Table 1). Using ELIS A, antibodies raised against CRF( 1 -21 ) as well as CRF( 1-41) detect high levels of epitope on cells transfected with CRF(1-16)/R1 ΔN . Therefore, the low level of stimulation of CRF(1-16)/R1 ΔN by urocortin is not due to an absence of membrane localized constructs. Mutational analysis of Rl has revealed two residues, namely His 199 and Met276 in the transmembrane segments 3 and 5, respectively, which affect the binding of NBI 27914, another non-peptide antagonist (Liaw, C. W., Grigoriadis, D. E., Lorang, M. T., De Souza, E. B., and Maki, R. A. (1997) Mol. Endocrinol. 11, 2048-2053), functionally similar to antalarmin. It is likely that similar segments are involved in the binding of antalarmin. The fact that antalarmin is able to inhibit the constitutive activation of CRF(1-16)/R1 Δ N is consistent with this assumption. The chimera in which the carboxy-terminal residues (17-41) of CRF replace the N-terminal domain of Rl, (CRF(17-41)/R1 ΔN ), is deficient in constitutive activation (Fig 4A and Table 1), yet produces a -50 fold (Fig 4B and Table 1) in response to 10 μM urocortin. These results are consistent with the amino-terminal portion of the CRF peptide being required for activation. The significance of the proximity between CRF(1-16) and the transmembrane region of the receptor was examined by inserting CRF(1-16) into the N-terminal domain of Rl between residues 28 and 29 within the intact receptor (Fig 3). This chimera, CRF(1-16)/R1, does not display constitutive activity (Fig 4 A and Table 1), but shows a large response to urocortin with similar potency EC50 - 0.2 nM to that of Rl (Fig 4B and Table 1). For CRF(1-16)/R1, both an anti-myc antibody, as well as the above-mentioned CRF antibodies, detect high expression levels of chimeras on cells transfected with this construct, indicating that the 16 residues of CRF are present in CRF(1-16)/R1. Therefore, the lack of constitutive activation of this chimera is not due to loss of the peptide during expression. The lack of constitutive activation of this construct is likely due to the ability of the N-terminal domain of the receptor to function as a spacer and thus diminish the proximity between the active portion of CRF and the body of the receptor. An analog of CRF in which Leu-8 is replaced by Ala retains full intrinsic activity, but has a 300-fold decrease in the relative potency compared to that of CRF as measured by in vitro ACTH release (Kornreich, W. D., Galyean, R., Hernandez, J.- F., Craig, A. G., Donaldson, C. J., Yamamoto, G., Rivier, C, Vale, W., and Rivier, J. (1992) J. Med. Chem.35, 1870-1876). Similar results for cAMP production are found for Rl . The corresponding modification in the peptide portion of the chimera, CRF(1-16[L8A])/R1 ΔN , abolishes the constitutive activation (Fig. 5B). This chimera, however, can be stimulated by urocortin (Fig. 5B). Using ELISA with antibodies raised against CRF(1-21) and CRF(1-41), the level of expression of CRF(1- 16[L8A])/R1 ΔN is similar to that of CRF(1-16)/R1 ΔN . These results indicate that the constitutive activation of the CRF(1-16)/R1 Δ N chimera is produced by specific interactions between the tethered amino-terminal part of CRF and the body of the receptor and that the specificity is reminiscent of that between CRF and native Rl. The present approach of tethering a ligand to the receptor will likely restrict the chimera to the same signaling pathways as the native receptor in responding to the ligand. This makes these novel ligand-dependent constitutively activated chimeras appropriate for transgenic mice using tissue-specific conditional expression ( see, e.g., Redfern, C. H., Coward, P., Degtyarev, M. Y., Lee, E. K., Kwa, A. T., Hennighausen, L., Bujard, H., Fishman, G. I., and Conklin, B. R. (1999) Nature Biotechnol. 17(2), 165-9) for the study of the pathophysiology associated with a specific receptor system. The constitutively activated chimera CRF(1- 16)/R1 Δ N is a new type of RASSL (receptor activated solely by a synthetic ligand). This chimera does not respond to endogenous levels of agonist like a RASSL. However, in contrast to a RASSL, which is only activated by administering a synthetic agonist, the CRF(1- 16)/R1 ΔN chimera will be constitutively activated upon expression. The activity of this chimera can be pharmacologically inhibited by an orally active, Rl -specific, non- peptide antagonist like antalarmin. This type of engineered chimera may be designated as a RISSL (receptor inhibited solely by a synthetic ligand). Example 2 ADDITIONAL CONSTITUTIVELY ACTIVE RECEPTOR CHIMERAS In a second approach, a constitutively active CRF receptor is a chimera in which the r/h CRF (1-41) (or a fragment thereof) is positioned downstream of an engineered eighth transmembrane helix fused to the intracellular C-terminal domain of the receptor. In this chimera, the peptide portion has a free C-terminus. The rat/human CRF, human urocortin, and astressin were kindly provided by Dr. J. Rivier (Salk Institute for Biological Studies). The non-peptide antagonist Antalarmin was kindly provided by Dr. G. Chrousos (NIH). The cDNAs encoding the human CRF-R 1 and CRF-R2αI were cloned by RT- PCR from human mRNA (Clontech) and inserted into the pCI vector as EcoRi/Xbal fragments. Both cDNAs contain a Kozak sequence upstream the initiating ATG codon. PCR was used to generate the desired fragments using Pfu polymerase (Stratagene) in a slightly modified version of methods previously described (Horton, R. M., Hunt, H. D., Ho, S. N., Pullen, J. K., and Pease, L. R. (1989) Gene 77(1), 61- 8). The generated fragment was inserted into the pCI expression vector containing appropriate parts of the wildtype receptor. All fragments generated by PCR, were verified by fluorescent dideoxynucleotide sequencing (Perkin Elmer). The constructs with an engineered eighth transmembrane helix consisting of 24 leucines fused to the C-terminal domain of the receptor (pSIS 180, 181, 222, 225, described below) were designed using a BamHI site which was introduced at the end of the C-terminus of the wildtype receptor in place of the stop-codon. This site was subsequently used to insert a BamHI Xbal fragment encoding residues as described below. For the hCRFl-TM8-CRFl-43 construct (pSIS 181), the BamHI Xbal fragment encoded the following 90 residues: GDPKKLLLLLLLLLLLLLLLLLLLLLLLLSNTSSNTSDDDDKSKLGGSEEPPISL DLTFIILLRHVLEMARAEQLAQQ AHSNRKLMEIIGK. (SEQ ID NO : 1 ) For the hCRFl-TM8 construct (pSIS 180), the BamHI/Xbal fragment encoded the following 47 residues: GDPKKLLLLLLLLLLLLLLLLLLLLLLLLSNTSSNTSDDDDKSDLGG. (SEQ ID NO:2) For the hCRFl-TM8-CRF 17-43 construct (pSIS 222), the BamHI/Xbal fragment encoded the following 74 residues: GDPKKLLLLLLLLLLLLLLLLLLLLLLLLSNTSSNTSDDDDKSDLGGEVLEMA RAEQLAQQAHSNRKLMEIIGK. (SEQ ID NO:3) For the hCRFl-TM8-glucagon construct (pSIS 225), the BamHI/Xbal fragment encoded the following 76 residues: GDPKKLLLLLLLLLLLLLLLLLLLLLLLLSNTSSNTSDDDDKSDLGGHSQGTFT SDYSKYLDSRRAQDFVQWLMNT (SEQ ID NO:4) The N-terminally modified constructs were designed by introducing the described residues followed by residues 112-415 in the hCR-Rl. Thus, residues 1- 111 in the wildtype receptor are deleted in these constructs. In the HA-myc-CRFl-41-[l 12-415] hCRFl (pSIS 215), the following residues were encoded instead of residue 1-111 in the wildtype receptor: MKTIIALSYLFCLVFADAEQKLISEEDLGSEEPPISLDLTFHLLREVLEMARAE QLAQQAHSNRKLMEII. (SEQ ID NO:5) In the HA-CRF1-41-[112-415] hCRFl (pSIS 216), the following residues were encoded instead of residue 1-111 in the wildtype receptor: MKTIIALSYIFCLVFASEEPPISLDLTFHLLREVLEMARAEQLAQQAHSNRKLM EII. (SEQ ID NO:6) In the HA-CRF1-41-[112-415] hCRFl (pSIS 217), the following residues were encoded instead of residue 1-11 1 in the wildtype receptor: MKTIIALSYIFCLVFASEEPPISLDLTFHLLR (SEQ ID NO:7) The purified plasmids (Qiagen maxi-prep) were transiently transfected into COS-M6 cells using the DEAE-dextran method as previously described (Perrin et al., U.S. Patent No. 5,728,545). The cells were grown in a humidified atmosphere with 5% CO 2 in, Dulbecco's modified Eagle's medium 1885, supplemented with 10% fetal calf serum, 2 mM glutamine and 0.2 mg/ml gentamicin. The day following the transfection, the cells were harvested, seeded into wells and 50 nM (final) of the PKA inhibitor H-89 (Calbiochem) was added. The next day, H-89 was added again two hours before the cAMP assay which was performed as previously described (Perrin et al.), except that cells were pre-treated with IB MX (Sigma) for 10 minutes before test compounds were applied. Incubation was continued for another 10 min., and the cells were extracted with 0.1 N HCL/95% EtOH and assayed for cAMP using a kit (Biome. Tech.) for before incubation with ligands for 10 minutes. The results of the cAMP assays are set out in Tables 2 A and 2B and show cAMP ratios for N-terminal modified (Table 2A) or C-terminal modified (Table 2B) hCRFl receptor constructs. Values are calculated as mean (+/-) s.e.m. For each construct, the calculated cAMP level with 1 uM Antalarmin added is defined as 1 in each experiment and all other cAMP levels are shown as ratio of this level. The number of independent experiments is indicated by (n). Table 2A Table 2B ro These results indicate that invention receptors having ligand replacing the N terminal 111 amino acids of the receptor have greater constitutive activation than if ligand is fused to the C terminus of the receptor. Each of the receptors remains responsive to natural ligands therefor (e.g., CRF and urocortin). The data further indicate that small molecule antagonists effectively inhibit the constitutive activity of the receptors, thereby demonstrating the utility of such receptors in assaying for antagonists of receptor function. While the invention has been described in detail with reference to certain preferred embodiments thereof, it will be understood that modifications and variations are within the spirit and scope of that which is described and claimed.

Claims: That which is claimed is: I. A constitutively active chimeric peptide comprising: domain(s) from at least one receptor, operatively fused to a ligand therefor, or functional fragment thereof. 2. A chimeric peptide according to claim 1, wherein at least one domain is responsive to an agonist or functional fragment thereof. 3. A chimeric peptide according to claim 2, wherein said agonist is a ligand for said receptor. 4. A chimeric peptide according to claim 3, wherein said receptor is a CRF receptor and said agonist is CRF or urocortin. 5. A chimeric peptide according to claim 4, wherein said CRF receptor is CRF-R1 and said functional fragment of CRF is amino acids 1-16 of SEQ ID NO:9. 6. A chimeric peptide according to claim 5, wherein said functional fragment of CRF replaces amino acids 1-111 of SEQ ID NO: 10. 7. A chimeric peptide according to claim 6, wherein said functional fragment of CRF has fused at an amino terminal end a hemaglutinin signal peptide. 8. A chimeric peptide according to claim 1, wherein said ligand is fused to the ligand binding domain of said receptor. 9. A chimeric peptide according to claim 8, wherein said ligand is CRF and said receptor is CRF-R 1. 10. A chimeric peptide according to claim 1, wherein said ligand, or functional fragment thereof, is positioned at the amino-terminus of said receptor. I I . A chimeric peptide according to claim 10, wherein said ligand is CRF and said receptor is CRF-R1. 12. A chimeric peptide according to claim 1, wherein said ligand, or functional fragment thereof, replaces the amino-terminal portion of said receptor. 13. A chimeric peptide according to claim 12, wherein said ligand is CRF and said receptor is CRF-R 1. 14. A chimeric peptide according to claim 1, wherein said ligand, or functional fragment thereof, is positioned at the carboxy-terminus of said receptor. 15. A chimeric peptide according to claim 1, wherein said ligand, or functional fragment thereof, is attached to said receptor through an engineered additional transmembrane helix. 16. A chimeric peptide according to claim 15, wherein said engineered additional transmembrane helix is attached at the carboxyl terminus of said receptor and comprises the amino acids of SEQ ID NO: l, SEQ ID NO:2, SEQ ID NO:3, or SEQ ID NO:4. 17. A chimeric peptide according to claim 16, wherein said receptor is a CRF receptor. 18. A chimeric peptide according to claim 17, wherein said CRF receptor is hCRF-Rl or hCRF-R2αl. 19. A chimeric peptide according to claim 15, wherein said wherein said engineered additional transmembrane helix is attached at the amino terminus of said receptor and comprises the amino acids of SEQ ID NO:5, SEQ ID NO:6, or SEQ ID NO:7. 20. A chimeric peptide according to claim 19, wherein said receptor is a CRF receptor. 21. A chimeric peptide according to claim 20, wherein said CRF receptor is hCRF-Rl or hCRF-R2αl. 22. A chimeric peptide according to claim 20, wherein said engineered transmembrane helix region replaces amino acids 1-1 1 1 of said CRF receptor. 23. A chimeric peptide according to claim 1, wherein said agonist, or functional fragment thereof, is inserted somewhere in the open reading frame of the receptor. 24. A chimeric peptide according to claim 1 , wherein said receptor is a cell surface receptor. 25. A nucleic acid encoding the chimeric peptide of claim 1. 26. A nucleic acid according to claim 25, further comprising an inducible response element that regulates transcription or translation of said chimeric peptide. 27. A method for producing a chimeric peptide wherein said method comprises expressing the nucleic acid of claim 26 in a host cell. 28. A method for converting an inducible receptor into a constitutively active receptor, said method comprising fusing a ligand, or functional fragment thereof, to said receptor. 29. A method according to claim 28, wherein at least one domain is responsive to an agonist or a functional fragment thereof. 30. A method according to claim 29, wherein said agonist is a ligand for said receptor. 31. A method according to claim 30, wherein said receptor is a CRF receptor and said agonist is CRF or urocortin. 32. A method according to claim 31, wherein said CRF receptor is CRF-Rl and said functional fragment of CRF is amino acids 1-16 of SEQ ID NO:9. 33. A method according to claim 32, wherein said functional fragment of CRF replaces amino acids 1-111 of SEQ ID NO: 10. 34. A method according to claim 33, wherein said functional fragment of CRF has fused at an amino terminal end a hemaglutinin signal peptide. 35. A method according to claim 28, wherein said ligand is fused to the ligand binding domain of said receptor. 36. A method according to claim 35, wherein said ligand is CRF and said receptor is CRF-Rl. 37. A method according to claim 28, wherein said ligand, or functional fragment thereof, is positioned at the amino-terminus of said receptor. 38. A method according to claim 37, wherein said ligand is CRF and said receptor is CRF-Rl. 39. A method according to claim 38, wherein said ligand, or functional fragment thereof, replaces the amino-terminal portion of said receptor. 40. A method according to claim 39, wherein said ligand is CRF and said receptor is CRF-Rl. 41. A method according to claim 28, wherein said ligand, or functional fragment thereof, is positioned at the carboxy-terminus of said receptor. 42. A method according to claim 28, wherein said ligand, or functional fragment thereof, is attached to said receptor through an engineered additional transmembrane helix. 43. A method according to claim 42, wherein said engineered additional transmembrane helix is attached at the carboxyl terminus of said receptor and comprises the amino acids of SEQ ID NO: l, SEQ ID NO:2, SEQ ID NO:3, or SEQ ID NO:4. 44. A method according to claim 43, wherein said receptor is a CRF receptor. 45. A method according to claim 44, wherein said CRF receptor is hCRF-Rl or hCRF-R2αl. 46. A method according to claim 42, wherein said wherein said engineered additional transmembrane helix is attached at the amino terminus of said receptor and comprises the amino acids of SEQ ID NO:5, SEQ ID NO:6, or SEQ ID NO:7. 47. A method according to claim 46, wherein said receptor is a CRF receptor. 48. A method according to claim 47, wherein said CRF receptor is hCRF-Rl or hCRF-R2αl. 49. A method according to claim 47, wherein said engineered transmembrane helix region replaces amino acids 1-111 of said CRF receptor. 50. A method according to claim 29, wherein said agonist, or functional fragment thereof, is inserted somewhere in the open reading frame of the receptor. 51. A method according to claim 28, wherein said receptor further comprises a post- translationally cleavable signal sequence. 52. A method according to claim 51 , wherein said post translationally cleavable sequence is a hemmaglutinin signal peptide. 53. A method for screening test compounds to identify modulators for receptors which are responsive to agonists, said method comprising: A) contacting a constitutively active peptide according to claim 1 with test compound, and B) determining the effect of test compound on receptor activity, wherein: an increase in signal transduction indicates that said test compound is an agonist, and a decrease in signal transduction indicates that said test compound is an antagonist. 54. A method for screening for peptide mutations that result in a peptide antagonist or a peptide with superagonist activity, said method comprising, A) mutating the chimeric peptide of claim 1 , and B) determining the effect of the mutation on receptor activity, wherein: an increase in signal transduction indicates that said mutation produces a peptide with superagonist activity, and a decrease in signal transduction indicates that said mutation produces a peptide with antagonist activity. 55. A method for studying the three-dimensional interaction of ligands and ligand binding domains of receptors, said method comprising evaluating the interaction between the ligand and the receptor of the chimeric receptor of claim 1. 56. A method for treating subjects affected by non-normal levels of signal transduction, said method comprising fransfecting said subjects with nucleic acid encoding the chimeric receptor according to claim 1. 57. A transgenic animal comprising the chimeric receptor of claim 1.

Inventors: NIELSEN LISA Z, NIELSEN SOREN M, PERRIN MARILYN H, VALE WYLIE W, NIELSEN, LISA, Z., NIELSEN, SOREN, M., PERRIN, MARILYN, H., VALE, WYLIE, W.
Applicants: NIELSEN LISA Z, NIELSEN SOREN M, PERRIN MARILYN H, SALK INST FOR BIOLOGICAL STUDI, VALE WYLIE W, NIELSEN, LISA, Z., NIELSEN, SOREN, M., PERRIN, MARILYN, H., THE SALK INSTITUTE FOR BIOLOGICAL STUDIES, VALE, WYLIE, W.
Application year: 2000
Main classification: C07K14
IPC codes: A61K48, C07K14, C07K19