Compartmentalization of ITAM and integrin signaling by adapter molecules

Summary:  Adapters are multidomain molecules that recruit effector proteins during signal transduction by immunoreceptors and integrins. The absence of these scaffolding molecules profoundly affects development and function of various hematopoietic lineages, underscoring their importance as regulators of signaling cascades. An emerging aspect of the mechanism by which engaged immunoreceptors and integrins transmit signals within the cell is by differential usage of various adapters that function to nucleate formation of distinct signaling complexes in a specific location within the cell. In this review, we discuss the mechanisms by which adapter proteins coordinate signal transduction with an emphasis on the role of subcellular compartmentalization in adapter function.     

In vitro regulation of beta1 and beta3 integrin subunits in endometrial epithelial cells from normal endometrium

OBJECTIVE: To evaluate the effect of prostaglandin E2 (PGE2) and interleukin-1beta (IL-1beta) on integrin expression. DESIGN Cultures of endometrial epithelial cells from normal endometrium. SETTING: All endometrial specimens were obtained from the Obstetrics and gynecology Department of the Catholic University, Rome, Italy. PATIENTS: Four patients were normal menstrual cycles undergoing operative laparoscopy for non-endometrial problems. INTERVENTION: Endometrial samples were collected by uterine courettings. MAIN OUTCOME MEASURE: Immunocytochemistry for beta1 and beta3 integrin subunits. RESULTS: PGE2 clearly enhances both beta1 and beta3 integrin subunit expression. IL-1beta seem to slightly increase only beta3 subunit expression. CONCLUSIONS: In light of the critical role played by eicosanoids in endometrial differentiation, we suggest that PGE2 is also involved in local paracrine regulation of integrin expression.     

The molecular assembly and organization of signaling active B-cell receptor oligomers

Summary:  In B cells, antigen drives the formation of B-cell receptor (BCR) clusters that initiate the formation of signaling complexes associated with the cytoplasmic domains of the BCRs. These signaling active complexes contain a number of protein and lipid kinases and phosphatases and adapter and scaffolding proteins that together function to trigger downstream signaling cascades leading to the activation of a variety of genes associated with B-cell activation. Although we are learning a considerable amount about the molecular details of the assembly of immune receptor signaling complexes, as reviewed in this volume, a fundamental question remains, namely how does antigen binding outside the cell initiate the assembly of signaling complexes inside the cell. For B cells, we do not yet understand how the information that the ectodomain of the BCR has bound to an antigen is translated across the membrane to induce changes in the cytoplasmic domains that trigger the assembly of signaling complexes. Here we describe what is known about the initiation of the antigen-driven BCR signal transduction in the newly emerging context of B-cell recognition of antigens presented by antigen-presenting cells in lymphoid tissues. We also discuss a recently proposed model for the initiation of BCR signaling termed the 'conformation-induced oligomerization model' and address the implications of this model for the mechanisms by which BCR signaling may be modulated by adapters and coreceptors.     

Inhibition of interferon induction and signaling by paramyxoviruses

Summary: The family Paramyxoviridae comprises a diverse group of viruses that includes several important human and veterinary pathogens. Members of this family have a non-segmented, single-stranded, negative sense RNA genome, a conserved gene order, and a similar replication strategy. Paramyxoviruses are divided into two subfamilies, Paramyxovirinae and Pneumovirinae, which comprise five genera and two genera, respectively. Viruses in each genus have developed strategies to circumvent the interferon (IFN) response by using a diverse array of proteins that are encoded within the phosphoprotein genes of the Paramyxovirinae or non-structural genes of the Pneumovirinae. This review focuses on the specific roles that these viral proteins play in the inhibition of IFN signaling and, to a lesser extent, on the mechanisms by which these proteins inhibit the induction pathways of IFN. An improved understanding of the interactions between viral proteins and the host innate immune response is critical to achieving a thorough comprehension of the pathogenesis of this important group of viruses. Hopefully this knowledge will support the development of more targeted vaccines and therapeutics to better prevent and control viral infection.     

The effects of live Streptococcus pneumoniae and tumor necrosis factor-α on neutrophil oxidative burst and β2-integrin expression

Objective   To study the effects of TNF- α and live Streptococcus pneumoniae on human neutrophil oxidative burst and β 2-integrin expression using flow cytometry.
Methods   Six clinical isolates of S. pneumoniae (serotypes 3, 19A, 22F, 6A, 33F and 9N) from patients with bacteremic pneumonia or upper respiratory tract infections were studied. Whole blood was incubated either alone, with TNF- α or with S. pneumoniae or with both TNF- α and pneumococci at a ratio of one neutrophil per 1–5 bacteria. After 30 min of incubation, the tubes were put into ice, fixed and analysed.
Results   S. pneumoniae caused an increase in oxidative burst but not greater than that caused by TNF- α alone. When whole blood was preincubated with TNF- α for 30 min before the addition of pneumococci, a further increase in the oxidative burst response was seen. The variation in CD11b expression was not significant. Both S. pneumoniae and TNF- α caused increases in CD18 expression. The addition of TNF- α directly with the bacteria caused no further increase, but preincubation of blood with TNF- α 30 min before the addition of the bacteria caused a significant increase in CD18 expression.
Conclusions   Live S. pneumoniae stimulates polymorphonuclear leukocytes to produce an oxidative burst and increases expression of CD18, and these effects are enhanced by TNF- α .     

Echistatin prevents posterior capsule opacification in diabetic rabbit model via integrin linked kinase signaling pathway

Purpose: To investigate the effect of disintegrin echistatin on integrin linked kinase (ILK) and subsequent PI3-K/Akt and ERK1/2 signaling pathways in the posterior capsule opacification (PCO) model of diabetic rabbit. Methods: 56 rabbits were injected alloxan to model diabetic. Then they accepted lens extraction surgery and randomly and intraoperatively injected distilled water (control group; n = 28) or 10.0 mg·L^-1 echistatin (echistatin-treated group; n = 28) into the anterior chamber. Each group was subdivided into ten days group (n = 14) and six weeks group (n = 14) respectively. The PCO severity was evaluated with a slit lamp microscope and light microscope for 10 days and 6 weeks postoperatively. The levels of ILK in the posterior capsule were determined by Q-PCR, Western blotting and Immunohistochemistry. Akt and ERK1/2 phosphorylation were analyzed by Western blotting. Results: 10 days and 6 weeks after surgery, the grades of PCO in the echistatin-treated group were lower than the control group. The lens epithelial cells (LECs) in the posterior capsule of echistatin-treated eyes had decreased degrees of proliferation and migration than the control group. And no significant side effects appeared after treated with echistatin. Echistatin could significantly reduce the expression of ILK in terms of both mRNA and protein levels. The phosphorylation levels of Akt and ERK1/2 were decreased in the echistatin-treated group compared with the control group. Conclusions: Echistatin could inhibit postoperative PCO occurrence and development in diabetic rabbit eyes, which may be related to down-regulation the expression of ILK and inhibition the PI3-K/Akt and ERK1/2 pathways.     

整合素调节Fak/SFKs与生长因子受体信号传导的研究进展

在胚胎发育和组织修复过程中,整合素介导的信号传导起着重要作用.整合素与细胞外配体结合,引发的细胞内信号可以产生两种主要作用,即肌动蛋白细胞骨架重构和调节细胞行为,如细胞存活、分化和生长.整合素也可以通过与细胞内衔接蛋白、胞质酪氨酸激酶、生长因子/细胞因子受体直接结合或功能联合,进入信号传导过程.了解有关整合素信号的最新实验和理论进展,尤其着重于整合素调节Fak/Src家族激酶（SFKs）活化和整合素与可溶性生长因子/细胞因子受体相互作用很有必要.     

The duplicitous nature of the Lyn tyrosine kinase in growth factor signaling

The Lyn tyrosine kinase is a unique member of the Src family of non-receptor protein tyrosine kinases whose principal role is to regulate signals through inhibitory receptors thereby promoting signal attenuation. Lyn is renowned for its role in B cell antigen receptor and FcεRI signaling; however, it is becoming increasingly apparent that Lyn also functions in signal transduction from growth factor receptors including the receptors for GM-CSF, IL-3, IL-5, SCF, erythropoietin, CSF-1, G-CSF, thrombopoietin and Flt3 ligand. Numerous studies have implicated Lyn in growth factor receptor signal amplification, while a number also suggest that Lyn participates in negative regulation of growth factor signaling. Indeed Lyn-deficient mice are hyper-responsive to myeloid growth factors and develop a myeloproliferative disorder that predisposes the mice to macrophage tumours, with loss of negative regulation through SHP-1 and SHIP-1 thought to be the major contributing factor to this phenotype. Developing a clear understanding of Lyn's role in establishing signaling thresholds in growth factor receptor signal amplification and signal inhibition may have important implications in the management of leukemias that may depend on Lyn activity.     

膀胱癌中整合素连接激酶及相关信号转导通路分子的表达及意义

目的 探讨整合素连接激酶(integrin linked kinase,ILK)及相关信号转导通路分子在膀胱癌中的表达及其临床意义.方法 采用免疫组化SP法检测47例膀胱癌及25例癌旁组织中ILK、AKT及β-catenin的表达.结果 ILK、AKT和β-eatenin在膀胱癌和癌旁组织中的阳性率分别为53.19％和12.00％、44.68％和20.00％、48.94％和12.00％,且差异均有显著性(x2=11.651,x2=4.309,x2=9.650,P＜0.05).同时,ILK、AKT及β-catenin在高分化膀胱癌组中的表达明显低于低分化癌组,差异有统计学意义(P＜0.05).AKT、β-catenin和ILK在膀胱癌中的表达均呈正相关.结论 ILK、AKT及β-catenin的异常表达在膀胱癌的恶性进展中起重要作用,三者联合检测可能为揭示膀胱癌发生、发展的有关机制提供理论依据.ILK、AKT及β-catenin的异常表达与膀胱癌的临床病理分级密切相关,可能参与膀胱癌的发生、发展,联合检测有助于判断膀胱癌的恶性程度.     

AMPK通过抑制STING调节干扰素免疫应答通路的研究

目的探索腺苷酸活化蛋白激酶(AMPK)与cGAS-STING通路之间的联系及其在先天免疫中扮演的角色。方法利用CRISPR/Cas9技术、蛋白质印迹、RT-qPCR等方法,探究AMPK对DNA相关免疫通路的调控机制。结果在HT-DNA和cGAMP刺激下,AMPK-/-细胞株的IFN-β的表达量明显高于野生型细胞株,但这种变化在RNA信号通路中并不明显;激活AMPK可以抑制细胞内的DNA信号通路;在DNA信号通路中,AMPK-/-细胞株相较于野生型细胞株,STING在RNA和蛋白水平上都明显升高,即AMPK对cGAS-STING通路的抑制很可能是通过抑制STING起作用。结论AMPK在调节cGAS-STING介导的干扰素免疫应答中起重要作用。     

Conformational snapshots of Tec kinases during signaling

Summary:  The control of cellular signaling cascades is of utmost importance in regulating the immune response. Exquisitely precise protein–protein interactions and chemical modification of substrates by enzymatic catalysis are the fundamental components of the signals that alert immune cells to the presence of a foreign antigen. In particular, the phosphorylation events induced by protein kinase activity must be spatially and temporally regulated by specific interactions to maintain a normal and effective immune response. High resolution structures of many protein kinases along with supporting biochemical data are providing significant insight into the intricate regulatory mechanisms responsible for controlling cellular signaling. The Tec family kinases are immunologically important kinases for which regulatory details are beginning to emerge. This review focuses on bringing together structural insights gained over the years to develop an understanding of how domain interactions both within the Tec kinases and between the Tec kinases and other signaling molecules control immune cell function.     

Autoinhibition and adapter function of Syk

Summary:  Development, survival, and activation of B lymphocytes are controlled by signals emanating from the B-cell antigen receptor (BCR). The BCR has an autonomous signaling function also known as tonic signaling that allows for long-term survival of B cells in the immune system. Upon binding of antigen to the BCR, the tonic signal is amplified and diversified, leading to alteration in gene expression and B-cell activation. The spleen tyrosine kinase (Syk) intimately cooperates with the signaling subunits of the BCR and plays a central role in the amplification and diversification of BCR signals. In this review, we discuss the molecular mechanisms by which Syk activity is inhibited and activated at the BCR. Importantly, Syk acts not only as a kinase that phosphorylates downstream substrates but also as an adapter that can bind to a diverse set of signaling proteins. Depending on its interactions and localization, Syk can signal opposing cell fate decisions such as proliferation or differentiation of B cells.     

IRAK-4 as the central TIR signaling mediator in innate immunity

Toll-like receptors (TLRs), which recognize pathogen-associated molecular patterns and members of the proinflammatory interleukin-1 receptor (IL-1R) family, share homologies in their cytoplasmic domains. Engagement of members of both of these families initiates a common intracellular signaling cascade, in which MyD88 and tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) are key adaptor proteins. Signaling between MyD88 and TRAF6 is mediated by members of the IL-1R-associated kinase (IRAK) family; however, the exact function of each IRAK protein remains controversial. IRAK-1 is required for the optimal transduction of IL-1R- and TLR-mediated signals, but IRAK-1 can be replaced by other IRAKs. Surprisingly, gene targeting studies show that the newest IRAK protein, IRAK-4, has an essential role in mediating signals initiated by IL-1R and TLR engagement. The kinase activity of IRAK-4 might be necessary to functionally modify IRAK-1 and perhaps other signal transducing substrates. Understanding the role of IRAK-4 in innate immunity will enable us to design novel strategies for therapeutic intervention in human infectious disease.     

Increased expression of integrin beta-4 in papillary thyroid carcinoma with gross lymph node metastasis

Although the prognosis is generally good for patients with papillary thyroid carcinoma, gross nodal metastasis of carcinoma has a poor prognosis. It is necessary to clarify how carcinoma progresses to gross nodal metastasis in order to establish a cure. The adhesion molecule integrin beta-4 is considered to be related to cell migration and metastasis in many carcinomas. In the present study, we examined integrin beta-4 expression in 65 cases of human papillary thyroid carcinoma using immunohistochemical methods. Expression of integrin beta-4 was found in all papillary carcinomas, but in few normal thyrocytes. Interestingly, integrin beta-4 expression in the carcinomas with gross (> or =3 cm) lymph node metastasis was significantly higher than that in carcinomas with small (<3 cm) or no lymph node metastasis. These results suggest that integrin beta-4 expression in thyroid carcinoma may play a role in the development of gross lymph node metastasis of papillary carcinomas.     

Novel Role and Regulation of the Interleukin-1 Receptor Associated Kinase （IRAK） Family Proteins

The interleukin-1 receptor associated kinases(IRAKs)sit at the bottle neck for the Toll-like-receptor(TLR)mediated signal transduetion process controlling host innate immune response.However,the exact role andregulation of IRAKs are still in the early stage and not fully understood.This review intends to summarize therecent advancement in this important topic and points out areas that need further intensive investigation.Cellular& Molecular Immunology.2005:2(1):36-39.     

Distinct signalling pathways promote phagocytosis of bacteria, latex beads and lipopolysaccharide in medfly haemocytes

In insects, phagocytosis is an important innate immune response against pathogens and parasites, and several signal transduction pathways regulate this process. The focal adhesion kinase (FAK)/Src and mitogen activated protein kinase (MAPK) pathways are of central importance because their activation upon pathogen challenge regulates phagocytosis via haemocyte secretion and activation of the prophenoloxidase (proPO) cascade. The goal of this study was to explore further the mechanisms underlying the process of phagocytosis. In particular, in this report, we used flow cytometry, RNA interference, enzyme-linked immunosorbent assay, Western blot and immunoprecipitation analysis to demonstrate that (1) phagocytosis of bacteria (both Gram-negative and Gram-positive) is dependent on RGD-binding receptors, FAK/Src and MAPKs, (2) latex bead phagocytosis is RGD-binding-receptor-independent and dependent on FAK/Src and MAPKs, (3) lipopolysaccharide internalization is RGD-binding-receptor-independent and FAK/Src-independent but MAPK-dependent and (4) in unchallenged haemocytes in suspension, FAK, Src and extracellular signal-regulated kinase (ERK) signalling molecules participating in phagocytosis show both a functional and a physical association. Overall, this study has furthered knowledge of FAK/Src and MAPK signalling pathways in insect haemocyte immunity and has demonstrated that distinct signalling pathways regulate the phagocytic activity of biotic and abiotic components in insect haemocytes. Evidently, the basic phagocytic signalling pathways among insects and mammals appear to have remained unchanged during evolution.     

Toll-like receptor signaling in neonatal sepsis and inflammation: a matter of orchestration and conditioning

Altered neonatal Toll-like receptor (TLR) function is hypothesized to contribute to the heightened susceptibility to infection and perpetuated inflammation in term and preterm neonates, clinically evident in neonatal sepsis and increased rates of inflammatory disorders. Current data indicate that basal TLR expression in term neonates equals adult expression patterns, while expression in preterm infants seems to increase, depending on gestational age. Regarding TLR signaling, some studies suggest TLR incompetence in neonates associated with impaired pro-inflammatory responses, others describe neonatal TLR function well developed and allude to its hyper-inflammation tendency. We discuss the competing positions and considerable limitations of research approaches and conclude that neonatal innate immunity is not generally less able to respond to TLR stimulation. Moreover, we describe pre-conditioning factors other than immaturity having a comparable impact. In the long term, better understanding of the complex interplay of pre- and postnatal conditions and maturation-dependent neonatal TLR function may provide new therapeutic approaches.     

Activation, coactivation, and costimulation of resting human natural killer cells

Summary:  Natural killer (NK) cells possess potent perforin- and interferon-γ-dependent effector functions that are tightly regulated. Inhibitory receptors for major histocompatibility complex class I display variegated expression among NK cells, which confers specificity to individual NK cells. Specificity is also provided by engagement of an array of NK cell activation receptors. Target cells may express ligands for a multitude of activation receptors, many of which signal through different pathways. How inhibitory receptors intersect different signaling cascades is not fully understood. This review focuses on advances in understanding how activation receptors cooperate to induce cytotoxicity in resting NK cells. The role of activating receptors in determining specificity and providing redundancy of target cell recognition is discussed. Using Drosophila insect cells as targets, we have examined the contribution of individual receptors. Interestingly, the strength of activation is not determined simply by additive effects of parallel activation pathways. Combinations of signals from different receptors can have different outcomes: synergy, no enhancement over individual signals, or additive effects. Cytotoxicity requires combined signals for granule polarization and degranulation. The integrin leukocyte function-associated antigen-1 contributes a signal for polarization but not for degranulation. Conversely, CD16 alone or in synergistic combinations, such as NKG2D and 2B4, signals for phospholipase-C-γ- and phosphatidylinositol-3-kinase-dependent degranulation.     

Post-synaptic density-93 mediates tyrosine-phosphorylation of the N-methyl-D-aspartate receptors

Src family protein kinases (SFKs) -mediated tyrosine-phosphorylation regulates N-methyl-d-aspartate (NMDA) receptor synaptic function. Some members of the membrane-associated guanylate kinase (MAGUK) family of proteins bind to both SFKs and NMDA receptors, but it is unclear whether the MAGUK family of proteins is required for SFKs-mediated tyrosine-phosphorylation of the NMDA receptors. Here, we showed by co-immunoprecipitation that post-synaptic density (PSD) -93, a member of the MAGUK family of proteins, interacts with the NMDA receptor subunits NR2A and NR2B as well as with Fyn, a member of the SFKs, in mouse cerebral cortex. Using a biochemical fractionation approach to isolate subcellular compartments revealed that the expression of Fyn, but not of other members of the SFKs (Lyn, Src, and Yes), was significantly decreased in synaptosomal membrane fractions derived from the cerebral cortex of PSD-93 knockout mice. Interestingly, we found that PSD-93 disruption causes reduction of tyrosine-phosphorylated NR2A and NR2B in the same fraction. Moreover, PSD-93 deletion markedly blocked the SFKs-mediated increase in tyrosine-phosphorylated NR2A and NR2B through the protein kinase C pathway after induction with 4-phorbol 12-myristate 13-acetate in cultured cortical neurons. Our findings indicate that PSD-93 appears to mediate tyrosine-phosphorylation of the NMDA receptors and synaptic localization of Fyn.     

Over-expression of integrin β3 can partially overcome the defect of integrin β3 signaling in transglutaminase 2 null macrophages

Transglutaminase 2 (TG2) is a protein crosslinking enzyme with many additional biological functions. We have previously shown that in TG2^−/− mice the in vivo clearance of apoptotic cells is defective leading to autoimmunity. TG2 contributes to the formation of phagocytic portals by binding to both integrin β₃, a known phagocytic receptor, and its bridging molecule, MFG-E8. In TG2 null macrophages integrin β₃ cannot accumulate around the apoptotic cells and its signaling is impaired. In the present study we describe a subline of TG2 null mice, in which a compensatory increase in integrin β₃ expression, which resulted alone in a high receptor concentration around the apoptotic cells without the requirement for accumulation, partially corrected the defect in integrin β₃ signaling. Our data provide a proof for the concept that the function of TG2 is to stabilize accumulated integrin β₃ concentration in the phagocytic cup.