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At least 505 records · Page 28Linked to original sources
PubMed2000

Human keratinocytes constitutively produce but do not process interleukin-18.

BACKGROUND: Interleukin (IL)-18 is a potent immunomodulatory cytokine which promotes T-helper (Th) 1 and cytotoxic responses. IL-18 signals through a two-chain receptor (IL-18R and accessory protein-like subunit, AcPL), and an inhibitory molecule, IL-18 binding protein (IL-18BP), has recently been characterized. OBJECTIVES: The aim of the present study was to define the production of IL-18 and its receptor by human keratinocytes. METHODS: The presence of IL-18 was determined using polymerase chain reaction in human keratinocyte cultures with or without treatment with potential inducers. RESULTS: The IL-18 gene was constitutively transcribed by primary human keratinocytes and cell lines and was not significantly altered following exposure to IL-1 beta, tumour necrosis factor-alpha, interferon (IFN)-gamma, phorbol myristate acetate or nickel sulphate. IL-18 protein was constitutively present at high levels in keratinocyte lysates and was detectable in supernatants exclusively in the unprocessed, 24-kDa form. Cytokine exposure failed to induce any change in protein levels or processing. Primary keratinocytes produced IL-18R and AcPL constitutively at the mRNA level, in addition to low levels of IL-18BP, which was transcriptionally inducible following treatment with IFN-gamma. CONCLUSIONS: These findings demonstrate that IL-18 is constitutively synthesized by human keratinocytes and is released in an unprocessed form in vitro. Release of IL-18 by human keratinocytes may permit them to regulate IFN-gamma production during cutaneous inflammatory responses and suggests that IL-18 may represent an attractive target for immunomodulatory intervention in Th1-mediated inflammatory diseases such as psoriasis.

Cell Culture Techniques↗
PubMed2003

Constitutive oligomerization of human D2 dopamine receptors expressed in Spodoptera frugiperda 9 (Sf9) and in HEK293 cells. Analysis using co-immunoprecipitation and time-resolved fluorescence resonance energy transfer.

Human D2Long (D2L) and D2Short (D2S) dopamine receptor isoforms were modified at their N-terminus by the addition of a human immunodeficiency virus (HIV) or a FLAG epitope tag. The receptors were then expressed in Spodoptera frugiperda 9 (Sf9) cells using the baculovirus system, and their oligomerization was investigated by means of co-immunoprecipitation and time-resolved fluorescence resonance energy transfer (FRET). [3H]Spiperone labelled D2 receptors in membranes prepared from Sf9 cells expressing epitope-tagged D2L or D2S receptors, with a pKd value of approximately 10. Co-immunoprecipitation using antibodies specific for the tags showed constitutive homo-oligomerization of D2L and D2S receptors in Sf9 cells. When the FLAG-tagged D2S and HIV-tagged D2L receptors were co-expressed, co-immunoprecipitation showed that the two isoforms can also form hetero-oligomers in Sf9 cells. Time-resolved FRET with europium and XL665-labelled antibodies was applied to whole Sf9 cells and to membranes from Sf9 cells expressing epitope-tagged D2 receptors. In both cases, constitutive homo-oligomers were revealed for D2L and D2S isoforms. Time-resolved FRET also revealed constitutive homo-oligomers in HEK293 cells expressing FLAG-tagged D2S receptors. The D2 receptor ligands dopamine, R-(-)propylnorapomorphine, and raclopride did not affect oligomerization of D2L and D2S in Sf9 and HEK293 cells. Human D2 dopamine receptors can therefore form constitutive oligomers in Sf9 cells and in HEK293 cells that can be detected by different approaches, and D2 oligomerization in these cells is not regulated by ligands.

Animals↗
PubMed2002

p75 neurotrophin receptor is required for constitutive and NGF-induced survival signalling in PC12 cells and rat hippocampal neurones.

We have previously shown that nerve growth factor (NGF)-induced activation of nuclear factor-kappaB increased neuronal expression of Bcl-xL, an anti-apoptotic Bcl-2 family protein. In the present study we determined the role of the p75 neurotrophin receptor in constitutive and NGF-induced survival signalling. Treatment of rat pheochromocytoma (PC12) cells with a blocking anti-rat p75 antibody or inhibition of p75 expression by antisense oligonucleotides reduced constitutive and NGF-induced bcl-xL expression. Treatment with the blocking anti-p75 antibody also inhibited NGF-induced activation of the survival kinase Akt. Inhibition of phosphatidylinositol-3-kinase (PI3 kinase) activity or overexpression of a dominant-negative mutant of Akt kinase inhibited NGF-induced nuclear factor-kappaB activation. Activation of Akt kinase by NGF was also observed in PC12nnr5 cells and cultured rat hippocampal neurones which both lack significant TrkA expression. Treatment of hippocampal neurones with the blocking anti-p75 antibody inhibited constitutive and NGF-induced Bcl-xL expression, activation of Akt, and blocked the protective effect of NGF against excitotoxic and apoptotic injury. Our data suggest that the p75 neurotrophin receptor mediates constitutive and NGF-induced survival signalling in PC12 cells and hippocampal neurones, and that these effects are mediated via the PI3-kinase pathway.

Animals↗
PubMed2001

The Constitutional Type Questionnaire: validation in the patient population of the Royal London Homoeopathic Hospital.

In homeopathy the choice of a medicine is based on the total 'picture' presented by the patient. This picture includes 'constitutional type' which comprises personality, and general physical features. The Constitutional Type Questionnaire (CTQ) is designed to systematically assess constitutional types. This study examines the reliability and validity of the CTQ. Four hundred and seventy-two outpatients attending clinics at the Royal London Homoeopathic Hospital completed the CTQ, a 152-item scale rating features traditionally considered typical of 19 constitutional homeopathic medicine 'pictures' on 5-point frequency or severity scales. A subsample was retested after 1 week to measure the test-retest reliability. Another subsample was prescribed a medicine by a homeopathic doctor. Prescriptions were compared with the CTQ, to assess the content validity of the scale. The construct validity was measured by Grade of Membership (GOM) analysis. The scale demonstrated good test-retest reliability (r=0.73), internal consistency (r=0.95). The correlation between CTQ results and the medicine prescribed by the homeopathic doctor was 75.8%. The GOM analyses are reported by Davidson et al elsewhere in this issue of the journal. Although the CTQ could be improved, the scale displays good reliability and validity.

Homeopathy↗
PubMed1996

Constitutively activating TSH receptor mutations as the cause of toxic thyroid adenoma, multinodular toxic goiter and autosomal dominant non autoimmune hyperthyroidism.

cAMP stimulates both the growth and differentiated thyroid function of the thyroid gland. In toxic nodules the clinical observation of hyperthyroidism together with TSH independent growth of the hot nodule suggests a chronic activation of the cAMP cascade. Somatic mutations in a gene of the cAMP regulatory cascade leading to constitutive activation of this cascade in toxic nodules were first detected in the G protein Gs alpha. Thereafter constitutively activating TSH receptor mutations were identified in 20-80% of toxic thyroid nodules and in 3 of 6 toxic multinodular goiters. Constitutively activating TSH receptor germline mutations are expected to lead to toxic hyperplasia. Sequencing of the TSH receptor gene in families with hereditary non autoimmune hyperthyroidism led to the identification of constitutively activating TSH receptor mutations in 7 families. Moreover, sporadic TSH receptor germline mutations have been identified in 3 children with severe congenital nonautoimmune hyperthyroidism. Therefore, in cases of clustering of non autoimmune hyperthyroidism in families and in cases of sporadic congenital hyperthyroidism with thyroid hyperplasia and no evidence for an autoimmune etiology a search for TSH receptor gene mutations is necessary to appropriately direct the therapy of these patients.

Adenoma↗
PubMed2004

Dynamers: polyacylhydrazone reversible covalent polymers, component exchange, and constitutional diversity.

Component exchange in reversible polymers allows the generation of dynamic constitutional diversity. The polycondensation of dihydrazides with dialdehydes generates polyacylhydrazones, to which the acylhydrazone functionality formed confers both hydrogen-bonding and reversibility features through the amide and imine groups, respectively. Polyacylhydrazones are thus dynamic polyamides. They are able to reversibly exchange either one or both of their repeating monomer units in the presence of different monomers, thus presenting constitutional dynamic diversity. The polymers subjected to monomer exchange/interchange may be brought to exhibit physical properties vastly different from those of the original polymer. The principle may be extended to other important classes of polymers, giving access, for instance, to dynamic polyureas or polycarbamates. These reversible polymers are therefore able to incorporate, decorporate, or reshuffle their constituting monomers, namely in response to environmental physical or chemical factors, an adaptability feature central to constitutional dynamic chemistry.

Journal Article↗
PubMed2005

Gelation-driven component selection in the generation of constitutional dynamic hydrogels based on guanine-quartet formation.

The guanosine hydrazide 1 yields a stable supramolecular hydrogel based on the formation of a guanine quartet (G-quartet) in presence of metal cations. The effect of various parameters (concentration, nature of metal ion, and temperature) on the properties of this gel has been studied. Proton NMR spectroscopy is shown to allow a molecular characterization of the gelation process. Hydrazide 1 and its assemblies can be reversibly decorated by acylhydrazone formation with various aldehydes, resulting in formation of highly viscous dynamic hydrogels. When a mixture of aldehydes is used, the dynamic system selects the aldehyde that leads to the most stable gel. Mixing hydrazides 1, 9 and aldehydes 6, 8 in 1:1:1:1 ratio generated a constitutional dynamic library containing the four acylhydrazone derivatives A, B, C, and D. The library constitution displayed preferential formation of the acylhydrazone B that yields the strongest gel. Thus, gelation redirects the acylhydrazone distribution in the dynamic library as guanosine hydrazide 1 scavenges preferentially aldehyde 8, under the pressure of gelation because of the collective interactions in the assemblies of G-quartets B, despite the strong preference of the competing hydrazide 9 for 8. Gel formation and component selection are thermoreversible. The process amounts to gelation-driven self-organization with component selection and amplification in constitutional dynamic hydrogels based on G-quartet formation and reversible covalent connections. The observed self-organization and component selection occur by means of a multilevel self-assembly involving three dynamic processes, two of supramolecular and one of reversible covalent nature. They extend constitutional dynamic chemistry to phase-organization and phase-transition events.

Aldehydes↗
PubMed2001

Distinct constitutive and low-CO2-induced CO2 uptake systems in cyanobacteria: genes involved and their phylogenetic relationship with homologous genes in other organisms.

Cyanobacteria possess a CO(2)-concentrating mechanism that involves active CO(2) uptake and HCO(3)(-) transport. For CO(2) uptake, we have identified two systems in the cyanobacterium Synechocystis sp. strain PCC 6803, one induced at low CO(2) and one constitutive. The low CO(2)-induced system showed higher maximal activity and higher affinity for CO(2) than the constitutive system. On the basis of speculation that separate NAD(P)H dehydrogenase complexes were essential for each of these systems, we reasoned that inactivation of one system would allow selection of mutants defective in the other. Thus, mutants unable to grow at pH 7.0 in air were recovered after transformation of a DeltandhD3 mutant with a transposon-bearing library. Four of them had tags within slr1302 (designated cupB), a homologue of sll1734 (cupA), which is cotranscribed with ndhF3 and ndhD3. The DeltacupB, DeltandhD4, and DeltandhF4 mutants showed CO(2)-uptake characteristics of the low CO(2)induced system observed in wild type. In contrast, mutants DeltacupA, DeltandhD3, and DeltandhF3 showed characteristics of the constitutive CO(2)-uptake system. Double mutants impaired in one component of each of the systems were unable to take up CO(2) and required high CO(2) for growth. Phylogenetic analysis indicated that the ndhD3/ndhD4-, ndhF3/ndhF4-, and cupA/cupB-type genes are present only in cyanobacteria. Most of the cyanobacterial strains studied possess the ndhD3/ndhD4-, ndhF3/ndhF4-, and cupA/cupB-type genes in pairs. Thus, the two types of NAD(P)H dehydrogenase complexes essential for low CO(2)-induced and constitutive CO(2)-uptake systems associated with the NdhD3/NdhF3/CupA-homologues and NdhD4/NdhF4/CupB-homologues, respectively, appear to be present in these cyanobacterial strains but not in other organisms.

Biological Transport↗
PubMed1987

Two promoters, one inducible and one constitutive, control transcription of the Streptomyces lividans galactose operon.

Galactose utilization in Streptomyces lividans was shown to be controlled by an operon that is induced in the presence of galactose and repressed by glucose. Two promoters, galP1 and galP2, which direct transcription of two distinct polycistronic transcripts, have been identified. galP1 is located immediately upstream of the operon and is induced in the presence of galactose. This promoter directs transcription of the galT, galE, and galK genes. The second promoter, galP2, is located within the operon just upstream of the galE gene. This promoter is responsible for constitutive transcription of the galE and galK genes. Comparison of the S. lividans gal operon to the Escherichia coli gal operon indicates the presence of a constitutive promoter positioned upstream of galE in both operons. We suggest that coupling the operon's constitutive promoter to the galE gene fulfills a physiological requirement for constitutive UDPgalactose 4-epimerase expression in Streptomyces.

Base Sequence↗
PubMed1991

Comparison of constitutive and inducible transcriptional enhancement mediated by kappa B-related sequences: modulation of activity in B cells by human T-cell leukemia virus type I tax gene.

The kappa B sequence (GGGACTTTCC) binds a factor, NF-kappa B, that is constitutively found in its functional, DNA binding form only in B lymphocytes. A factor with apparently indistinguishable sequence specificity can be induced in many other cell types, where it is used to regulate inducible gene expression. For example, kappa B-related sequences have been shown to be important for the transcription of a few inducible genes, such as the interleukin 2 receptor alpha-chain gene and the beta-interferon gene. However, these genes are not constitutively active in B lymphocytes, suggesting that other regulatory mechanisms must play a role in determining the patterns of expression. We have investigated the constitutive and inducible transcriptional activity mediated by five kappa B-related sequence elements in two different cell types. We show that in S194 plasma cells the activity of each element correlates well with the relative affinity of B-cell-derived NF-kappa B for that element. This leads to significantly lower transcription enhancement by sites derived from the interleukin 2 receptor or T-cell receptor genes in S194 cells. However, in either EL-4 (T) cells or S194 cells, both lower-affinity sites can be significantly induced by the tax gene product of human T-cell leukemia virus type I, showing that NF-kappa B activity can be modulated even in a B-cell line that constitutively expresses this factor.

Animals↗
PubMed1994

Constitutive and inducible nitric oxide synthase gene expression, regulation, and activity in human lung epithelial cells.

Histochemical activity and immunoreactivity of nitric oxide synthase (NOS, EC 1.14.13.39) have been recently demonstrated in human lung epithelium. However, the molecular nature of NOS and the regulation and function of the enzyme(s) in the airway is not known. A549 cells (human alveolar type II epithelium-like), BEAS 2B cells (transformed human bronchial epithelial cells), and primary cultures of human bronchial epithelial cells all exhibited constitutive NOS activity that was calcium dependent and inhibitable by the NOS inhibitor NG-monomethyl-L-arginine. Nitric oxide production by epithelial cells was enhanced by culture in the presence of interferon gamma, interleukin 1 beta, tumor necrosis factor alpha, and lipopolysaccharide; the NOS activity expressed under these conditions showed less dependence on calcium, reminiscent of other inducible forms of NOS. Two distinct NOS mRNA species, homologous to previously identified constitutive brain (type I) and inducible hepatic (type II) NOS, were demonstrated by reverse transcription-polymerase chain reaction in all cell lines. Northern analysis confirmed the expression of inducible NOS mRNA. Cell culture with epidermal growth factor, a principal regulator of epithelial cell function, decreased inducible NOS activity by posttranscriptional action but did not affect constitutive NOS activity. The coexistence of constitutive and inducible NOS in human alveolar and bronchial epithelial cells is consistent with a complex mechanism evolved by epithelial cells to protect the host from microbial assault at the air/surface interface while shielding the host from the induction of airway hyperreactivity.

Adenocarcinoma↗
PubMed1996

Nonuniformity of the constitutive law parameters for shear rupture and quasistatic nucleation to dynamic rupture: a physical model of earthquake generation processes.

Based on the recent high-resolution laboratory experiments on propagating shear rupture, the constitutive law that governs shear rupture processes is discussed in view of the physical principles and constraints, and a specific constitutive law is proposed for shear rupture. It is demonstrated that nonuniform distributions of the constitutive law parameters on the fault are necessary for creating the nucleation process, which consists of two phases: (i) a stable, quasistatic phase, and (ii) the subsequent accelerating phase. Physical models of the breakdown zone and the nucleation zone are presented for shear rupture in the brittle regime. The constitutive law for shear rupture explicitly includes a scaling parameter Dc that enables one to give a common interpretation to both small scale rupture in the laboratory and large scale rupture as earthquake source in the Earth. Both the breakdown zone size Xc and the nucleation zone size L are prescribed and scaled by Dc, which in turn is prescribed by a characteristic length lambda c representing geometrical irregularities of the fault. The models presented here make it possible to understand the earthquake generation process from nucleation to unstable, dynamic rupture propagation in terms of physics. Since the nucleation process itself is an immediate earthquake precursor, deep understanding of the nucleation process in terms of physics is crucial for the short-term (or immediate) earthquake prediction.

Journal Article↗
PubMed1997

Constitutive and impaired signaling of leptin receptors containing the Gln --> Pro extracellular domain fatty mutation.

Leptin (OB), an adipocyte-secreted circulating hormone, and its receptor (OB-R) are key components of an endocrine loop that regulates mammalian body weight. In this report we have analyzed signal transduction activities of OB-R containing the fatty mutation [OB-R(fa)], a single amino acid substitution at position 269 (Gln --> Pro) in the OB-R extracellular domain that results in the obese phenotype of the fatty rat. We find that this mutant receptor exhibits both ligand-independent transcriptional activation via interleukin 6 and hematopoietin receptor response elements and ligand-independent activation of signal transducer and activator of transcription (STAT) proteins 1 and 3. However, OB-R(fa) is unable to constitutively activate STAT5B and is highly impaired for ligand induced activation of STAT5B compared with OB-R(wt). Introduction of the fatty mutation into a OB-R/G-CSF-R chimera generates a receptor with constitutive character that is similar but distinct from that of OB-R(fa). Constitutive mutant OB-R(fa) receptor signaling is repressed by coexpression of OB-R(wt). The implications of an extracellular domain amino acid substitution generating a cytokine receptor with a partially constitutive phenotype are discussed both in terms of the mechanism of OB-R triggering and the biology of the fatty rat.

Animals↗
PubMed1997

Synthesis and characterization of a novel retinylamine analog inhibitor of constitutively active rhodopsin mutants found in patients with autosomal dominant retinitis pigmentosa.

Two different mutations of the active-site Lys-296 in rhodopsin, K296E and K296M, have been found to cause autosomal dominant retinitis pigmentosa (ADRP). In vitro studies have shown that both mutations result in constitutive activation of the protein, suggesting that the activated state of the receptor may be responsible for retinal degeneration in patients with these mutations. Previous work has highlighted the potential of retinylamine analogs as active-site directed inactivators of constitutively active mutants of rhodopsin with the idea that these or related compounds might be used therapeutically for cases of ADRP involving mutations of the active-site Lys. Unfortunately, however, amine derivatives of 11-cis-retinal, although highly effective against a K296G mutant of rhodopsin, were without affect on the two naturally occurring ADRP mutants, presumably because of the greater steric bulk of Glu and Met side chains in comparison to Gly. For this reason we synthesized a retinylamine analog one carbon shorter than the parent 11-cis-retinal and show that this compound is indeed an effective inhibitor of both the K296E and K296M mutants. The 11-cis C19 retinylamine analog 1 inhibits constitutive activation of transducin by these mutants and their constitutive phosphorylation by rhodopsin kinase, and it does so in the presence of continuous illumination from room lights.

Animals↗
PubMed1997

Targeted expression of constitutively active receptors for parathyroid hormone and parathyroid hormone-related peptide delays endochondral bone formation and rescues mice that lack parathyroid hormone-related peptide.

Mice in which the genes encoding the parathyroid hormone (PTH)-related peptide (PTHrP) or the PTH/PTHrP receptor have been ablated by homologous recombination show skeletal dysplasia due to accelerated endochondral bone formation, and die at birth or in utero, respectively. Skeletal abnormalities due to decelerated chondrocyte maturation are observed in transgenic mice where PTHrP expression is targeted to the growth plate, and in patients with Jansen metaphyseal chondrodysplasia, a rare genetic disorder caused by constitutively active PTH/PTHrP receptors. These and other findings thus indicate that PTHrP and its receptor are essential for chondrocyte differentiation. To further explore the role of the PTH/PTHrP receptor in this process, we generated transgenic mice in which expression of a constitutively active receptor, HKrk-H223R, was targeted to the growth plate by the rat alpha1 (II) collagen promoter. Two major goals were pursued: (i) to investigate how constitutively active PTH/PTHrP receptors affect the program of chondrocyte maturation; and (ii) to determine whether expression of the mutant receptor would correct the severe growth plate abnormalities of PTHrP-ablated mice (PTHrP-/-). The targeted expression of constitutively active PTH/PTHrP receptors led to delayed mineralization, decelerated conversion of proliferative chondrocytes into hypertrophic cells in skeletal segments that are formed by the endochondral process, and prolonged presence of hypertrophic chondrocytes with delay of vascular invasion. Furthermore, it corrected at birth the growth plate abnormalities of PTHrP-/- mice and allowed their prolonged survival. "Rescued" animals lacked tooth eruption and showed premature epiphyseal closure, indicating that both processes involve PTHrP. These findings suggest that rescued PTHrP-/- mice may gain considerable importance for studying the diverse, possibly tissue-specific role(s) of PTHrP in postnatal development.

Animals↗
PubMed1995

The human leukocyte antigen A2 interferon-stimulated response element consensus sequence binds a nuclear factor required for constitutive expression.

Both constitutive and interferon-inducible enhancer-like elements have been identified previously in the promoter of human leukocyte antigen (HLA) class I genes. One of these sites is termed the interferon-stimulated response element (ISRE). We have tested the function of an ISRE consensus sequence in the human HLA class I gene HLA-A2 and confirmed previous studies that showed that the HLA-A2 ISRE consensus sequence does not mediate a response to interferons. However, deletion of the ISRE consensus sequence caused a several-fold reduction in the constitutive expression of the HLA-A2 gene in K562 and Jurkat cells. Mobility shift assays performed with the HLA-A2 ISRE revealed the presence of a constitutive binding protein (ISRE/CBP). This protein binds specifically to the HLA-A2 ISRE sequence, and binding is not efficiently competed by the ISRE sequences of the HLA-B7 or ISG54 genes. Substitution of the HLA-B7 or ISG54 ISRE sequences for the HLA-A2 ISRE sequence caused a severalfold reduction in the constitutive expression of the HLA-A2 gene. Mass determinations showed the ISRE/CBP to be 105 kDa, different than any previously characterized ISRE binding proteins. We propose that ISRE/CBP is a novel positive transcriptional regulatory factor for the HLA-A2 gene that may contribute to the differential expression of HLA-A versus HLA-B genes.

Base Sequence↗
PubMed1995

Cell type-specific secretion of parathyroid hormone-related protein via the regulated versus the constitutive secretory pathway.

Parathyroid hormone-related protein (PTHrP) is endoproteolytically processed to yield a family of mature secretory forms. These include an amino-terminal, a mid-region, and a carboxyl-terminal form. Prior studies suggested that the mid-region form is secreted via the regulated secretory pathway, whereas the amino- and carboxyl-terminal forms are secreted via the constitutive pathway. Further, PTHrP is unusual in that it is produced under normal circumstances by neuroendocrine cell types as well as by prototypical constitutively secreting cell types. The potential for cell-specific secretory pathway use by PTHrP has not been explored. Using immunohistochemical and perifusion techniques, we demonstrate that all three PTHrP daughter peptides are secreted via the regulated pathway in neuroendocrine cells. In contrast, all three daughter peptides are secreted in a constitutive fashion by non-neuroendocrine cells. Thus, the secretion of PTHrP is unique in that it appears to be cell-specific. When it is expressed in neuroendocrine cells that contain the regulated pathway, it is secreted in a regulated fashion; when it is expressed in non-neuroendocrine cells, it defaults to the constitutive pathway. This phenomenon has not previously been described for a polypeptide hormone in naturally occurring cells.

Animals↗
PubMed1996

Transmembrane regions V and VI of the human luteinizing hormone receptor are required for constitutive activation by a mutation in the third intracellular loop.

Gonadotropin receptors are members of the seven-transmembrane (TM) receptor family. Several point mutations in TM V and VI and the intracellular loop 3 (i3) have been identified in the luteinizing hormone (LH) receptor gene, leading to constitutive activation of the receptor. Because gonadotropin receptors are highly conserved, we mutated the follicle-stimulating hormone (FSH) receptor at the corresponding amino acids. However, the FSH receptor mutants showed minimal increases in basal cAMP production. Taking advantage of this difference between the two receptors, we designed chimeric receptors with or without a point mutation in the i3 to identify the region in the LH receptor important for its constitutive activation. Introduction of the point mutation into chimeric receptors containing only TM V to VI from the LH receptor led to major increases in ligand-independent cAMP production. Furthermore, a chimeric receptor with only TM V and VI derived from the LH receptor can be rendered constitutively active by the mutation in the i3 from the FSH receptor. These results suggest that interactions between TM V and VI of the FSH receptor are essential for maintaining the receptor in the more constrained state, whereas interactions between these domains of the LH receptor are permissive for constitutively activating mutations in the i3.

Amino Acid Sequence↗