Insulin Receptor-related Receptor Research Articles

A Comparative Kidney Transcriptome Analysis of Bicarbonate-Loaded insrr-Null Mice.

The maintenance of plasma pH is critical for life in all organisms. The kidney plays a critical role in acid-base regulation in vertebrates by controlling the plasma concentration of bicarbonate. The receptor tyrosine kinase IRR (insulin receptor-related receptor) is expressed in renal β-intercalated cells and is involved in alkali sensing due to its ability to autophosphorylate under alkalization of extracellular medium (pH > 7.9). In mice with a knockout of the insrr gene, which encodes for IRR, urinary bicarbonate secretion in response to alkali loading is impaired. The specific regulatory mechanisms in the kidney that are under the control of IRR remain unknown. To address this issue, we analyzed and compared the kidney transcriptomes of wild-type and insrr knockout mice under basal or bicarbonate-loaded conditions. Transcriptomic analyses revealed a differential regulation of a number of genes in the kidney. Using TaqMan real-time PCR, we confirmed different expressions of the slc26a4, rps7, slc5a2, aqp6, plcd1, gapdh, rny3, kcnk5, slc6a6 and atp6v1g3 genes in IRR knockout mice. Also, we found that the expression of the kcnk5 gene is increased in wild-type mice after bicarbonate loading but not in knockout mice. Gene set enrichment analysis between the IRR knockout and wild-type samples identified that insrr knockout causes alterations in expression of genes related mostly to the ATP metabolic and electron transport chain processes.

Structural basis of the alkaline pH-dependent activation of insulin receptor-related receptor.

The insulin receptor (IR) family is a subfamily of receptor tyrosine kinases that controls metabolic homeostasis and cell growth. Distinct from IR and insulin-like growth factor 1 receptor, whose activation requires ligand binding, insulin receptor-related receptor (IRR)-the third member of the IR family-is activated by alkaline pH. However, the molecular mechanism underlying alkaline pH-induced IRR activation remains unclear. Here, we present cryo-EM structures of human IRR in both neutral pH inactive and alkaline pH active states. Combined with mutagenesis and cellular assays, we show that, upon pH increase, electrostatic repulsion of the pH-sensitive motifs of IRR disrupts its autoinhibited state and promotes a scissor-like rotation between two protomers, leading to a T-shaped active conformation. Together, our study reveals an unprecedented alkaline pH-dependent activation mechanism of IRR, opening up opportunities to understand the structure-function relationship of this important receptor.

Hyperinsulinism Gateway Between Diabetes and Cancer: New Roles for Bifunctional Antidiabetic - Anticancer Active Somatostatins, IGF - 1 Inhibitors and Insulin Sensitizers

Hyperinsulinism refers to a condition of abnormally high concentration of circulating insulin in the body commonly seen in obese and type 2 diabetic patients due to insulin resistance. Elevated circulating insulin is the main factor linking obesity, diabetes and cancer and an important mechanism of cancer therapeutic resistance and failure. The insulin receptor subfamily which is made up of the insulin receptor (IR), the type 1 IGF receptor (IGF-1) which binds insulin-like growth factors I and II, and the orphan insulin receptor-related receptor (IRRR) have been identified and linked to the process of carcinogenesis, as cancer cells require insulin for growth. IR receptors are overexpressed in various cancers and insulin activates growth mostly through these receptors and their interplays through mainly the PI3K/AKT/mTOR, MAPK and RAS signal transduction pathways. Estrogens and androgens synergistic effects worsen hyperinsulinism and accelerates its carcinogenesis properties while progesterone and Sex Hormone Binding Globulins exert modulatory effects. This review examined the various factor interplays in hyperinsulinism that links diabetes with cancer and the biomarkers involved. It equally identified the potentials and therapeutic roles for Somatostatins, IGF – 1 inhibitors and Insulin sensitizing antidiabetics in preventing and reversing the cancer linked trends.

Relationship between Condition Index Values and Expression Levels of Gene and Protein in the Adductor Muscle of Diploid and Triploid Oysters Crassostrea gigas

Three proteins [myosin heavy chain (MHC), filamin-C fragment (FIL-C), and actin 2 (ACT2)] were identified in adductor muscle from diploid and triploid Pacific oysters (Crassostrea gigas) and the relationship between the condition index (CI) and mRNA expression of these genes was investigated, together with the mRNA expression of molluscan insulin-related peptide (MIP), C. gigas insulin receptor-related receptor (CIR), and insulin-like growth factor binding protein complex acid labile subunit (IGFBP-ALS). Monthly changes in the CI were similar to the changes in the tissue weight rate in both groups. ACT2 and MHC mRNA expression was statistically higher in the triploid than the diploid, while FIL-C mRNA expression was significantly higher in the diploid (p<0.05). The MIP, CIR, and IGFBP-ALS mRNA expression of the diploid oysters were all significantly higher in July than in other months (p<0.05). The MIP, CIR, and IGFBP-ALS mRNA expression in the triploid oysters was high in July, but there were no significant differences (p>0.05). Changes in the expression levels of the genes investigated in this study could be used as intrinsic indicators of the annual growth, maturity, and spawning period of cultured diploid and triploid C. gigas in Tongyeong, Korea.

Insulin Receptor-Related Receptor Regulates the Rate of Early Development in Xenopus laevis.

The orphan insulin receptor-related receptor (IRR) encoded by insrr gene is the third member of the insulin receptor family, also including the insulin receptor (IR) and the insulin-like growth factor receptor (IGF-1R). IRR is the extracellular alkaline medium sensor. In mice, insrr is expressed only in small populations of cells in specific tissues, which contain extracorporeal liquids of extreme pH. In particular, IRR regulates the metabolic bicarbonate excess in the kidney. In contrast, the role of IRR during Xenopus laevis embryogenesis is unknown, although insrr is highly expressed in frog embryos. Here, we examined the insrr function during the Xenopus laevis early development by the morpholino-induced knockdown. We demonstrated that insrr downregulation leads to development retardation, which can be restored by the incubation of embryos in an alkaline medium. Using bulk RNA-seq of embryos at the middle neurula stage, we showed that insrr downregulation elicited a general shift of expression towards genes specifically expressed before and at the onset of gastrulation. At the same time, alkali treatment partially restored the expression of the neurula-specific genes. Thus, our results demonstrate the critical role of insrr in the regulation of the early development rate in Xenopus laevis.

The insulin receptor family in the heart: new light on old insights.

Insulin was discovered over 100 years ago. Whilst the first half century defined many of the physiological effects of insulin, the second emphasised the mechanisms by which it elicits these effects, implicating a vast array of G proteins and their regulators, lipid and protein kinases and counteracting phosphatases, and more. Potential growth-promoting and protective effects of insulin on the heart emerged from studies of carbohydrate metabolism in the 1960s, but the insulin receptors (and the related receptor for insulin-like growth factors 1 and 2) were not defined until the 1980s. A related third receptor, the insulin receptor-related receptor remained an orphan receptor for many years until it was identified as an alkali-sensor. The mechanisms by which these receptors and the plethora of downstream signalling molecules confer cardioprotection remain elusive. Here, we review important aspects of the effects of the three insulin receptor family members in the heart. Metabolic studies are set in the context of what is now known of insulin receptor family signalling and the role of protein kinase B (PKB or Akt), and the relationship between this and cardiomyocyte survival versus death is discussed. PKB/Akt phosphorylates numerous substrates with potential for cardioprotection in the contractile cardiomyocytes and cardiac non-myocytes. Our overall conclusion is that the effects of insulin on glucose metabolism that were initially identified remain highly pertinent in managing cardiomyocyte energetics and preservation of function. This alone provides a high level of cardioprotection in the face of pathophysiological stressors such as ischaemia and myocardial infarction.

The Ambiguity Issue in Solving Inverse Problems of Small-Angle Scattering: A Consistent Approach Using an Insulin Receptor-Related Receptor as an Example. Methods for Interpreting SAXS Data

The construction of three-dimensional models of protein macromolecules is a serious challenge due to the possible ambiguity of solving the inverse problem of reconstructing a three-dimensional structure from a one-dimensional small-angle scattering profile. The target function of this task can have several local minima, which leads to the dependence of the solution on the initial values of the model parameters and on the method of finding the global minimum. The problem of creating structural models is also complicated by averaging the scattering pattern over all orientations of particles in space and by the size and shape distribution of scattering objects in the case of polydispersity and/or polymorphism. In this study, the issue of ambiguity in solving inverse problems and restoring the three-dimensional structure of a protein is considered using the structure of the ectodomain of an insulin receptor-related receptor (ectoIRR) in solution as an example. The paper presents a consistent approach to solving this problem, starting from the determination of general structural parameters and ab initio reconstruction of shape to modeling by rigid bodies (using molecular tectonics), hybrid methods, and analysis of scattering profiles by singular vector decomposition.

Abstract P-13: Structural Studies of Insulin Receptor-Related Receptor Ectodomain

Background: The insulin receptor-related receptor (IRR) was originally discovered due to its high homology to the other family members (insulin receptor and insulin-like growth factor 1 receptor). We determined that IRR can be activated by mildly alkaline extracellular media and has typical features of the ligand-receptor interaction, including its specificity and dose-dependence. Since pH-sensitive properties of IRR are determined by its ectodomain; therefore, we chose as an option to study the soluble extracellular domain IRR. Methods: The investigation carried out in Titan Krios 60-300 TEM/STEM (FEI, USA) CryoEM, equipped with direct electron detector Falcon II (FEI, USA) and Cs image corrector (CEOS, Germany), at an accelerating voltage of 300 kV. Data processing and 3D reconstruction were carried out using computing resources of the Federal Collective Usage Center Complex for Simulation and Data Processing for Mega-Science Facilities at NRC “Kurchatov Institute.” Results: The obtained 2D classifications of particles of the ectodomain IRR at a neutral pH form several 3D models. This indicates that the ectodomain has several possible conformations, which is consistent with our previously obtained data using SEC-SAXS and AFM. In the future, additional careful data processing is required, as well as studies of the IRR ectodomain in mildly alkaline pH. Conclusion: In this study, we presented the structural characteristics of the IRR ectodomain obtained by CryoEM. These results are an important step towards understanding the mechanism of functioning of the IRR.

Activity-dependent conformational transitions of the insulin receptor–related receptor

The insulin receptor (IR), insulin-like growth factor 1 receptor (IGF-1R), and insulin receptor-related receptor (IRR) form a mini family of predimerized receptor-like tyrosine kinases. IR and IGF-1R bind to their peptide agonists triggering metabolic and cell growth responses. In contrast, IRR, despite sharing with them a strong sequence homology, has no peptide-like agonist but can be activated by mildly alkaline media. The spatial structure and activation mechanisms of IRR have not been established yet. The present work represents the first account of a structural analysis of a predimerized receptor-like tyrosine kinase by high-resolution atomic force microscopy in their basal and activated forms. Our data suggest that in neutral media, inactive IRR has two conformations, where one is symmetrical and highly similar to the inactive Λ/U-shape of IR and IGF-1R ectodomains, whereas the second is drop-like and asymmetrical resembling the IRR ectodomain in solution. We did not observe complexes of IRR intracellular catalytic domains of the inactive receptor forms. At pH 9.0, we detected two presumably active IRR conformations, Γ-shaped and T-shaped. Both of conformations demonstrated formation of the complex of their intracellular catalytic domains responsible for autophosphorylation. The existence of two active IRR forms correlates well with the previously described positive cooperativity of the IRR activation. In conclusion, our data provide structural insights into the molecular mechanisms of alkali-induced IRR activation under mild native conditions that could be valuable for interpretation of results of IR and IGF-IR structural studies.

Probing Structure and Function of Alkali Sensor IRR with Monoclonal Antibodies.

To study the structure and function of the pH-regulated receptor tyrosine kinase insulin receptor-related receptor (IRR), а member of the insulin receptor family, we obtained six mouse monoclonal antibodies against the recombinant IRR ectodomain. These antibodies were characterized in experiments with exogenously expressed full-length IRR by Western blotting, immunoprecipitation, and immunocytochemistry analyses. Utilizing a previously obtained set of IRR/IR chimeras with swapped small structural domains and point amino acid substitutions, we mapped the binding sites of the obtained antibodies in IRR. Five of them showed specific binding to different IRR domains in the extracellular region, while one failed to react with the full-length receptor. Unexpectedly, we found that 4D5 antibody can activate IRR at neutral pH, and 4C2 antibody can inhibit activation of IRR by alkali. Our study is the first description of the instruments of protein nature that can regulate activity of the orphan receptor IRR and confirms that alkali-induced activation is an intrinsic property of this receptor tyrosine kinase.

Role of the insulin receptor‐related receptor in Xenopus laevis development

The orphan insulin receptor‐related receptor (IRR), in contrast to its close homologs, the insulin receptor (IR) and insulin‐like growth factor receptor (IGF‐IR) can be activated by mildly alkaline extracellular medium. Unlike ubiquitously expressed IR and IGF‐IR, IRR is found in specific sets of cells in only some tissues, most of them being exposed to extracorporal liquids of extreme pH. In the evolution, IRR is highly conserved since its divergence from the insulin and insulin‐like growth factor receptors in amphibia. IR and IGF‐IR signaling are important in growth and development in zebrafish or frog embryogenesis. In contrast, the role of IRR activation during embryogenesis is unknown although Xenopus embryo have strong IRR expression. To address this, we examined the function of the IRR during Xenopus laevis development by morpholino‐mediated selective knockdown of IRR. We demonstrated that inhibition of IRR expression in Xenopus laevis leads to delayed development, but this phenotype can be restored by incubation of embryos in alkaline medium. Also, using RNA‐seq of total RNA we showed that IRR inhibition dramatically changed the embryo transcriptome, and partially restored after alkali treatment. We have identified several hundred genes (eomes, frzb, pax6 etc.) whose expression changes after IRR knockdown and restores by alkali exposure. Our results clearly demonstrate that IRR plays previously unidentified important role in frog embryogenesis and growth.Support or Funding InformationThis work was financially supported by the Russian Foundation for Basic Research (grants No.19‐04‐00815, 17‐00‐00486)To understand which role IRR could play in the embryonic development, we downregulated it by injecting anti‐sense morpholino oligonucleotide to IRR mRNA (MO1 xIRR) into the two‐cells embryos (Fig. 1A). As a result, we observed significant retardation of development of these embryos comparing to their siblings injected by the control MO (Fig. 1B,C). As IRR in mouse is known as a receptor which activity depends on pH, we decided to verify if pH different from the neutral one could interfere with the effects observed in embryos with downregulated pH. To this end, we incubated embryos injected with IRR MO and by the control one in solutions with pH 5.5, 7.2 and 8.4.Whereas clear retardation of development was seen in the MO1 xIRR injected embryos incubated in pH 5.2 and 7.2 (Fig, 1B,C,D,E). Surprisingly, the embryos incubated at pH 8.4 developed approximately synchronously with the control ones (Fig. 1F,G). In other words, unexpectedly alkaline pH “rescues” embryos with downregulated IRR from retardation of development.We represented in table 1 top of 15 up‐regulated and 15 down‐regulated genes after MO injection in embryos at pH 7.2, which included several key transcription factors such as eomes, pou5f3.2 (or oct25), hhex and pax6. Also, we see that genes with the largest modified expression are actin or myosin coding genes.Figure 1 gene Fold change MO xIRR 7.2/control 7.2 Fold change MO xIRR 8.4/MO xIRR 7.2 mylpf.L 0,15 2,95 myosin light chain, phosphorylatable, fast skeletal muscle S homeolog(mylpf.S) act3.L 0,15 1,89 actin, alpha skeletal muscle MGC64484 0,15 1,89 Actin, alpha cardiac muscle 1 act2 0,20 2,32 actin, alpha sarcomeric/cardiac myl1.S 0,25 1,93 myosin light chain 1 S homeolog MGC53823 0,25 1,99 Actin, alpha cardiac muscle 2‐like col2a1.L 0,26 2,07 collagen, type II, alpha 1 L homeolog des.1.L 0,26 2,11 desmin, gene 1 L homeolog fbxl22.S 0,30 1,94 F‐box and leucine‐rich repeat protein 22 S homeolog tnnc2.L 0,30 1,68 troponin C type 2 (fast) L homeolog pax6.S 0,30 2,43 paired box 6 S homeolog des.1.S 0,30 1,95 desmin, gene 1 S homeolog smyd1.L 0,31 2,21 SET and MYND domain containing 1 L homeolog tnnt3.L 0,32 2,26 troponin T3, fast skeletal type L homeolog nr2f5.S 0,32 1,87 nuclear receptor subfamily 2, group F, member 5 S homeolog dctn2.L 1,92 0,58 dynactin subunit 2 L homeolog pou5f3.2.L 1,93 0,51 POU class 5 homeobox 3, gene 2 L homeolog ywhag.L 1,93 0,50 tyrosine 3‐monooxygenase/tryptophan 5‐monooxygenase activation protein, gamma L homeolog ppp2r2b.L 1,93 0,52 protein phosphatase 2 regulatory subunit B, beta L homeolog chm.L 1,97 0,56 choroideremia (Rab escort protein 1) L homeolog hhex.L 1,97 0,62 hematopoietically expressed homeobox L homeolog slc22a15.2.S 1,97 0,63 solute carrier family 22, member 15, gene 2 S homeolog gramd1c.L 1,97 0,70 GRAM domain containing 1C L homeolog ccna1.L 1,99 0,48 cyclin A1 L homeolog neu1.L 2,00 0,56 neuraminidase 1 (lysosomal sialidase) L homeolog cdk5r2.S 2,03 0,57 cyclin‐dependent kinase 5, regulatory subunit 2 (p39) S homeolog cer1.S 2,11 0,54 cerberus 1, DAN family BMP antagonist S homeolog xpo6.S 2,19 0,67 exportin 6 S homeolog eomes.S 2,42 0,36 eomesodermin S homeolog frzb.S 2,71 0,39 frizzled‐related protein S homeolog

Role of the insulin-like growth factor system in gonad sexual maturation in Pacific oyster Crassostrea gigas

BackgroundThe IGF system plays important roles in controlling growth, development, reproduction, and aging of organisms.MethodsTo estimate maturation of the Pacific oyster Crassostrea gigas, we investigated the expression of insulin-like growth factor (IGF) system components and sex-specific genes. To determine the role of the IGF system in the growth and spawning period of female and male oysters, we examined mRNA expression levels of the C. gigas insulin receptor-related receptor (CIR), IGF binding protein complex acid labile subunit (IGFBP_ALS), and molluscan insulin-related peptide (MIP), as well as those of vitellogenin (Vg) and receptor-type guanylate cyclase (Gyc76C) in gonads of C. gigas collected between April and October, when sex can be determined visually in this species.ResultsWe found that MIP, IGFBP_ALS, and CIR mRNA expression levels were dependent on sex and month and were greater in males than in females. CIR and Vg mRNA expression levels were very similar among females, whereas IGF system components and Gyc76C were very similarly expressed among males. The highest expression values were observed in May, when oysters are mature; CIR and Vg mRNA expression levels were highest in females, and those of MIP, IGFBP_ALS, CIR, and Gyc76C were highest in males. Interestingly, we observed a 1:1 proportion of females to males during this period.ConclusionOur results suggest that IGF system components, as well as Vg and Gyc76C, are associated with sexual maturation in C. gigas.

The dimeric ectodomain of the alkali-sensing insulin receptor–related receptor (ectoIRR) has a droplike shape

Insulin receptor-related receptor (IRR) is a receptor tyrosine kinase of the insulin receptor family and functions as an extracellular alkali sensor that controls metabolic alkalosis in the regulation of the acid-base balance. In the present work, we sought to analyze structural features of IRR by comparing them with those of the insulin receptor, which is its closest homolog but does not respond to pH changes. Using small-angle X-ray scattering (SAXS) and atomic force microscopy (AFM), we investigated the overall conformation of the recombinant soluble IRR ectodomain (ectoIRR) at neutral and alkaline pH. In contrast to the well-known inverted U-shaped (or λ-shaped) conformation of the insulin receptor, the structural models reconstructed at different pH values revealed that the ectoIRR organization has a "droplike" shape with a shorter distance between the fibronectin domains of the disulfide-linked dimer subunits within ectoIRR. We detected no large-scale pH-dependent conformational changes of ectoIRR in both SAXS and AFM experiments, an observation that agreed well with previous biochemical and functional analyses of IRR. Our findings indicate that ectoIRR's sensing of alkaline conditions involves additional molecular mechanisms, for example engagement of receptor juxtamembrane regions or the surrounding lipid environment.

IRR is involved in glucose-induced endocytosis after insulin secretion

Endocytosis after insulin secretion plays a pivotal role in the regulation of insulin secretion in pancreatic β-cells. Our recent study suggested that EPI64, a GTPase activating protein for Rab27a, contributes to the regulation of glucose-induced endocytosis, which is mediated by the GDP-bound form of Rab27a. Here, we identified insulin receptor-related receptor (IRR) as an EPI64-interacting protein. Knockdown of IRR inhibited glucose-induced uptake of transferrin, a marker of endocytosis, translocation of the guanine-nucleotide-exchange factor ARNO to the plasma membrane, and generation of phosphatidylinositol 3,4,5-trisphosphate (PIP3). These results suggest that IRR functions upstream of PIP3 generation and controls endocytosis after insulin secretion.

Optimization of heterological expression of insulin receptor-related receptor ectodomain

In this paper, we present an approach to optimize the heterologous expression of the receptor tyrosine kinase IRR, which further simplifies the purification of the IRR from the medium and increases the final yield. The approach proposed by us can find application in the biotechnological production of other large-scale recombinant proteins produced for medical purposes.

The Hybrid Protein of the Alkaline Sensor IRR and the Fluorescent Protein GFP Retains the Functional Activity of the Receptor

The insulin receptor-related receptor (IRR) is a cellular sensor of a weakly alkaline medium. Its spatial structure and the mechanism of activation have not been yet established. In the present work, a system of heterologous expression of full-length IRR tyrosine kinase has been created. A plasmid construct encoding the cDNA of the full-length human IRR fused at the C-terminus with the green fluorescent protein (GFP) has been generated, and the hybrid protein has been expressed in HEK293 line cells. It has been shown that the receptor incorporated in the hybrid protein is expressed on the cytoplasmic membrane and retains its functional activity. The resulting protein can be used in further studies of the structure and function of IRR.

Abstract 443: α 1 -adrenergic Receptor Agonists Activate Akt Signalling via the Insulin Receptor Family of Receptors

Introduction: Akt signalling was delineated in the context of insulin stimulation and is cardioprotective. Akt is activated by insulin or insulin-like growth factor 1 (IGF1), acting through their receptors (IRs, IGF1Rs). Other receptors including α 1 -adrenergic receptors (α 1 ARs) activate Akt, but the mechanism is unclear. Hypothesis: We hypothesise that α 1 -ARs activate Akt via insulin receptor family receptors (InsRFs). Our aims were to establish if this is so, and if linsitinib [a highly selective inhibitor of InsRFs developed for cancer] affects cardiac responses to α 1 -AR agonists. Methods: Rat neonatal cardiomyocytes were exposed to or adult rat hearts perfused with α 1 AR agonists [50 nM A61603; 100 μM phenylephrine (PE)] with/without linsitinib. Effects on signalling were assessed by immunoblotting. Protein synthesis was assessed using 3 H-Phe. Effects of linsitinib (2 mg/kg/d) on cardiac function/hypertrophy induced by PE (40 mg/kg/d, 4 d) in mice (male C57Bl6; 10 wks) in vivo were determined. Echocardiography was used to assess cardiac function and pulmonary/aortic flow. Results: We identified a third InsRF in heart, the insulin receptor-related receptor (IRR). IRRs are alkali-sensitive and required for pH regulation in kidney. Alkaline pH (pH 9.0) activated IRRs and Akt, and increased protein synthesis in perfused hearts, being activated in a complex with IR/IGF1Rs. α 1 ARs activated Akt in cardiomyocytes and perfused hearts and increased protein synthesis. Linsitinib inhibited activation of Akt and the increase in protein synthesis induced by alkaline pH or α 1 AR agonists. Thus, α 1 ARs signal via InsRFs to Akt and protein synthesis. In perfused hearts, linsitinib enhanced the increase in developed pressure induced by A61603 and, in vivo, linsitinib enhanced PE-induced left ventricular (LV) hypertrophy with increased LV wall thickness and decreased internal diameter. Aortic flow was compromised in the presence of PE/linsitinib. Conclusions: Our data demonstrate a novel signalling paradigm in which α 1 -ARs signal through one or more InsRFs in cardiomyocytes to activate Akt and increase protein synthesis. However, functionally, the InsRF signal appears to mitigate some of the hypertrophic effects of α 1 -AR signalling in the whole heart.

Involvement of insulin-like growth factor in intraspecific variation in growth of Pacific oyster Crassostrea gigas during winter

Insulin-like growth factor I (IGF-I) plays a critical role in regulating proliferation, differentiation, growth, and survival in fish as well as in humans. Few studies have investigated the involvement of IGFs in bivalves. In Korea, the Pacific oyster Crassostrea gigas is an important aquaculture species, and its growth is dependent on environmental conditions. Intrinsic factors that can be used as indicators to assess oyster growth based on individual size are limited. This study analyzed IGF-I, IGF-I receptor (IGF-IR), and IGF binding protein (IGFBP) in adductor muscle of C. gigas in winter using immunoblotting and reverse transcriptase-polymerase chain reaction. Levels of IGF-I and the IGF-IR β subunit, and expression of C. gigas insulin receptor-related receptor and IGFBP messenger RNA were significantly higher in large individuals (LI) than small individuals (SI). The IGF-IR β subunit was detected in the adductor muscle of both groups; however, phosphorylation of the IGF-IR β subunit and extracellular signal-regulated kinase was greater in LI than SI. The condition index of C. gigas showed a similar pattern. The results of this study suggest that the IGF system is associated with growth rate differences in C. gigas during winter in Korea.

Behavioral Characteristics of Mice with Knockout of the IRR Alkali Sensor Gene

The tyrosine kinase receptor IRR (insulin receptor-related receptor) can be activated on alkalinization of the cell medium. IRR is detected in organs in contact with fluids with extreme pH values, as well as in certain central nervous system cells, where its role is unclear. We report here our analysis of the behavioral characteristics of mice with knockout of the IRR gene in a series of ethological tests. The control group included animals with the IRR gene from the same litters as knockout mice. In the social interest test, knockout animals showed a reduction in the number of social contacts. The forced swimming test revealed no statistically significant differences in the duration of immobility, though there were more animals showing longer periods of immobility among knockout mice. In the resident-intruder test, wild-type mice showed typical aggression levels, while seven of 16 knockout animals remained inert and, conversely, were attacked by the “stranger.” These data provide evidence that deletion of the IRR gene leads to impairment of the aggressive-defensive behavior typical of the parental strain.

Insulin receptor‐related receptor as an extracellular pH sensor regulating bicarbonate excretion

The acid‐base balance in the organism at the system level is controlled primarily by the kidneys and lungs. In the kidneys, this function is performed by the intercalated cells of two types; alpha cells excrete excess acids while beta cells excrete excess bases as bicarbonate. In a result of the biochemical analysis of the insulin receptor‐related receptor (IRR), a receptor‐like tyrosine kinase enriched in beta‐intercalated cells, we discovered its ability to activate upon the extracellular treatment with mildly alkaline media. IRR is a close homolog of the receptors of insulin and insiln‐like growth factor but cannot be activated by to their agonists whereas IRR homologs do not respond to extracellular pH changes. This pH‐sensing property of IRR is evutionary conserved from amphibians to mammals. The IRR activation by alkali resembles a typical ligand‐receptor interaction since it is dose‐dependent and saturable, specific in relation to other homologous receptors, defined by the structure of the IRR ectodomain and accompanied by its conformational changes. Mutagenesis revealed two sites in the IRR ectodomain that act synergystically to induce the receptor activation. IRR activation triggers intracellular signaling resembling the insulin receptor pathway. We obtained a panel of monoclonal antibodies against the IRR ectodomain, one of them being able to activate the receptor, and another – to inhibit its response to alkaline media. Altogether, the obtained data suggest that IRR is a cellular pH sensor. We used kcnockout mice to address IRR systemic functions. The knockout mice are viable in normal conditions but fail to excrete excees bicarbonate upon experimentally induced alkalosis. Nevertheless, the increased blood bicarbonate is compensated by CO2 retention that keeps the blood pH within the physiological range. We performed comparative analyses of knockout and wild type mice kidney and brain transcriptomes and identified statistically valid changes in the expression of several proteins, in particular, ion transporters. Also, a behavioral phenotype of the knockout mice was revealed. They appeared to be less socially active and less agressive as compared to their wild‐type littermates.Support or Funding InformationThis work was financially supported by the Russian Science Foundation (grant № 14‐50‐00131)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.