GenScript Biotech

The KiSS1-derived peptide receptor (also known as GPR54 or the Kisspeptin receptor) is a G_q-coupled receptor which binds the peptide hormone kisspeptin (metastin). The KiSS1 gene inhibits metastatic activity of melanoma and other tumor cell lines, and clinical evidence supports a role for KiSS1 in inhibition of metastasis in human cancer. Kisspeptins and GPR54 also play a central role in hypothalamic regulation of puberty, by directly governing the release of gonadotropin-releasing hormone from the hypothalamus. In addition, mutations in GPR54 in mice and humans result in hypogonadotropic hypogonadism.

Lymphocyte-activation gene 3, also known as LAG-3, is a protein which in humans is encoded by the LAG3 gene.[1] LAG3, which was discovered in 1990[2] and was designated CD223 (cluster of differentiation 223) after the Seventh Human Leucocyte Differentiation Antigen Workshop in 2000[3]. LAG-3 is a cell surface molecule with diverse biologic effects on T cell function. It is an immune checkpoint receptor and as such is the target of various drug development programs by pharmaceutical companies seeking to develop new treatments for cancer and autoimmune disorders.

The dihydroxy-leukotriene, leukotriene B4 (LTB4) is a member of the G protein-coupled receptor (GPCR) family, in a subfamily of GPCRs that includes receptors for chemokines and other chemotactic factors. Recently, a second, lower affinity receptor for LTB4 (BLT2) has also been cloned, with broader ligand specificity for various eicosanoids. LTB4 stimulates neutrophil chemotaxis and secretion but may also affect immunomodulation, contraction of certain smooth muscles via an indirect mechanism and activation of the nuclear transcription factor PPARα (peroxisome proliferation activated receptor alpha). Chemotaxis, the principal effects of LTB4 and related dihydroxy-acids on leukocytes, occurs via activation of BLT1 receptors.

M1 was expressed in the CNS such as cerebral cortex, basal ganglia, limbic areas, vestibular system and esophageal smooth muscle. Synaptic transmission by muscarinic acetylcholine receptors (mAChRs) is employed throughout the central and peripheral nervous systems to elicit a large and diverse array of neurophysiological actions. An important aspect of mAChR functional diversity is reflected by the multitude of biochemical and electrophysiological actions evoked by acetylcholine binding to mAChRs, which include the regulation of intracellular levels of cAMP, cGMP and inositol phospholipids, and the opening or closing of the potassium, calcium, and chloride ion channels found in certain tissues.

Muscarinic acetylcholine receptors belong to a superfamily of seven-TM-domain receptors that interact with G-proteins to initiate intracellular responses. Five muscarinic receptor subtypes have been identified and named from M1 to M5. The M2 muscarinic receptor couples to G_i/o to inhibit cAMP production. GenScript co-transfected human M2 with Gα15 in the CHO-K1 which supports high levels of recombinant M2 expression on the cell surface and contains high levels of Gα15 to couple the receptor to the calcium signaling pathway.

Muscarinic acetylcholine receptors belong to a superfamily of seven-TM-domain receptors that interact with G-proteins to initiate intracellular responses. Five muscarinic receptor subtypes have been identified and named from M1 to M5. The M3 muscarinic receptors are located at many places in the body, e.g. smooth muscles, endocrine, exocrine glands, as well as lungs. They are also found in the CNS, where it induces emesis. They generally cause smooth muscle contraction and increased glandular secretions.

Muscarinic acetylcholine receptors belong to a superfamily of seven-TM-domain receptors that interact with G-proteins to initiate intracellular responses. Five muscarinic receptor subtypes have been identified and named from M1 to M5. The M4 muscarinic receptor couples to G_i/o to inhibit cAMP production. GenScript co-transfected human M4 with Gα15 in the CHO-K1 which supports high levels of recombinant M4 expression on the cell surface and contains high levels of Gα15 to couple the receptor to the calcium signaling pathway.

Muscarinic acetylcholine receptors belong to a superfamily of seven-TM-domain receptors that interact with G-proteins to initiate intracellular responses. Five muscarinic receptor subtypes have been identified, named M1 through M5. Receptors of the M5 receptor subtype couple through the G_q/11 class of G-proteins and activate the phospholipase C pathway. Activation of this pathway in turn leads to increases in free intracellular calcium levels as inositol triphosphate mediates release of calcium from the endoplasmic reticulum. RT-PCR reveals that M5 mRNA is quite uniformly expressed in brain. However, there is little data regarding the expression and function of the M5 receptor in peripheral tissues. Currently, it is clear that the M5 receptor, due to the high likelihood that its distribution is restricted to the CNS, probably plays a discrete role in dopaminergic transmission. Although the identification of M5 expression in salivary glands and iris-ciliary muscle suggests a broader role, the data on this is sparse and requires extensive confirmation.

The melanocortin receptor 2, MC2 receptor, is G_s-coupled GPCRs expressed in zona fasciculata of the adrenal cortex placental and stimulates production of cortisol. MC2 is a member of the rhodopsin family of 7-transmembrane and it’s also known as the ACTH receptor or corticotropin receptor because it is specific for ACTH alone. Activation of the MC2 receptor initiates a cascade of events affecting multiple steps in corticoid steroidogenesis. Mutations in MC2 may result in familial glucocorticoid deficiency, a group of autosomal recessive disorders characterized by resistance to ACTH.

The melanocortin receptor 3, MC3 receptor, is Gs-coupled GPCRs expressed in brain, placental, and gut tissues but not in melanoma cells or in the adrenal gland. MC3 receptor knockouts exhibit a metabolic syndrome. At 4-6 months old, MC3 receptor knockout mice show increased fat mass, reduced lean mass and a higher feed efficiency, with normal metabolic rates. MC3 receptor knockout mice are hyperleptinaemic and males are often mildly hyperinsulinaemic.

Human Recombinant CHO-K1/MC4/Gα15 (cAMP) Stable Cell Line stably overexpress the melanocortin 4 receptor gene (MC4). This stable cell line product is designed for measuring the In Vitro function of MC4.

The melanocortin receptor 5, MC5 receptor, is Gs-coupled GPCRs expressed in the adrenal gland, fat cells, kidney, leukocytes, lung, lymph node, mammary gland, ovary, pituitary, testis and uterus. Mice with targeted deletion of the MC5 receptor have a defect in water and thermoregulation secondary to their decreased production of sebaceous lipids.

Coupled exclusively Gq, MCH2 is a receptor for melanin-concentrating hormone. Other than regulating skin color, two subtypes (MCHR1 and MCHR2) are involved in energy homeostasis and social behaviors. In contrast to MCHR1 that is conserved among all mammals, functional MCHR2 is only expressed in carnivores and primates, but not in rodents.

Recombinant CHO-K1 cells stably overexpress tumor necrosis factor receptor superfamily member 9 isoform 1 precursor (Mus musculus) on the surface. The surface expression of mouse 4-1BB is validated by FACS analysis. This cell line is designed for cell-based binding for screening antibodies binding with mouse 4-1BB and evaluating target binding affinity.

The adenosine receptors ADORA2A are Gs-coupled GPCRs expressed in the thymus gland, heart, lung, kidney, brain, platelets, spleen and leukocytes. ADORA2A down-regulates chemokine receptor function and inhibits platelet aggregation. ADORA2A antagonists may be useful as therapy for Parkinson’s disease.

CD155, commonly known as PVR (poliovirus receptor) and Necl-5 (nectin-like molecule-5), is a type I transmembrane single-span glycoprotein and belongs to the nectins and nectin-like (Necl) subfamily. CD155 was originally identified based on its ability to mediate the cell attachment and entry of poliovirus (PV), an etiologic agent of the central nervous system disease poliomyelitis. The normal cellular function is in the establishment of intercellular adherens junctions between epithelial cells. CD155 may assist in an efficient humoral immune response generated within the intestinal immune system. It’s been shown that CD155 can be recognized and bind by DNAM-1 and CD96, which promotes the adhension, migration, and NK-cell killing. Thus, inducing cell-mediated tumor-specific immunity.

CD38 (cluster of differentiation 38), also known as cyclic ADP ribose hydrolase is a glycoprotein found on the surface of many immune cells (white blood cells), including CD4⁺, CD8⁺, B lymphocytes and natural killer cells. CD38 also functions in cell adhesion, signal transduction and calcium signaling. In humans, the CD38 protein is encoded by the CD38 gene which is located on chromosome 4. CD38 is a multifunctional ectoenzyme that catalyzes the synthesis and hydrolysis of cyclic ADP-ribose (cADPR) from NAD+ to ADP-ribose. These reaction products are essential for the regulation of intracellular Ca²⁺.

This gene encodes a membrane protein, which is involved in the increase in intracellular calcium concentration that occurs upon cell adhesion to extracellular matrix. The encoded protein is also a receptor for the C-terminal cell binding domain of thrombospondin, and it may play a role in membrane transport and signal transduction. This gene has broad tissue, distribution, and is reduced in expression on Rh erythrocytes. Alternatively spliced transcript variants have been found for this gene.

Recombinant CHO-K1 cells stably overexpress Mus musculus cytotoxic T-lymphocyte-associated protein 4 (CTLA4) on the surface. The surface expression of mouse CTLA4 is validated by FACS analysis. This cell line is designed for cell-based binding for screening antibodies binding with mouse CTLA4 and for evaluating target binding affinity.

HVEM is found on the surface of various cell types, including hematopoietic and non-hematopoietic cells, and is expressed mainly in spleen, thymus, bone marrow, lung and intestines. It has been identified as a canonical TNF receptor, signaling through the TNF receptor-associated factors (TRAFs) leading to NFκB activation. Furthermore, in addition to acting as a signaling receptor, in binding to Ig superfamily members it acts as a ligand for these inhibitory receptors. Therefore, bidirectional signaling is possible for the HVEM-mediated signaling network, which can be involved in positive or negative immunological reactions under different contexts.

Programmed death-ligand 1 (PD-L1) also known as cluster of differentiation 274 (CD274) or B7 homolog 1 (B7-H1) is a protein that in humans is encoded by the CD274 gene. The formation of PD-1 receptor / PD-L1 or B7.1 receptor /PD-L1 ligand complex transmits an inhibitory signal which reduces the proliferation of these CD8+ T cells at the lymph nodes. After that PD-1 is also able to control the accumulation of foreign antigen specific T cells in the lymph nodes through apoptosis, which is further mediated by a downregulation of the Bcl-2 gene.

TIGIT (also called T cell immunoreceptor with Ig and ITIM domains) is one newly discovered immune receptor on some percentage of T cells and Natural Killer Cells (NK). It is also identified as WUCAM and Vstm3. TIGIT binds to CD155 (PVR) on dendritic cells (DCs), macrophages, etc. with high affinity, and also to CD112 (PVRL2) with lower affinity. Research has shown that TIGIT-Fc fusion proteins interact with PVR on dendritic cells and increase IL-10 secretion, while decreasing IL-12 secretion under LPS stimulation. TIGIT-Fc fusion proteins also inhibit T cell activation in vivo. TIGIT’s inhibition of NK cytotoxicity can be blocked by antibodies against its interaction with PVR, which is directed through its ITIM domain.

V-domain Ig suppressor of T cell activation (VISTA) is a potent negative regulator of T-cell function that is expressed on hematopoietic cells. VISTA levels are heightened within the tumor microenvironment, in which its blockade can enhance antitumor immune responses in mice. In humans, blockade of the related programmed cell death 1 (PD-1) pathway has shown great potential in clinical immunotherapy trials.

Recombinant CHO-K1 cells stably overexpress mas-related G-protein coupled receptor member X2 (MRGPRX2). MRGPRX2 couples to both Gi and Gq alpha subunits in mast cells naturally, so that the PLC signaling can be triggered within the cells by agonist binding. This cell line is recommended for agonist screening and functional validations with calcium flux assay.

Melatonin binds to two specific G-protein coupled receptors (GPCR), MT1 (MTNR1A/MEL1A) and MT2 (MTNR1B/MEL1B). MT1 receptors signal via inhibitory G proteins (Gα_i and Gα_o) leading to adenylate cyclase inhibition and possibly inositol phosphate stimulation in recombinant systems. In certain native tissues (e.g. sheep pars tuberalis, rat cerebral and caudal arteries) melatonin responses are presumably mediated through activation of MT1 receptors. The hypothalamic suprachiasmatic nucleus appears to be involved in circadian rhythm while the hypophysial pars tuberalis may be responsible for the reproductive effects of melatonin.

Melatonin is a neurohormone that plays a key role in the synchronisation of circadian and seasonal functions with cyclic environmental variations. In mammals, two melatonin receptors, MT1 and MT2, have been cloned. Activation of MT2 melatonin receptors phase shift circadian rhythms of neuronal firing in the suprachiasmatic nucleus, inhibit dopamine release in retina, induce vasodilation and inhibition of leukocyte rolling in arterial beds, and enhance immune responses.

The motilin receptor (MTLR) represents a clinically useful pharmacological target, as agonists binding to the MTLR have gastroprokinetic properties. Motilin is a 22 amino acid peptide that potently stimulates gastrointestinal contractility. The biological effects of motilin are mediated by a G_q-coupled seven transmembrane protein, currently termed motilin receptor (MR) that shares significant sequence similarity with the ghrelin receptor. The motilin receptor is also activated by the antibiotic erythromycin; this interaction appears to mediate some of the gastrointestinal side effects of erythromycin. Although motilides (non-antibiotic derivatives of erythromycin) such as ABT-229 have been investigated for treatment of diabetic gastroporesis, the effectiveness has been limited by tachyphylaxis (decreased response to ligand) resulting from receptor downregulation. Agonists of the motilin receptor with reduced densensitization activity remain a potential treatment for disorders of gastric motility.

GPR109A is a high affinity receptor for nicotinic acid (niacin) and is a member of the nicotinic acid receptor family of G protein-coupled receptors (the other identified member being GPR109B). GPR109A is a G_i/o protein-coupled receptor with high affinity for nicotinic acid. In GPR109A knockout mice, the effects of niacin on both lipids and flushing are eliminated. Furthermore in arrestin beta 1 knockout mice, niacin’s effect on flushing is greatly reduced while the lipid modifying effects are maintained. GPR109A is believed to be an important biomolecular target of niacin which is a widely prescribed drug for the treatment of dyslipidemia and to increase HDL cholesterol but whose therapeutic use is limited by flushing.

Tachykinins are peptides sharing the common C-terminal amino acid sequence Phe-X-Gly-Leu-Met-NH₂. This neuropeptide family is composed of substance P, neurokinin A, and neurokinin B, which are widely distributed in mammalian central and peripheral nervous systems. It plays a role as both a neurotransmitter and a neuromodulator. Their actions are mediated by binding with three distinct receptors, namely, NK1, NK2, and NK3. NK1 has high affinity with substance P. In the CNS, NK1 has been implicated to play a role in regulating neuronal survival and degeneration. In the cardiovascular system, NK1 mediates endothelium-dependent vasodilatation and plasma protein extravasations. In the gastrointestinal system, NK1 receptors mediate intestinal motility, secretion, and vascular functions. SP-NK1 receptor communication is also involved in glioma development and progression. NK1 receptor antagonists may have several therapeutic applications in diseases mediated by tachykinins, such as pulmonary disorders, gut disorders, and the pathophysiology of depression.

Tachykinins are peptides sharing a common C-terminal amino acid sequence: Phe-X-Gly-Leu-Met-NH₂. This neuropeptide family is composed of substance P, neurokinin A, and neurokinin B, which are widely distributed in mammalian central and peripheral nervous systems. These three molecules serve as both neurotransmitters and neuromodulators. Their actions are mediated by binding with three distinct receptors, namely NK1, NK2, and NK3. In particular, NK2 is expressed in gastrointestinal tract. Activation of NK2 is chiefly responsible for the regulation of intestinal motor functions (both excitatory and inhibitory), secretions, inflammation, and visceral sensitivity. Antagonists of NK2 may be useful in the treatment of irritable bowel syndrome.

Tachykinins are peptides sharing a common C-terminal amino acid sequence: Phe-X-Gly-Leu-Met-NH₂. This neuropeptide family is composed of substance P, neurokinin A, and neurokinin B, which are widely distributed in mammalian central and peripheral nervous systems. These three molecules serve as both neurotransmitters and neuromodulators. Their actions are mediated by binding with three distinct receptors, namely NK1, NK2, and NK3. NK3 receptors show affinity for neurokinin B. They are predominantly expressed in both the peripheral and central nervous systems. NK3 receptors appear to modulate monoaminergic and amino acid neurotransmission. Studies show that manipulating modulation of NK3 receptor activity may have therapeutic utility in psychiatric diseases such as schizophrenia and affective disorders.

Neuromedin U receptor 1 (NMUR1) is a member of G-protein coupled receptor family A and is the most abundant in peripheral tissue, particularly in small intestines and stomach. This protein is a receptor for Neuromedin U (NMU-25). The first biological activity ascribed to NmU was smooth muscle contraction. Neuromedin U has also been reported to increase arterial blood pressure and modify ion transport in the intestinal tract. Finally, NmU injected subcutaneously into rats has been reported to result in a short term increase in circulating ACTH levels and a long term increase in serum corticosterone levels, suggesting a role in regulation of the hypothalamo-pituitary-adrenal axis.

The neuropeptide S receptor isoform a (NPS1a) is G_q/11 and Gs-coupled GPCRs expressed in the bronchial smooth muscle cells, basally in colon epithelium and in occasional basal keratinocytes in skin. An Asn107->Ile mutation is significantly up-regulated in a mouse model of ovalbumen-induced lung inflammation supporting a role in the pathogenesis of asthma.

The NPS receptor is a typical GPCR, also known as GPR154, vasopressin-receptor related receptor 1 (VRR1), or GPRA. NPSR was found mainly expressed in the central nervous system of rats by using in-situ hybridization. NPS receptor mRNA is widely distributed in many brain areas with high expression levels in cortex, hypothalamus, amygdala and multiple midline thalamic nuclei. Many of these areas have been functionally associated with arousal and processing of emotional behavior. In 2004, the NPS receptor was identified as an asthma susceptibility gene in a genome wide screen in Finnish and Canadian patients. The study showed that a number of polymorphic variants of the NPS receptor exist in human and that particular sets of these variants (haplotypes) are associated with an increased risk of asthma and possibly allergic diseases characterized by high IgE serum levels. A carboxy-terminal splice variant of human NPS receptor was found to be over-expressed in asthmatic airway tissue. Expression of NPS receptor mRNA was also found upregulated in a mouse model of airway inflammation.

The NPY family consists of three 36-amino acid peptides, neuropeptide Y (NPY), peptide YY (PYY) and pancreatic polypeptide (PP), which bind to the NPY receptors. At present five distinct NPY receptors, Y1, Y2, Y4, Y5, and y6, have been established by receptor cloning studies and all of them are G_i-coupled GPCRs. Activation of NPY receptors mediate a variety physiological effects including stimulation of food intake, inhibition of anxiety in the CNS, presynaptic inhibition of neurotransmitter release in the CNS and periphery, modulation of circadian rhythm, release of pituitary hormones, modulation of hippocampal activity, pain transmission, vasoconstriction, inhibition of insulin release and modulation of renal function. With regard to endogenous agonists, the receptors Y2 preferentially bind NPY and PYY.

The NPY family consists of three 36-amino acid peptides, neuropeptide Y (NPY), peptide YY (PYY) and pancreatic polypeptide (PP), which bind to the NPY receptors. At present five distinct NPY receptors, Y1, Y2, Y4, Y5, and y6, have been established by receptor cloning studies and all of them are G_i-coupled GPCRs. Activation of NPY receptors mediate a variety physiological effects including stimulation of food intake, inhibition of anxiety in the CNS, presynaptic inhibition of neurotransmitter release in the CNS and periphery, modulation of circadian rhythm, release of pituitary hormones, modulation of hippocampal activity, pain transmission, vasoconstriction, inhibition of insulin release and modulation of renal function. Y4 receptor preferentially binds PP, with significant binding to NPY and PYY.

Neurotensin receptor 1 (NTS1) is a member of G-protein coupled receptor family A and is a receptor for Neurotensin (NT). Neurotensin (NT) exerts its intracellular effect by interacting with 3 different receptors. Two of these receptors (NTR1 and NTR2) belong to the G protein-coupled receptor family, whereas the third one (NTR3) is a type I receptor with a single transmembrane domain. The NTS1 is expressed in CNS such as cerebral cortex, basal ganglia, limbic areas, vestibular system, and esophageal smooth muscle.

Opioid receptor family includes three classic receptors, μ, δ, and κ, also known as OP1, OP2 and OP3, respectively. The receptors are G_i/o-coupled GPCRs which will reduce intracellular cAMP levels after activation. δ-opioid receptor modulates many kinase cascades including ERKs, Akts, JNKs, STAT3, P38 involving Src, Ras, Rac, Raf-1, Cdc42, RTKs. In addition, δ-opioid receptor has also been proposed to interact with μ receptors. The observed pharmacological cross-talk may partially arise from agonist cross-reactivity.

Opioid receptors and their endogenous peptide ligands play important roles in the reward and reinforcement of drugs such as heroin, cocaine, and alcohol. The κ-opioid receptor is a type of opioid receptor which binds the peptide opioid dynorphin as the primary endogenous ligand. κ-opioid receptors are widely distributed in the brain, spinal cord, and in pain neurons. They are associated with the risk for alcohol dependence.

Following the cloning of the classical opioid receptors (mu, delta and kappa), the opiate receptor like-1 (ORL1) was identified as a G-protein coupled receptor (GPCR) with 65% structure homology to the other members of the opioid family. OPRL1 is a receptor for the 17 aa neuropeptide nociceptin/orphanin FQ and may be involved in the regulation of numerous brain activities, particularly instinctive and emotional behaviors. Recently, new study results are consistent with the reported high density of ORL1 receptor mRNA in dorsal raphe nucleus and with inhibitory actions of nociceptin in cells expressing ORL1.

The µ-opioid receptor (oprm1) is the principal site of action in the brain by which morphine, other opiate drugs of abuse, and endogenous opioid peptides effect analgesia and alter mood. Opioid receptors belong to the rhodopsin family of G protein-coupled receptors (GPCRs). The three types of opioid receptors (μ, δ, and κ) have been shown to associate with each other in a homotypic or heterotypic fashion when expressed in heterologous cells. A member of the seven-transmembrane domain (TM) G protein-coupled receptor (GPCR) superfamily, the µ-opioid receptor modulates ion channels and second messenger effectors in an opioid agonist-dependent fashion that is reversible by the classic opiate antagonist naloxone.

OX40L is the ligand for CD134 and is expressed on such cells as DC2s (a subtype of dendritic cells) enabling amplification of Th2 cell differentiation. OX40L has also been designated CD252 (cluster of differentiation 252). Various single nucleotide polymorphisms (SNPs) of the OX40L gene have been identified. For some of them association with systemic lupus erythematosus has been reported.

The orexin/hypocretin peptides orexin A and orexin B (also known as hcrt-1 and hcrt-2) are 33- and 28-amino acid peptides, respectively. They are preferentially expressed in hypothalamus. The orexins have a range of physiological functions including feeding control, energy metabolism, neuroendocrine function modulation, and sleep-wake cycle regulation. The two orexin receptor subtypes OX1 and OX2 both mediate the action of orexin-A and orexin-B and couple efficiently through Gq/11 to activate phospholipase C and lead to elevation of intracellular calcium.

Hypocretin (orexin) receptor 2, also known as OX2, is a human protein encoded by the HCRTR2 gene. Orexin A and orexin B are neuropeptides originally identified as endogenous ligands for OX2. Orexin neuropeptides are produced by a small group of neurons in the lateral hypothalamic and perifornical areas, a region that is classically implicated with the control of mammalian feeding behavior. Orexin neurons that project throughout the central nervous system to nuclei are known to be important in the control of feeding, sleep-wakefulness, neuroendocrine homeostasis, and autonomic regulation.

OX40 (CD134) is a member of the TNFR/TNF superfamily. Costimulatory signals from OX40 to a conventional T cell promotes division and survival, augmenting the clonal expansion of effector and memory populations as they are being generated to antigens. OX40 additionally suppresses the differentiation and activity of Treg, further amplifying this process.

Oxytocin is an abundant neuropeptide that exerts a wide spectrum of central and peripheral effects. In the context of human reproduction, oxytocin promotes uterine contractions and lactation. It is the most commonly used drug for labor induction. Oxytocin receptors are expressed in the uterus and in mammary glands, where they mediate functions related to partutition, such as contraction of the uterine myometrium during labor and milk letdown. In addition, oxytocin receptors are expressed in a variety of other peripheral tissues and in the brain, where oxytocin mediates a variety of functions. OXTR antagonists may be promising candidates to prevent preterm labor and dysmenorrheal and its agonists may also be useful for treatment of psychiatric illnesses such as anxiety, drug abuse, sexual dysfunctions, eating disorders, and autism.

The Vasoactive intestinal Peptide/Pituitary Adenylate Cyclase Activating Polypeptide (VPAC) receptor is G protein-coupled and has been divided into at least three types: PAC1, VPAC1, and VPAC2. High expression of PAC1 is observed in CNS and the adrenal medulla. Several splice variants of PAC1 result in proteins that differ at the N-terminus and third intracellular loop; these variants differ in their affinities for PACAP and abilities to activate Gq and Gs. This manual describes establishment of a cell line and a protocol of pharmacologically validated human PAC1 GPCR receptor (Genebank Accession Number: NM_003382).

Proteinase-activated receptors (PAR) are a subfamily of G-protein coupled, seven-transmembrane domain receptors, which are cleaved within the aminoterminal exodomain by certain serine proteinases at a specific peptide bond. Trypsin and mast cell tryptase, and more recently, the activated coagulation factors VIIa and Xa, have been identified as serine proteinases able to activate mammalian PAR-2. As already indicated, PAR-2 is believed to be involved in inflammation. This role for PAR-2 implies that elastase and cathepsin G would paradoxically display an anti-inflammatory property by disarming PAR-2.

Protease-activated receptor (PAR)-4 is a member of a unique family of GPCRs. The protease-activated receptors (PARs) that are activated by proteolytic cleavage of the N-terminal domain of the receptor reveal a tethered ligand. The PAR family consists of 4 receptors; PAR1 and PAR3 are activated by thrombin, and PAR2 and PAR4 are activated by several serine proteases (Macfarlane et al., 2001). PAR4 is a recently identified low-affinity thrombin receptor that plays a pathophysiological role in many types of tissues including the lung. Mice lacking PAR4 are protected from mesenteric arteriole thrombosis, indicating that PAR4 is a potential target for treatment of thrombosis in humans.

Programmed cell death protein 1, also known as PD-1 and CD279 (cluster of differentiation 279), is a protein that in humans is encoded by the PDCD1 gene. PD-1, functioning as an immune checkpoint, plays an important role in down regulating the immune system by preventing the activation of T-cells, which in turn reduces autoimmunity and promotes self-tolerance. The inhibitory effect of PD-1 is accomplished through a dual mechanism of promoting apoptosis (programmed cell death) in antigen specific T-cells in lymph nodes while simultaneously reducing apoptosis in regulatory T cells (suppressor T cells).