Kindlin-3 in the immune system

Kindlin-3 is a member of the kindlin family of focal adhesion proteins which bind to integrin beta-chain cytoplasmic domains to regulate integrin function. In contrast to kindlin-1 and kindlin-2 proteins, kindlin-3 is expressed mainly in the hematopoietic system. Mutations in kindlin-3 result in the rare genetic disorder, leukocyte adhesion deficiency type III (LAD-III), which is characterized by bleeding and recurrent infections due to deficient beta1, beta2 and beta3 integrin activation in platelets and leukocytes. Here, we review the role of kindlin-3 in integrin activation and in different immune cell functions

Regulation of leukocyte integrin binding to Ig-family ligands

The adhesion molecules of blood cells are of great importance in the regulation of many of the most central processes of the human body, e.g. haematopoiesis, immune functions, haemostasis and wound healing, and the delivery of oxygen to the tissues. Leukocyte β2 integrins, VLA-4 integrin and members of the immunoglobulin superfamily like ICAMs (intercellular adhesion molecules) and VCAM (vascular cell adhesion molecule) are the most essential adhesion molecules of blood cells. The adhesion molecules on blood cells have many requirements that they need to fulfil in order to maintain a physiological system: they need to stay in an inactive, non-binding state for most of the time, and to be activated and thus become adhesive only when needed. In addition, they should specifically recognise their binding partners or ligands, as unnecessary binding could lead for example to clogging of the blood vessels, autoimmune diseases or allergic reactions. Still one important feature of blood cell adhesion is the ability to let go and release the adhesion, when the cell needs to move forward or continue patrolling the circulation. In my thesis work I have analysed the properties of leukocyte integrins and their ligands as well as the regulation of their interactions. We observed that the red cell adhesion molecule ICAM-4 can bind to CR4, a leukocyte integrin expressed on monocytes and macrophages, and that the I domain is the ICAM-4 binding site on leukocyte integrins (LFA-1, Mac-1 and CR4). We also characterised the phosphorylation of the cytoplasmic tail of CR4, and found that αX chain is phosphorylated on Ser1158, and that this phosphorylation is essential for CR4 inside-out activation, adhesion and phagocytosis but not for outside-in signaling initiated by CR4. Finally we analysed the regulation of VLA-4 mediated adhesion to VCAM-1 that is controlled by the β2 integrins. The findings of my studies show how leukocyte integrins are involved in numerous blood cell functions and that their functions are tightly regulated. Due to their multifold roles, they also offer attractive targets for therapeutic use. The specificity of phosphorylations or ligands may serve as distinctive factors between different integrins, even members of the same family.Veren solut hoitavat kukin omia tarkoin säädeltyjä tehtäviään elimistössä. Punasolut kuljettavat happea ja hiilidioksidia, verihiutaleet tukkivat verenvuodon ja valkosolut huolehtivat elimistön puolustusmekanismeista. Yhteistä näille kaikille soluille on että pääosan verenkierrossa viettämästään ajasta ne kulkevat huimaa vauhtia eteenpäin Toisaalta tärkeää solujen toiminnalle on pystyä tarvittaessa aistimaan muutoksia ympäristössään ja tunnistamaan tärkeitä rakenteita toisten solujen pinnalla tai solunulkoisessa materiaalissa. Myös tarttumista eli adheesiota tarvitaan jossain vaiheessa kaikkien verisolujen elämänkaaren aikana. Verisolujen tarttumistapahtumista huolehtivat suurelta osin soluadheesiomolekyylit, suuri proteiiniryhmä joka voidaan jakaa useampaan perheeseen molekyylin rakenteen ja tehtävän perusteella. Verisolujen adheesiotapahtumissa tärkeimpiä molekyylejä ovat integriinit sekä ICAM-perheen (intercellular adhesion molecules) proteiinit. Väitöskirjassani olen tutkinut näiden molekyylien välistä tarttumista sekä sen erityyppisiä säätelymekanismeja. Ensimmäisissä osajulkaisuissa selvitimme punasoluissa sekä niiden esiasteissa esiintyvän adheesiomolekyyli ICAM-4:n sitoutumista valkosolujen integriineihin ja löysimme ICAM-4:lle uuden tarttumiskumppanin, integriini CR4:n. Seuraavaksi tutkimme CR4:n toimintaan vaikuttavia solunsisäisiä mekanismeja. CR4-molekyylin on aiemmin raportoitu vaikuttavan useisiin valkosolujen tehtäviin kuten vanhentuneiden, infektoituneiden tai muuten viallisten solujen tuhoamiseen. Tutkimuksissamme huomasimme, että mikäli CR4:n solunsisäinen osa ei pysty fosforyloitumaan (siihen ei voida liittää fosfaattiryhmää), solun ulkopuolinen osa ei puolestaan pysty tarttumaan vastinmolekyyleihinsä. Viimeisessä osajulkaisussa tutkimme erityyppisten integriinien välistä vuoropuhelua saman solun sisällä. Huomasimme, että yhden integriinin fosforylaatio voi estää toisen, samassa solussa ilmentyvän integriinin tarttumista vastinmolekyyliinsä. Verisolujen tarttumistapahtumien mekanismien ja säätelyn tunnistaminen on tärkeää erilaisten sairauksien tutkimuksessa. Esim. sirppisoluanemiassa punasolut tarttuvat juurikin ICAM-4:n kautta liikaa verisuonen seinämiin, mikä aiheuttaa kiputiloja. Monissa autoimmuunisairauksissa (kuten MS-tauti, psoriasis ja Crohnin tauti) integriinit taas osallistuvat tulehdusreaktioihin joissa valkosolut hyökkäävät potilaan omia kudoksia vastaan, aiheuttaen vakavia vaurioita. Tarttumisen säätelyn tunteminen on ensiarvoisen tärkeää uusien hoitomenetelmien kehittämisessä

The versatile functions of complement C3-derived ligands

The complement system is a major component of immune defense. Activation of the complement cascade by foreign substances and altered self-structures may lead to the elimination of the activating agent, and during the enzymatic cascade, several biologically active fragments are generated. Most immune regulatory effects of complement are mediated by the activation products of C3, the central component. The indispensable role of C3 in opsonic phagocytosis as well as in the regulation of humoral immune response is known for long, while the involvement of complement in T-cell biology have been revealed in the past few years. In this review, we discuss the immune modulatory functions of C3-derived fragments focusing on their role in processes which have not been summarized so far. The importance of locally synthesized complement will receive special emphasis, as several immunological processes take place in tissues, where hepatocyte-derived complement components might not be available at high concentrations. We also aim to call the attention to important differences between human and mouse systems regarding C3-mediated processes. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Lt

Structural specializations of α4β7\alpha_4 \beta_7, an integrin that mediates rolling adhesion

The lymphocyte homing receptor integrin $\alpha_4 \beta_7$ is unusual for its ability to mediate both rolling and firm adhesion. $\alpha_4 \beta_1$ and $\alpha_4 \beta_7$ are targeted by therapeutics approved for multiple sclerosis and Crohn’s disease. Here, we show by electron microscopy and crystallography how two therapeutic Fabs, a small molecule (RO0505376), and mucosal adhesion molecule-1 (MAdCAM-1) bind α4β7. A long binding groove at the $\alpha_4 -\beta_7$interface for immunoglobulin superfamily domains differs in shape from integrin pockets that bind Arg-Gly-Asp motifs. RO0505376 mimics an Ile/Leu-Asp motif in $\alpha_4$ ligands, and orients differently from Arg-Gly-Asp mimics. A novel auxiliary residue at the metal ion–dependent adhesion site in $\alpha_4 \beta_7$ is essential for binding to MAdCAM-1 in $Mg^{2+}$ yet swings away when RO0505376 binds. A novel intermediate conformation of the $\alpha_4 \beta_7$ headpiece binds MAdCAM-1 and supports rolling adhesion. Lack of induction of the open headpiece conformation by ligand binding enables rolling adhesion to persist until integrin activation is signaled

Heparan Sulfate: A Ubiquitous Glycosaminoglycan with Multiple Roles in Immunity

Heparan sulfate (HS) is a highly acidic linear polysaccharide with a very variable structure. It is ubiquitously expressed on cell surfaces and in the extracellular matrix and basement membrane of mammalian tissues. Synthesized attached to various core proteins to form HS-proteoglycans, HS is capable of interacting with various polypeptides and exerting diverse functions. In fact, a bioinformatics analysis of mammalian proteins that express a heparin/HS-binding motif and are associated with the immune system identified 235 candidate proteins, the majority having an intracellular location. This simple analysis suggests that HS may, in fact, interact with many more components of the immune system than previously realized. Numerous studies have also directly demonstrated that HS plays multiple prominent functional roles in the immune system that are briefly reviewed in this article. In particular, the molecule has been shown to regulate leukocyte development, leukocyte migration, immune activation, and inflammatory processes

Structural Correlates of Rotavirus Cell Entry

Cell entry by non-enveloped viruses requires translocation into the cytosol of a macromolecular complex—for double-strand RNA viruses, a complete subviral particle. We have used live-cell fluorescence imaging to follow rotavirus entry and penetration into the cytosol of its ∼700 Å inner capsid particle (“double-layered particle”, DLP). We label with distinct fluorescent tags the DLP and each of the two outer-layer proteins and track the fates of each species as the particles bind and enter BSC-1 cells. Virions attach to their glycolipid receptors in the host cell membrane and rapidly become inaccessible to externally added agents; most particles that release their DLP into the cytosol have done so by ∼10 minutes, as detected by rapid diffusional motion of the DLP away from residual outer-layer proteins. Electron microscopy shows images of particles at various stages of engulfment into tightly fitting membrane invaginations, consistent with the interpretation that rotavirus particles drive their own uptake. Electron cryotomography of membrane-bound virions also shows closely wrapped membrane. Combined with high resolution structural information about the viral components, these observations suggest a molecular model for membrane disruption and DLP penetration

Beta2-Integrins and Interacting Proteins in Leukocyte Trafficking, Immune Suppression, and Immunodeficiency Disease

Beta2-integrins are complex leukocyte-specific adhesion molecules that are essential for leukocyte (e.g., neutrophil, lymphocyte) trafficking, as well as for other immunological processes such as neutrophil phagocytosis and ROS production, and T cell activation. Intriguingly, however, they have also been found to negatively regulate cytokine responses, maturation, and migratory responses in myeloid cells such as macrophages and dendritic cells, revealing new, and unexpected roles of these molecules in immunity. Because of their essential role in leukocyte function, a lack of expression or function of beta2-integrins causes rare immunodeficiency syndromes, Leukocyte adhesion deficiency type I, and type III (LAD-I and LAD-III). LAD-I is caused by reduced or lost expression of beta2-integrins, whilst in LAD-III, beta2-integrins are expressed but dysfunctional because a major integrin cytoplasmic regulator, kindlin-3, is mutated. Interestingly, some LAD-related phenotypes such as periodontitis have recently been shown to be due to an uncontrolled inflammatory response rather than to an uncontrolled infection, as was previously thought. This review will focus on the recent advances concerning the regulation and functions of beta2-integrins in leukocyte trafficking, immune suppression, and immune deficiency disease.Peer reviewe