How GPCR Phosphorylation Patterns Orchestrate Arrestin-Mediated Signaling.

Binding of arrestin to phosphorylated G-protein-coupled receptors (GPCRs) controls many aspects of cell signaling. The number and arrangement of phosphates may vary substantially for a given GPCR, and different phosphorylation patterns trigger different arrestin-mediated effects. Here, we determine how GPCR phosphorylation influences arrestin behavior by using atomic-level simulations and site-directed spectroscopy to reveal the effects of phosphorylation patterns on arrestin binding and conformation. We find that patterns favoring binding differ from those favoring activation-associated conformational change. Both binding and conformation depend more on arrangement of phosphates than on their total number, with phosphorylation at different positions sometimes exerting opposite effects. Phosphorylation patterns selectively favor a wide variety of arrestin conformations, differently affecting arrestin sites implicated in scaffolding distinct signaling proteins. We also reveal molecular mechanisms of these phenomena. Our work reveals the structural basis for the long-standing "barcode" hypothesis and has important implications for design of functionally selective GPCR-targeted drugs.

The myelination-associated G protein-coupled receptor 37 is regulated by Zfp488, Nkx2.2, and Sox10 during oligodendrocyte differentiation.

Oligodendrocyte differentiation and myelination in the central nervous system are controlled and coordinated by a complex gene regulatory network that contains several transcription factors, including Zfp488 and Nkx2.2. Despite the proven role in oligodendrocyte differentiation little is known about the exact mode of Zfp488 and Nkx2.2 action, including their target genes. Here, we used overexpression of Zfp488 and Nkx2.2 in differentiating CG4 cells to identify aspects of the oligodendroglial expression profile that depend on these transcription factors. Although both transcription factors are primarily described as repressors, the detected changes argue for an additional function as activators. Among the genes activated by both Zfp488 and Nkx2.2 was the G protein-coupled receptor Gpr37 that is important during myelination. In agreement with a positive effect on Gpr37 expression, downregulation of the G protein-coupled receptor was observed in Zfp488- and in Nkx2.2-deficient oligodendrocytes in the mouse. We also identified several potential regulatory regions of the Gpr37 gene. Although Zfp488 and Nkx2.2 both bind to one of the regulatory regions downstream of the Gpr37 gene in vivo, none of the regulatory regions was activated by either transcription factor alone. Increased activation by Zfp488 or Nkx2.2 was only observed in the presence of Sox10, a transcription factor continuously present in oligodendroglial cells. Our results argue that both Zfp488 and Nkx2.2 also act as transcriptional activators during oligodendrocyte differentiation and cooperate with Sox10 to allow the expression of Gpr37 as a modulator of the myelination process.

Genome-wide identification of G-protein coupled receptors (GPCRs) and their expression profile in response to ß-cypermethrin stress in Zeugodacus cucurbitae.

G-protein coupled receptors (GPCRs) are the largest and most diverse transmembrane receptor family in the cell. They are involved in regulating a wide range of biological processes, including behavior, reproduction, and development. However, GPCRs have not yet been identified in Zeugodacus cucurbitae. The current study focuses on the GPCRs identification, classification, distribution, and their expression analysis under ß-cypermethrin stress to uncover novel targets for pest management and assist in the development of effective strategies for controlling the melon fly population. We identified 80 GPCRs genes including 50 GPCRs identified in family A, 17 GPCRs identified in family B, 8 identified in family C, and 5 identified in family F. Z. cucurbitae GPCRs showed significant differences in both the number of genes in families or subfamilies, as well as the sequencing of the genes. Interestingly, newly identified GPCRs genes are expressed differently at various developmental stages of Z. cucurbitae. Further, we evaluated these 80 GPCRs using Realtime quantitative PCR to confirm their expression between ß-cypermethrin-resistant (RS) strain and susceptible strain (SS) of Z. cucurbitae. We identified 50 GPCR genes were highly overexpressed in a RS. Among these genes, eight genes were strongly induced by the 30% lethal concentration (LC) while two genes were significantly increased by the 50% LC of ß-cypermethrin. This first genome-wide profiling and characterization of GPCRs could lay foundation for unraveling detoxification mechanism and target site modifications which may improve the insect resistance and could be effective insecticide targets for Z. cucurbitae management.

Redox properties and PAS domain structure of the Escherichia coli energy sensor Aer indicate a multistate sensing mechanism.

The Per-Arnt-Sim (PAS; named for the representative proteins: Period, Aryl hydrocarbon receptor nuclear translocator protein and Single-minded) domain of the dimeric Escherichia coli aerotaxis receptor Aer monitors cellular respiration through a redox-sensitive flavin adenine dinucleotide (FAD) cofactor. Conformational shifts in the PAS domain instigated by the oxidized FAD (FADOX)/FAD anionic semiquinone (FADASQ) redox couple traverse the HAMP (histidine kinases, adenylate cyclases, methyl-accepting chemotaxis proteins, and phosphatases) and kinase control domains of the Aer dimer to regulate CheA kinase activity. The PAS domain of Aer is unstable and has not been previously purified. Here, residue substitutions that rescue FAD binding in an FAD binding-deficient full-length Aer variant were used in combination to stabilize the Aer PAS domain. We solved the 2.4 Å resolution crystal structure of this variant, Aer-PAS-GVV, and revealed a PAS fold that contains distinct features associated with FAD-based redox sensing, such as a close contact between the Arg115 side chain and N5 of the isoalloxazine ring and interactions of the flavin with the side chains of His53 and Asn85 that are poised to convey conformational signals from the cofactor to the protein surface. In addition, we determined the FADox/FADASQ formal potentials of Aer-PAS-GVV and full-length Aer reconstituted into nanodiscs. The Aer redox couple is remarkably low at -289.6 ± 0.4 mV. In conclusion, we propose a model for Aer energy sensing based on the low potential of Aer-PAS-FADox/FADASQ couple and the inability of Aer-PAS to bind to the fully reduced FAD hydroquinone.

A rare non-canonical splice site in Trema orientalis SYMRK does not affect its dual symbiotic functioning in endomycorrhiza and rhizobium nodulation.

BACKGROUND: Nitrogen-fixing nodules occur in ten related taxonomic lineages interspersed with lineages of non-nodulating plant species. Nodules result from an endosymbiosis between plants and diazotrophic bacteria; rhizobia in the case of legumes and Parasponia and Frankia in the case of actinorhizal species. Nodulating plants share a conserved set of symbiosis genes, whereas related non-nodulating sister species show pseudogenization of several key nodulation-specific genes. Signalling and cellular mechanisms critical for nodulation have been co-opted from the more ancient plant-fungal arbuscular endomycorrhizal symbiosis. Studies in legumes and actinorhizal plants uncovered a key component in symbiotic signalling, the LRR-type SYMBIOSIS RECEPTOR KINASE (SYMRK). SYMRK is essential for nodulation and arbuscular endomycorrhizal symbiosis. To our surprise, however, despite its arbuscular endomycorrhizal symbiosis capacities, we observed a seemingly critical mutation in a donor splice site in the SYMRK gene of Trema orientalis, the non-nodulating sister species of Parasponia. This led us to investigate the symbiotic functioning of SYMRK in the Trema-Parasponia lineage and to address the question of to what extent a single nucleotide polymorphism in a donor splice site affects the symbiotic functioning of SYMRK. RESULTS: We show that SYMRK is essential for nodulation and endomycorrhization in Parasponia andersonii. Subsequently, it is revealed that the 5'-intron donor splice site of SYMRK intron 12 is variable and, in most dicotyledon species, doesn't contain the canonical dinucleotide 'GT' signature but the much less common motif 'GC'. Strikingly, in T. orientalis, this motif is converted into a rare non-canonical 5'-intron donor splice site 'GA'. This SYMRK allele, however, is fully functional and spreads in the T. orientalis population of Malaysian Borneo. A further investigation into the occurrence of the non-canonical GA-AG splice sites confirmed that these are extremely rare. CONCLUSION: SYMRK functioning is highly conserved in legumes, actinorhizal plants, and Parasponia. The gene possesses a non-common 5'-intron GC donor splice site in intron 12, which is converted into a GA in T. orientalis accessions of Malaysian Borneo. The discovery of this functional GA-AG splice site in SYMRK highlights a gap in our understanding of splice donor sites.

Membrane receptor activation mechanisms and transmembrane peptide tools to elucidate them.

Single-pass membrane receptors contain extracellular domains that respond to external stimuli and transmit information to intracellular domains through a single transmembrane (TM) α-helix. Because membrane receptors have various roles in homeostasis, signaling malfunctions of these receptors can cause disease. Despite their importance, there is still much to be understood mechanistically about how single-pass receptors are activated. In general, single-pass receptors respond to extracellular stimuli via alterations in their oligomeric state. The details of this process are still the focus of intense study, and several lines of evidence indicate that the TM domain (TMD) of the receptor plays a central role. We discuss three major mechanistic hypotheses for receptor activation: ligand-induced dimerization, ligand-induced rotation, and receptor clustering. Recent observations suggest that receptors can use a combination of these activation mechanisms and that technical limitations can bias interpretation. Short peptides derived from receptor TMDs, which can be identified by screening or rationally developed on the basis of the structure or sequence of their targets, have provided critical insights into receptor function. Here, we explore recent evidence that, depending on the target receptor, TMD peptides cannot only inhibit but also activate target receptors and can accommodate novel, bifunctional designs. Furthermore, we call for more sharing of negative results to inform the TMD peptide field, which is rapidly transforming into a suite of unique tools with the potential for future therapeutics.

The conserved ancestral signaling pathway from cilium to nucleus.

Many signaling molecules are localized to both the primary cilium and nucleus. Localization of specific transmembrane receptors and their signaling scaffold molecules in the cilium is necessary for correct physiological function. After a specific signaling event, signaling molecules leave the cilium, usually in the form of an endocytic vesicle scaffold, and move to the nucleus, where they dissociate from the scaffold and enter the nucleus to affect gene expression. This ancient pathway probably arose very early in eukaryotic evolution as the nucleus and cilium co-evolved. Because there are similarities in molecular composition of the nuclear and ciliary pores the entry and exit of proteins in both organelles rely on similar mechanisms. In this Hypothesis, we propose that the pathway is a dynamic universal cilia-based signaling pathway with some variations from protists to man. Everywhere the cilium functions as an important organelle for molecular storage of certain key receptors and selection and concentration of their associated signaling molecules that move from cilium to nucleus. This could also have important implications for human diseases such as Huntington disease.

Buried ionizable networks are an ancient hallmark of G protein-coupled receptor activation.

Seven-transmembrane receptors (7TMRs) have evolved in prokaryotes and eukaryotes over hundreds of millions of years. Comparative structural analysis suggests that these receptors may share a remote evolutionary origin, despite their lack of sequence similarity. Here we used structure-based computations to compare 221 7TMRs from all domains of life. Unexpectedly, we discovered that these receptors contain spatially conserved networks of buried ionizable groups. In microbial 7TMRs these networks are used to pump ions across the cell membrane in response to light. In animal 7TMRs, which include light- and ligand-activated G protein-coupled receptors (GPCRs), homologous networks were found to be characteristic of activated receptor conformations. These networks are likely relevant to receptor function because they connect the ligand-binding pocket of the receptor to the nucleotide-binding pocket of the G protein. We propose that agonist and G protein binding facilitate the formation of these electrostatic networks and promote important structural rearrangements such as the displacement of transmembrane helix-6. We anticipate that robust classification of activated GPCR structures will aid the identification of ligands that target activated GPCR structural states.

Juxtamembrane contribution to transmembrane signaling.

Signaling across the cell membrane mediated by transmembrane receptors plays an important role in diverse biological processes. Recent studies have indicated that, in a number of single-span transmembrane receptors, the intracellular juxtamembrane (JM) sequence linking the transmembrane helix with the rest of the cytoplasmic domain participates directly in the signaling process via several novel mechanisms. This review briefly highlights several modes of JM dynamics in the context of signal transduction that are shared by different types of transmembrane receptors.

Development of allosteric modulators of GPCRs for treatment of CNS disorders.

The discovery of allosteric modulators of G protein-coupled receptors (GPCRs) provides a promising new strategy with potential for developing novel treatments for a variety of central nervous system (CNS) disorders. Traditional drug discovery efforts targeting GPCRs have focused on developing ligands for orthosteric sites which bind endogenous ligands. Allosteric modulators target a site separate from the orthosteric site to modulate receptor function. These allosteric agents can either potentiate (positive allosteric modulator, PAM) or inhibit (negative allosteric modulator, NAM) the receptor response and often provide much greater subtype selectivity than orthosteric ligands for the same receptors. Experimental evidence has revealed more nuanced pharmacological modes of action of allosteric modulators, with some PAMs showing allosteric agonism in combination with positive allosteric modulation in response to endogenous ligand (ago-potentiators) as well as "bitopic" ligands that interact with both the allosteric and orthosteric sites. Drugs targeting the allosteric site allow for increased drug selectivity and potentially decreased adverse side effects. Promising evidence has demonstrated potential utility of a number of allosteric modulators of GPCRs in multiple CNS disorders, including neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and Huntington's disease, as well as psychiatric or neurobehavioral diseases such as anxiety, schizophrenia, and addiction.

Engineering transcriptional regulation for cell-based therapies.

A major aim in the field of synthetic biology is developing tools capable of responding to user-defined inputs by activating therapeutically relevant cellular functions. Gene transcription and regulation in response to external stimuli are some of the most powerful and versatile of these cellular functions being explored. Motivated by the success of chimeric antigen receptor (CAR) T-cell therapies, transmembrane receptor-based platforms have been embraced for their ability to sense extracellular ligands and to subsequently activate intracellular signal transduction. The integration of transmembrane receptors with transcriptional activation platforms has not yet achieved its full potential. Transient expression of plasmid DNA is often used to explore gene regulation platforms in vitro. However, applications capable of targeting therapeutically relevant endogenous or stably integrated genes are more clinically relevant. Gene regulation may allow for engineered cells to traffic into tissues of interest and secrete functional proteins into the extracellular space or to differentiate into functional cells. Transmembrane receptors that regulate transcription have the potential to revolutionize cell therapies in a myriad of applications, including cancer treatment and regenerative medicine. In this review, we will examine current engineering approaches to control transcription in mammalian cells with an emphasis on systems that can be selectively activated in response to extracellular signals. We will also speculate on the potential therapeutic applications of these technologies and examine promising approaches to expand their capabilities and tighten the control of gene regulation in cellular therapies.

Improvements in Glucose Regulation in Children and Young People with Cystic Fibrosis-Related Diabetes following Initiation of Elexacaftor/Tezacaftor/Ivacaftor.

INTRODUCTION: Cystic fibrosis transmembrane conductance regulator (CFTR) modulators are increasingly used in children and young people with cystic fibrosis (CF). Data in adults show there may be an impact on glycaemic control in those with CF-related diabetes (CFRD). Paediatric data are rare. Case Series/Presentation: Children aged >12 years with CFRD, who were eligible for elexacaftor/tezacaftor/ivacaftor (ELX/TEZ/IVA) were commenced on treatment. Glucose monitoring via the FreeStyle Libre system was commenced prior to, immediately after, and several months after commencing ELX/TEZ/IVA. Glycaemic control, shown by time in range (3-10 mmol/L), percentage of time spent hypoglycaemic (<3 mmol/L), and percentage of time spent hyperglycaemic (>10 mmol/L) on Insulin doses were recorded. Following ELX/TEZ/IVA, four of seven children stopped insulin, two required substantially reduced doses of insulin, one showed no response. Glycaemic control remained similar on lower doses or no insulin. Hypoglycaemia was detected in those not requiring insulin. CONCLUSION: ELX/TEZ/IVA has a positive impact on glycaemic control and insulin requirements in children with CFRD. Close monitoring is required when commencing treatment. Children with CFRD need counselling regarding possible reductions in insulin requirement and re-education regarding symptoms, signs, and management of hypoglycaemia.

Complex formation of immunoglobulin superfamily molecules Side-IV and Beat-IIb regulates synaptic specificity.

Neurons establish specific synapses based on the adhesive properties of cell-surface proteins while also retaining the ability to form synapses in a relatively non-selective manner. However, comprehensive understanding of the underlying mechanism reconciling these opposing characteristics remains incomplete. Here, we have identified Side-IV/Beat-IIb, members of the Drosophila immunoglobulin superfamily, as a combination of cell-surface recognition molecules inducing synapse formation. The Side-IV/Beat-IIb combination transduces bifurcated signaling with Side-IV's co-receptor, Kirre, and a synaptic scaffold protein, Dsyd-1. Genetic experiments and subcellular protein localization analyses showed the Side-IV/Beat-IIb/Kirre/Dsyd-1 complex to have two essential functions. First, it narrows neuronal binding specificity through Side-IV/Beat-IIb extracellular interactions. Second, it recruits synapse formation factors, Kirre and Dsyd-1, to restrict synaptic loci and inhibit miswiring. This dual function explains how the combinations of cell-surface molecules enable the ranking of preferred interactions among neuronal pairs to achieve synaptic specificity in complex circuits in vivo.

Role of vascular endothelial progenitor cells in construction of new vascular loop.

Since bone marrow-derived endothelial progenitor cells (EPCs) have been detected in adult peripheral blood, the mode of vasculogenesis in the adult tissue has caught attention in field of vascularization research. To confirm the role of EPCs in construction of new vascular loop, we took the biomaterial scaffold implanted into adult rat as an experimental model to observe and examine the actions of the EPCs in neovascularization of the material by immunohistochemistry and transmission electron microscopy. Additionally, by establishing a chemotactic migration model for vascular endothelial cells (ECs) and EPCs, the migrations of ECs and EPCs were explored in simulations. The results of 20,000 simulations showed that the number of the vascular loops assisted by the EPCs was 2-5 times that of the vascular sprouts being naturally joined. Based on the results of experiments and simulations, we conclude that the EPCs are able to assist the angiogenic sprouts in joining under the condition of plenty of the EPCs being mobilized, which aggregate at sites close to sprout tips, forming a cell cord and differentiating to ECs in situ, and become vessel segments between neighboring sprouts. This suggests that there is a difference between the adult and embryo in the manner of vasculogenesis and that a small number of EPCs can play an important role to make the new blood vessels achieve rapid functionalization.

Reprogramming of Activated Pancreatic Stellate Cells via Mechanical Modulation of Transmembrane Force-sensitive N-cadherin Receptor.

Cancer has been the leading cause of death due mainly to tumor metastasis. The tumor microenvironment plays a key role in tumor metastasis. As the main stromal cells in tumor microenvironment originated from activated fibroblast, cancer-associated fibroblasts (CAFs) play a major role in promoting tumor metastasis. A promising therapeutic avenue is reprogramming of CAFs into tumor-restraining quiescence state. In this study, we observed that CAF-like active pancreatic stellate cells (PSCs) interact with each other via N-cadherin, a force-sensitive transmembrane receptor. Since N-cadherin ligation mediated mechanotransduction has been reported to restrict integrin mediated signalling, we thus hypothesized that the reprogramming of activated PSCs by mechanical modulation of N-cadherin ligation might be possible. To test this hypothesis, we grafted N-cadherin ligand (HAVDI peptide) onto soft polyethylene glycol hydrogel substrate prior to cell adhesion to mimic cell-cell interaction via N-cadherin ligation. We found that the activated PSCs could be reprogrammed to their original quiescent state when transferred onto the substrate with immobilized HAVDI peptide. These results reveal a key role of mechanosensing by intercellular transmembrane receptor in reprogramming of activated PSCs, and provide a potential way for designing novel therapeutic strategies for cancer treatment.

Prognostic Value of the Expression of Receptor Tyrosine Kinase-Like Orphan Receptor 1 (ROR-1) in Chronic Lymphocytic Leukemia.

Background: The transmembrane receptor tyrosine kinase-like orphan receptor 1 (ROR1) has acted on the causation and sustentation of mature B-cell lymphomagenesis for chronic lymphocytic leukemia (CLL) cells. The study attempted to show whether there is a relationship between the level of ROR1 surface expression in CLL cells and disease findings. Materials and Methods: The level of ROR1 cell surface expression was determined in accordance with the flow cytometric analysis of CLL patients at the first diagnosis time.  Two groups were formed according to the high and low ROR1 levels. The cut-off point for the ROR1 level was calculated for advanced-stage disease using receiver operating characteristic (ROC) curves. A two-sided p-value <0,05 was considered statistically significant. Results: 108 CLL cases with a median age of 60 were enrolled. The median percentage of ROR1 cell surface marker positivity in the CD5/CD19 positive leukemic cell was 62%. The CLL cases with high ROR1 levels have thrombocytopenia (p=0.042), anemia (p=0.028), and high beta-2 microglobulin value ≥3 mg/dL (p=0.002) and the need for first-line treatment (p=0.043). Conclusion: The poor prognostic parameters such as splenomegaly, anemia, higher beta-2 microglobulin levels, intermediate/advanced RAI stage disease, and need for first-line treatment had associated high-level ROR 1 expression of our CLL patients. It needs to be investigated for its effect on predicting disease burden and aggressiveness with more comprehensive studies on ROR1 expression levels in CLL cases.

Model of ligand-triggered information transmission in G-protein coupled receptor complexes.

We present a model for the effects of ligands on information transmission in G-Protein Coupled Receptor (GPCR) complexes. The model is built ab initio entirely on principles of statistical mechanics and tenets of information transmission theory and was validated in part using agonist-induced effector activity and signaling bias for the angiotensin- and adrenergic-mediated signaling pathways, with in vitro observations of phosphorylation sites on the C tail of the GPCR complex, and single-cell information-transmission experiments. The model extends traditional kinetic models that form the basis for many existing models of GPCR signaling. It is based on maximizing the rates of entropy production and information transmission through the GPCR complex. The model predicts that (1) phosphatase-catalyzed reactions, as opposed to kinase-catalyzed reactions, on the C-tail and internal loops of the GPCR are responsible for controlling the signaling activity, (2) signaling favors the statistical balance of the number of switches in the ON state and the number in the OFF state, and (3) biased-signaling response depends discontinuously on ligand concentration.

ROBO1 Promotes Homing, Dissemination, and Survival of Multiple Myeloma within the Bone Marrow Microenvironment.

The bone marrow (BM) microenvironment actively promotes multiple myeloma (MM) pathogenesis and therapies targeting both cancer cells and the niche are highly effective. We were interested in identifying novel signaling pathways supporting MM-BM crosstalk. Mutations in the transmembrane receptor Roundabout 1 (ROBO1) were recently identified in MM patients, however their functional consequences are uncertain. Through protein structure-function studies, we discovered that ROBO1 is necessary for MM adhesion to BM stromal and endothelial cells and ROBO1 knock out (KO) compromises BM homing and engraftment in a disseminated mouse model. ROBO1 KO significantly decreases MM proliferation in vitro and intra- and extramedullary tumor growth, in vivo. Mechanistically, ROBO1 C-terminus is cleaved in a ligand-independent fashion and is sufficient to promote MM proliferation. Viceversa, mutants lacking the cytoplasmic domain, including the human-derived G674* truncation, act dominantly negative. Interactomic and RNA sequencing studies suggest ROBO1 may be involved in RNA processing, supporting further studies.

Melatonin and Its Homologs Induce Immune Responses via Receptors trP47363-trP13076 in Nicotiana benthamiana.

Melatonin (N-acetyl-5-methoxytryptamine), a naturally occurring small molecule, can protect plants against abiotic stress after exogenous treatmenting with it. It is not known if melatonin homologs, such as 5-methoxytryptamine and 5-methoxyindole, that are easy and more cost-effective to synthesize can stimulate the plant immune system in the same manner as melatonin. In the present study, we assessed the biological activity of the melatonin homologs, 5-methoxytryptamin and 5-methoxyindole. The results showed that melatonin and its homologs all induced disease resistance against Phytophthora nicotianae in Nicotiana benthamiana plants. The application of all three compounds also induced stomatal closure and the production of reactive oxygen species. Gene expression analysis indicated that the expression of genes involved in hydrogen peroxide (H2O2), nitric oxide (NO) production, and salicylic acid (SA) biosynthesis was significantly upregulated by all three compounds. Four homologs of the melatonin receptors were identified by blasting search with the phytomelatonin receptor in Arabidopsis. Molecular docking studies were also used to identify four putative melatonin receptors in N. benthamiana. Further experimentation revealed that silencing of the melatonin receptors trP47363 and trP13076 in N. benthamiana compromised the induction of stomatal closure, PR-1a gene expression and SA accumulation by all three compounds. Collectively, our data indicate that the induction of defense responses in N. benthamiana by melatonin, 5-methoxytryptamine, and 5-methoxyindole involves the melatonin receptors trP47363 and trP13076.

Interferon Receptor Trafficking and Signaling: Journey to the Cross Roads.

Like most plasma membrane proteins, type I interferon (IFN) receptor (IFNAR) traffics from the outer surface to the inner compartments of the cell. Long considered as a passive means to simply control subunits availability at the plasma membrane, an array of new evidence establishes IFNAR endocytosis as an active contributor to the regulation of signal transduction triggered by IFN binding to IFNAR. During its complex journey initiated at the plasma membrane, the internalized IFNAR complex, i.e. IFNAR1 and IFNAR2 subunits, will experience post-translational modifications and recruit specific effectors. These finely tuned interactions will determine not only IFNAR subunits destiny (lysosomal degradation vs. plasma membrane recycling) but also the control of IFN-induced signal transduction. Finally, the IFNAR system perfectly illustrates the paradigm of the crosstalk between membrane trafficking and intracellular signaling. Investigating the complexity of IFN receptor intracellular routes is therefore necessary to reveal new insight into the role of IFNAR membrane dynamics in type I IFNs signaling selectivity and biological activity.