Intracellular Domain Research Articles

A leucine-rich-repeat receptor-like kinase regulates pollen aperture formation in rice.

Apertures in pollen grains exhibit species-specific patterns and provide an ideal model for studying cell-surface patterning. Pollen apertures are critical for cereal crop fertility, and while DEFECTIVE IN APERTURE FORMATION1 (OsDAF1) and INAPERTURATE POLLEN1 (OsINP1) have been documented to participate in pollen aperture formation in rice (Oryza sativa), the molecular transduction pathway regulating aperture formation is largely unknown. Here, we report that a leucine-rich-repeat receptor-like kinase (LRR-RLK), AM1, plays a key role in rice pollen aperture formation. Mutations of OsAM1 lead to complete sterility due to disappearance of the pollen aperture and failure in pollen tube germination. OsAM1 encodes a LRR-RLK that belongs to the STRUBBELIG-receptor family. Similar to other reported aperture regulators, OsAM1 assembles to future aperture sites on tetrads after meiosis to regulate aperture formation. The extracellular and intracellular domain of OsAM1 interacts with OsINP1 and OsDAF1, respectively. However, despite their interaction and the absence of aperture formation in osam1 pollen grains, OsINP1 and OsDAF1 localize to future aperture sites at the tetrad stage. Mutation of OsINP1, however, disrupts normal localization of OsAM1, indicating that OsAM1 acts downstream of OsINP1. Our findings reveal the role of a LRR-RLK protein in pollen aperture formation and shed light on the regulatory network of pollen aperture formation.

Revealing a hidden conducting state by manipulating the intracellular domains in KV10.1 exposes the coupling between two gating mechanisms.

The KCNH family of potassium channels serves relevant physiological functions in both excitable and non-excitable cells, reflected in the massive consequences of mutations or pharmacological manipulation of their function. This group of channels shares structural homology with other voltage-gated K+ channels, but the mechanisms of gating in this family show significant differences with respect to the canonical electromechanical coupling in these molecules. In particular, the large intracellular domains of KCNH channels play a crucial role in gating that is still only partly understood. Using KCNH1(KV10.1) as a model, we have characterized the behavior of a series of modified channels that could not be explained by the current models. With electrophysiological and biochemical methods combined with mathematical modeling, we show that the uncovering of an open state can explain the behavior of the mutants. This open state, which is not detectable in wild-type channels, appears to lack the rapid flicker block of the conventional open state. Because it is accessed from deep closed states, it elucidates intermediate gating events well ahead of channel opening in the wild type. This allowed us to study gating steps prior to opening, which, for example, explain the mechanism of gating inhibition by Ca2+-Calmodulin and generate a model that describes the characteristic features of KCNH channels gating.

Revealing a hidden conducting state by manipulating the intracellular domains in KV10.1 exposes the coupling between two gating mechanisms

Revealing a hidden conducting state by manipulating the intracellular domains in KV10.1 exposes the coupling between two gating mechanisms

Cannabinoid combination targets NOTCH1-mutated T-cell acute lymphoblastic leukemia through the integrated stress response pathway.

In T-cell acute lymphoblastic leukemia (T-ALL), more than 50% of cases display autoactivation of Notch1 signaling, leading to oncogenic transformation. We have previously identified a specific chemovar of Cannabis that induces apoptosis by preventing Notch1 maturation in leukemia cells. Here, we isolated three cannabinoids from this chemovar that synergistically mimic the effects of the whole extract. Two were previously known, cannabidiol (CBD) and cannabidivarin (CBDV), whereas the third cannabinoid, which we termed 331-18A, was identified and fully characterized in this study. We demonstrated that these cannabinoids act through cannabinoid receptor type 2 and TRPV1 to activate the integrated stress response pathway by depleting intracellular Ca2+. This is followed by increased mRNA and protein expression of ATF4, CHOP, and CHAC1, which is hindered by inhibiting the upstream initiation factor eIF2α. The increased abundance of CHAC1 prevents Notch1 maturation, thereby reducing the levels of the active Notch1 intracellular domain, and consequently decreasing cell viability and increasing apoptosis. Treatment with the three isolated molecules resulted in reduced tumor size and weight in vivo and slowed leukemia progression in mice models. Altogether, this study elucidated the mechanism of action of three distinct cannabinoids in modulating the Notch1 pathway, and constitutes an important step in the establishment of a new therapy for treating NOTCH1-mutated diseases and cancers such as T-ALL.

Exploring the constitutive activation mechanism of the class A orphan GPR20.

GPR20, an orphan G protein-coupled receptor (GPCR), shows significant expression in intestinal tissue and represents a potential therapeutic target to treat gastrointestinal stromal tumors. GPR20 performs high constitutive activity when coupling with Gi. Despite the pharmacological importance of GPCR constitutive activation, determining the mechanism has long remained unclear. In this study, we explored the constitutive activation mechanism of GPR20 through large-scale unbiased molecular dynamics simulations. Our results unveil the allosteric nature of constitutively activated GPCR signal transduction involving extracellular and intracellular domains. Moreover, the constitutively active state of the GPR20 requires both the N-terminal cap and Gi protein. The N-terminal cap of GPR20 functions like an agonist and mediates long-range activated conformational shift. Together with the previous study, this study enhances our knowledge of the self-activation mechanism of the orphan receptor, facilitates the drug discovery efforts that target GPR20.

Isolation and analysis of the exosomal membrane proteins in bullous pemphigoid

ABSTRACT Background Bullous pemphigoid (BP) is a severe autoimmune sub-epidermal bullous disease. Exosomes are small extracellular vesicles secreted by most cell types. The exosomal membrane proteins are implicated in various biological and pathological pathways. This study aims to explore the potential roles of exosomes in BP pathomechanism. Research design We collected plasma samples from 30 BP patients and 31 healthy controls. Nanoparticle tracking analysis (NTA) was used to analyze the size and concentration of exosomes. The immunogold labelling experiment and extracellular vesicle (EV) array were performed to detect the content and distribution of exosomes. Results The exosomes from both the BP and control groups’ plasma were successfully extracted. EV Array showed that CD63 and CD9 levels were significantly higher in the BP group than in the control group (p < 0.05). Expression levels of the BP180 NC16A and intracellular domain (ICD) were higher in the anti-BP180 positive group versus the controls (p < 0.05). The active BP group exhibits higher CD63 and BP180 ICD protein concentrations than the control or inactive BP groups (p < 0.05). Conclusion BP180 autoantigen fragments were expressed on the exosomal membrane in BP patients. The BP180 ICD and CD63 on exosomes could potentially be novel biomarkers for monitoring disease activity.

Functional Aptamers In Vitro Evolution for Intranuclear Blockage of RNA-Protein Interaction.

Over the last 30 years, despite considerable research and endeavors aimed at harnessing aptamers as pharmaceutical molecules, the progress in developing aptamer-based drugs has been falling short of expectations. Sequential steps of affinity molecule acquisition and functional screening are typically required for discovering affinity-based macromolecule therapeutics, which can be time-consuming and limiting in candidate selection. Additionally, aptamers often necessitate tedious postselection modifications to overcome pharmacokinetic limitations, which usually impede the binding affinity. Herein, we propose a novel in vitro screening platform termed Functional Aptamers in vitro Evolution (FAIVE), which integrates affinity molecule acquisition with functional screening and introduces chemical diversity during the process. This platform aims to rapidly generate functional aptamers capable of binding to target proteins and regulating their functions. Illustrated by targeting intranuclear RNA-protein interactions involving HIV-1 Tat protein and TAR RNA, FAIVE demonstrates a selection of functional aptamers with significant intracellular blocking effects. The study also explores lipid nanoparticle delivery systems to enhance intracellular delivery efficiency, expanding aptamer targeting potential to broader intracellular and intranuclear domains. This study emphasizes the potential of FAIVE to expedite the development of aptamer-based drugs and facilitate the creation of more versatile and effective therapeutics.

Finely ordered intracellular domain harbors an allosteric site to modulate physiopathological function of P2X3 receptors

P2X receptors, a subfamily of ligand-gated ion channels activated by extracellular ATP, are implicated in various physiopathological processes, including inflammation, pain perception, and immune and respiratory regulations. Structural determinations using crystallography and cryo-EM have revealed that the extracellular three-dimensional architectures of different P2X subtypes across various species are remarkably identical, greatly advancing our understanding of P2X activation mechanisms. However, structural studies yield paradoxical architectures of the intracellular domain (ICD) of different subtypes (e.g., P2X3 and P2X7) at the apo state, and the role of the ICD in P2X functional regulation remains unclear. Here, we propose that the P2X3 receptor’s ICD has an apo state conformation similar to the open state but with a less tense architecture, containing allosteric sites that influence P2X3’s physiological and pathological roles. Using covalent occupancy, engineered disulfide bonds and voltage-clamp fluorometry, we suggested that the ICD can undergo coordinated motions with the transmembrane domain of P2X3, thereby facilitating channel activation. Additionally, we identified a novel P2X3 enhancer, PSFL77, and uncovered its potential allosteric site located in the 1α3β domain of the ICD. PSFL77 modulated pain perception in P2rx3+/+, but not in P2rx3−/−, mice, indicating that the 1α3β, a “tunable” region implicated in the regulation of P2X3 functions. Thus, when P2X3 is in its apo state, its ICD architecture is fairly ordered rather than an unstructured outward folding, enabling allosteric modulation of the signaling of P2X3 receptors.

CD28 hinge used in chimeric antigen receptor (CAR) T-cells exhibits local structure and conformational exchange amidst global disorder

T-cell therapies based on chimeric antigen receptor (CAR) targeting of a tumor-specific antigen offer hope for patients with relapsed or refractory cancers. CAR hinge and transmembrane regions link antigen recognition domains to intracellular signal transduction domains. Here, we apply biophysical methods to characterize the structure and dynamic properties of the CD28 CAR hinge (CD28H) used in an FDA-approved CD19 CAR for the treatment of B-lineage leukemia/lymphoma. By using nuclear Overhauser effect spectroscopy (NOESY), which detects even transiently occupied structural motifs, we observed otherwise elusive local structural elements amidst overall disorder in CD28H, including a conformational switch from a native β-strand to a 310-helix and polyproline II helix-like structure. These local structural motifs contribute to an overall loosely formed extended geometry that could be captured by NOESY data. All FDA-approved CARs use prolines in the hinge region, which we find in CD28, and previously in CD8α, isomerize to promote structural plasticity and dynamics. These local structural elements may function in recognition and signaling events and constrain the spacing between the transmembrane and antigen recognition domains. Our study thus demonstrates a method for detecting local and transient structure within intrinsically disordered systems and moreover, our CD28H findings may inform future CAR design.

Neuregulin1 Nuclear Signaling Influences Adult Neurogenesis and Regulates a Schizophrenia Susceptibility Gene Network within the Mouse Dentate Gyrus.

Neuregulin1 (Nrg1) signaling is critical for neuronal development and function from fate specification to synaptic plasticity. Type III Nrg1 is a synaptic protein which engages in bidirectional signaling with its receptor ErbB4. Forward signaling engages ErbB4 phosphorylation, whereas back signaling engages two known mechanisms: (1) local axonal PI3K-AKT signaling and (2) cleavage by γ-secretase resulting in cytosolic release of the intracellular domain (ICD), which can traffic to the nucleus (Bao et al., 2003; Hancock et al., 2008). To dissect the contribution of these alternate signaling strategies to neuronal development, we generated a transgenic mouse with a missense mutation (V321L) in the Nrg1 transmembrane domain that disrupts nuclear back signaling with minimal effects on forward signaling or local back signaling and was previously found to be associated with psychosis (Walss-Bass et al., 2006). We combined RNA sequencing, retroviral fate mapping of neural stem cells, behavioral analyses, and various network analyses of transcriptomic data to investigate the effect of disrupting Nrg1 nuclear back signaling in the dentate gyrus (DG) of male and female mice. The V321L mutation impairs nuclear translocation of the Nrg1 ICD and alters gene expression in the DG. V321L mice show reduced stem cell proliferation, altered cell cycle dynamics, fate specification defects, and dendritic dysmorphogenesis. Orthologs of known schizophrenia (SCZ)-susceptibility genes were dysregulated in the V321L DG. These genes coordinated a larger network with other dysregulated genes. Weighted gene correlation network analysis and protein interaction network analyses revealed striking similarity between DG transcriptomes of V321L mouse and humans with SCZ.

Children with hemoglobin C or S trait have low serologic responses to a subset of malaria variant surface antigens

Children with hemoglobin AC or AS have decreased susceptibility to clinical malaria. Parasite variant surface antigen (VSA) presentation on the surface of infected erythrocytes is altered in erythrocytes with hemoglobin C (Hb AC) or sickle trait (Hb AS) mutations in vitro. The protective role of incomplete or altered VSA presentation against clinical malaria in individuals with Hb AC or AS is unclear. Using a high-throughput protein microarray, we sought to use serological responses to VSAs as a measure of host exposure to VSAs among Malian children with Hb AC, Hb AS, or wildtype hemoglobin (Hb AA). In uncomplicated malaria, when compared to Hb AA children, Hb AC children had significantly lower serological responses to extracellular Plasmodium falciparum erythrocyte membrane protein-1 (PfEMP1) domains but did not differ in responses to intracellular PfEMP1 domains and other VSAs, including members of the repetitive interspersed family (RIFIN) and subtelomeric variable open reading frame (STEVOR) family. Healthy children with Hb AC and Hb AS genotypes recognized fewer extracellular PfEMP1s compared to children with Hb AA, especially CD36-binding PfEMP1s. These reduced serologic responses may reflect reduced VSA presentation or lower parasite exposure in children with Hb AC or AS and provide insights into mechanisms of protection.

A Non-Canonical p75HER2 Signaling Pathway Underlying Trastuzumab Action and Resistance in Breast Cancer.

Overexpression of HER2 occurs in 25% of breast cancer. Targeting HER2 has proven to be an effective therapeutic strategy for HER2-positive breast cancer. While trastuzumab is the most commonly used HER2 targeting agent, which has significantly improved outcomes, the overall response rate is low. To develop novel therapies to boost trastuzumab efficacy, it is critical to identify the mechanisms underlying trastuzumab action and resistance. We recently showed that the inhibition of breast cancer cell growth by trastuzumab is not through the inhibition of HER2 canonical signaling. Here we report the identification of a novel non-canonical HER2 signaling pathway and its interference by trastuzumab. We showed that HER2 signaled through a non-canonical pathway, regulated intramembrane proteolysis (RIP). In this pathway, HER2 is first cleaved by metalloprotease ADAM10 to produce an extracellular domain (ECD) that is released and the p95HER2 that contains the transmembrane domain (TM) and intracellular domain (ICD). p95HER2, if further cleaved by an intramembrane protease, γ-secretase, produced a soluble ICD p75HER2 with nuclear localization signal (NLS). p75HER2 is phosphorylated and translocated to the nucleus. Nuclear p75HER2 promotes cell proliferation. Trastuzumab targets this non-canonical HER2 pathway via inhibition of the proteolytic cleavage of HER2 by both ADAM10 and γ-secretase. However, p75HER2 pathway also confers resistance to trastuzumab once aberrantly activated. Combination of trastuzumab with ADAM10 and γ-secretase inhibitors completely blocks p75HER2 production in both BT474 and SKBR3 cells. We concluded that HER2 signals through the RIP signaling pathway that promotes cell proliferation and is targeted by trastuzumab. The aberrant HER2 RIP signaling confers resistance to trastuzumab that could be overcome by the application of inhibitors to ADAM10 and γ-secretase.

Dachsous and Fat coordinately repress the Dachs-Dlish-Approximated complex to control growth.

Two protocadherins, Dachsous (Ds) and Fat (Ft), regulate organ growth in Drosophila via the Hippo pathway. Ds and Ft bind heterotypically to regulate the abundance and subcellular localization of a 'core complex' consisting of Dachs, Dlish and Approximated. This complex localizes to the junctional cortex where it promotes growth by repressing the pathway kinase Warts. Ds is believed to promote growth by recruiting and stabilizing the core complex at the junctional cortex, while Ft represses growth by promoting degradation of core complex components. Here, we examine the functions of intracellular domains of Ds and Ft and their relationship to the core complex. While Ds promotes accumulation of the core complex proteins in cortical puncta, it is not required for core complex assembly. Indeed, the core complex assembles maximally in the absence of both Ds and Ft. Furthermore, while Ds promotes growth in the presence of Ft, it represses growth in the absence of Ft by removing the core complex from the junctional cortex. Ft similarly recruits core complex components, however it normally promotes their degradation. Our findings reveal that Ds and Ft constrain tissue growth by repressing the default 'on' state of the core complex.

Development of bioassay platforms for biopharmaceuticals using Jurkat-CAR cells by AICD

The assessment of bioactivity for therapeutic antibody release assay poses challenges, particularly when targeting immune checkpoints. An in vitro bioassay platform was developed using the chimeric antigen receptor on Jurkat cells (Jurkat-CAR) to analyze antibodies targeting immune checkpoints, such as CD47/SIRPα, VEGF/VEGFR1, PD-1/PD-L1, and CD70/CD27. For CD47/SIRPα, the platform involved a Jurkat-CAR cell line expressing the chimeric SIRPα receptor (CarSIRPα). CarSIRPα was created by sequentially fusing the SIRPα extracellular region with the CD8α hinge region, the transmembrane (TM) and intracellular (IC) domains of CD28, and the intracellular signaling domain of CD3ζ. The resulting Jurkat-CarSIRPα cells can undergo “activation-induced cell death (AICD)” upon incubation with purified or cellular CD47, as evidenced by the upregulation of CD69, IL-2, and IFN-γ. Similar results also appeared in Jurkat CarVEGFR1, Jurkat CarPD1 and Jurkat CARCD27 cells. These cells are perfectly utilized for the bioactivity analysis of therapeutic antibody. Our study indicates that the established in vitro assay platform based on Jurkat-CAR has been confirmed repeatedly and has shown robust reproducibility; thus, this platform can be used for screening or for release assays of given antibody drugs targeting immune checkpoints.

Twisted Intramolecular Charge Transfer-Based Viscosity-Responsive Probe Reveals Lysosomal Degradation Process of Endocytosed Foreign Bodies.

The optimization of nanomedicines requires a thorough understanding of nanocarrier attrition during lysosome-mediated biological processes. Real-time monitoring of endocytosis provides valuable insights into the lysosomal effects on nanocarriers and the release of nanodrugs. We report the development of a coresponsive probe that detects changes in the spatial viscosity of the intracellular domain caused by lysosomal degradation of foreign bodies. The probe, based on a benzofuro[2,3-d]pyrimidine structure, exhibits torsional intramolecular charge transfer (TICT) and responds to ambient viscosity changes with a sensitive fluorescence intensity. The antidiffused fluorescence transition of the probe in the spatially restricted domain serves as a key indicator for real-time monitoring. When encapsulated with diverse foreign bodies and emitted into macrophages by endocytosis, the probe forms nanoparticles. Lysosomes uptake these materials for intracellular digestion, causing alterations in the aggregation or depolymerization state of the nanoparticles, leading to viscosity changes manifested by the probe's fluorescence. By studying the spatial viscosity changes caused by lysosomal degradation of foreign bodies, our monitoring strategy contributes to understanding the digestion or escape capabilities of potential pharmaceutical-carrying nanocarriers, providing guidelines to design more effective nanocarriers that navigate lysosomal degradation to achieve precise drug payloads and release.

Apoptotic Receptors and CD107a Expression by NK Cells in an Interaction Model with Trophoblast Cells.

Natural killer cells (NK cells) exert cytotoxicity towards target cells in several ways, including the expression of apoptosis-mediating ligands (TRAIL, FasL). In addition, NK cells themselves may be susceptible to apoptosis due to the expression of TRAIL receptors. These receptors include TRAIL-R1 (DR4), TRAIL-R2 (DR5), capable of inducing apoptosis, and TRAIL-R3 (DcR1), TRAIL-R4 (DcR2), the so-called "decoy receptors", which lack an intracellular domain initiating activation of caspases. Of particular interest is the interaction of uterine NK cells with cells of fetal origin, trophoblasts, which are potential targets for natural killer cells to carry out cytotoxicity. The aim of this work was to evaluate the expression of proapoptotic receptors and their ligands as well as CD107a expression by NK cells in a model of interaction with trophoblast cells. To evaluate NK cells, we used cells of the NK-92 line; cells of the JEG-3 line were used as target cells. The cytokines IL-1β, IL-15, IL-18, TNFα, IL-10, TGFβ and conditioned media (CM) of the first and third trimester chorionic villi explants were used as inducers. We established that cytokines changed the expression of apoptotic receptors by NK cells: in the presence of TNFα, the amount and intensity of Fas expression increased, while in the presence of TGFβ, the amount and intensity of expression of the DR5 receptor decreased. Soluble chorionic villi factors alter the expression of TRAIL and FasL by NK-92 cells, which can reflect the suppression of the TRAIL-dependent mechanism of apoptosis in the first trimester and stimulating the Fas-dependent mechanism in the third trimester. In the presence of trophoblast cells, the expression of TRAIL and DcR1 by NK cells was reduced compared to intact cells, indicating an inhibitory effect of trophoblast cells on NK cell cytotoxicity. In the presence of chorionic villi CM and trophoblast cells, a reduced number of NK-92 cells expressing DR4 and DR5 was found. Therefore, soluble factors secreted by chorionic villi cells regulate the resistance of NK cells to death by binding TRAIL, likely maintaining their activity at a certain level in case of contact with trophoblast cells.

LOXL2-induced PEAR1 Ser891 phosphorylation suppresses CD44 degradation and promotes triple-negative breast cancer metastasis.

CD44 is associated with a high risk of metastasis, recurrence, and drug resistance in various cancers. Here we report that platelet endothelial aggregation receptor 1 (PEAR1) is a CD44 chaperone protein that protected CD44 from endocytosis-mediated degradation and enhances cleavage of the CD44 intracellular domain (CD44-ICD). Furthermore, we found that lysyl oxidase-like protein 2 (LOXL2), an endogenous ligand of PEAR1, bound to the PEAR1-EMI domain and facilitated the interaction between PEAR1 and CD44 by inducing PEAR1 Ser891 phosphorylation in a manner that was independent of its enzyme activity. Levels of PEAR1 protein and PEAR1 phosphorylation at Ser891 were increased in patients with triple-negative breast cancer (TNBC), were positively correlated with expression of LOXL2 and CD44, and were negatively correlated with overall survival. The level of PEAR1 Ser891 phosphorylation was identified as the best independent prognostic factor in TNBC patients. The prognostic efficacy of the combination of PEAR1 phosphorylation at Ser891 and CD44 expression was superior to that of PEAR1 phosphorylation at Ser891 alone. Blocking the interaction between LOXL2 and PEAR1 with monoclonal antibodies significantly inhibited TNBC metastasis, representing a promising therapeutic strategy for TNBC.

A Chimeric IL-7Rα/IL-2Rβ Receptor Promotes the Differentiation of T Cell Progenitors into B Cells and Type 2 Innate Lymphoid Cells.

IL-7 and IL-2 are evolutionarily related cytokines that play critical roles in the development and expansion of immune cells. Although both IL-7R and IL-2R activate similar signaling molecules, whether their signals have specific or overlapping functions during lymphocyte differentiation remains unclear. To address this question, we generated IL-7R α-chain (IL-7Rα)/IL-2R β-chain (IL-24β) (72R) knock-in mice expressing a chimeric receptor consisting of the extracellular domain of IL-7Rα and the intracellular domain of IL-2Rβ under the control of the endogenous IL-7Rα promoter. Notably, this 72R receptor induced higher levels of STAT5 and Akt phosphorylation in T cells. In the periphery of 72R mice, the number of T cells, B cells, and type 2 innate lymphoid cells (ILC2s) was increased, whereas early T cell progenitors and double-negative 2 thymocytes were reduced in the thymus. In addition, cell proliferation and Notch signaling were impaired in the early thymocytes of 72R mice, leading to their differentiation into thymic B cells. Interestingly, ILC2s were increased in the thymus of 72R mice. Early T cell progenitors from 72R mice, but not from wild-type mice, differentiated into NK cells and ILC2-like cells when cocultured with a thymic stromal cell line. Thus, this study indicates that the chimeric 72R receptor transduces more robust signals than the authentic IL-7Rα, thereby inducing the alternative differentiation of T cell progenitors into other cell lineages. This suggests that cytokine receptors may provide instructive signals for cell fate decisions.

Development of a highly sensitive platform for protein–protein interaction detection and regulation of T cell function

We developed a highly sensitive assay for detecting protein-protein interaction using chimeric receptors comprising two molecules of interest in the extracellular domain and interferon alpha and beta receptor subunit 1 or 2 (IFNAR1/2) in the intracellular domain. This intracellular IFNAR1/2 reconstitution system (IFNARRS) proved markedly more sensitive than the NanoBiT system, currently considered one of the best detection systems for protein interaction. Employing chimeric receptors with extracellular domains from the IFNγ or IL-2 receptor and the intracellular domains of IFNAR1/2, the IFNARRS system effectively identifies low IFNγ or IL-2 levels. Cells stably expressing these chimeric receptors responded to IFNγ secreted by activated T cells following various stimuli, including a specific peptide-antigen. The activation signals were further enhanced by the expression of relevant genes, such as costimulators, via IFN-stimulated response elements in the promoters. Besides IFNγ or IL-2, the IFNARRS system demonstrated the capability to detect other cytokines by using the corresponding extracellular domains from these target cytokine receptors.

G protein coupled receptor in apoptosis and apoptotic cell clearance.

Apoptosis is a genetically programmed form of cell death that is substantially conserved across the evolutionary tree. Apoptotic cell elimination includes recognition, phagocytosis, and degradation. Failure to clear apoptotic cells can ultimately cause a series of human diseases, such as systemic lupus erythematosus, Alzheimer's disease, atherosclerosis, and cancer. Consequently, the timely and effective removal of apoptotic cells is crucial to maintaining the body's homeostasis. GPCRs belong to the largest membrane receptor family. Its intracellular domain exerts an effect on the trimer G protein. By combining with a variety of ligands, the extracellular domain of G protein initiates the dissociation of G protein trimers and progressively transmits signals downstream. Presently, numerous G protein-coupled receptors (GPCRs) have been identified as participants in the apoptosis signal transduction pathway and the apoptotic cell clearance pathway. Therefore, studies on the mechanism of GPCRs in the clearance of apoptotic cells is important for the development of GPCRs therapeutics.