Biophysical Characterization of Oxidized Phospholipids : Implications for Altered Membrane Structure and Function

The present study aims to elucidate the modifications in the structure and functionality of the phospholipid matrix of biological membranes brought about by free radical-mediated oxidative damage of its molecular constituents. To this end, the surface properties of two oxidatively modified phospholipids bearing an aldehyde or carboxyl function at the end of truncated sn-2 acyl chain were studied using a Langmuir balance. The results obtained reveal both oxidized species to have a significant impact on the structural dynamics of phospholipid monolayers, as illustrated by the progressive changes in force-area isotherms with increasing mole fraction of the oxidized lipid component. Moreover, surface potential measurements revealed considerable modifications in the electric properties of oxidized phospholipid containing monolayers during film compression, suggesting a packing state-controlled reorientation of the intramolecular electric dipoles of the lipid headgroups and acyl chains. Based on the above findings, a model describing the conformational state of oxidized phospholipid molecules in biological membranes is proposed, involving the protrusion of the acyl chains bearing the polar functional groups out from the hydrocarbon phase to the surrounding aqueous medium. Oxidative modifications alter profoundly the physicochemical properties of unsaturated phospholipids and are therefore readily anticipated to have important implications for their interactions with membrane-associating molecules. Along these lines, the carboxyl group bearing lipid was observed to bind avidly the peripheral membrane protein cytochrome c. The binding was reversed following increase in ionic strength or addition of polyanionic ATP, thus suggesting it to be driven by electrostatic interactions between cationic residues of the protein and the deprotonated lipid carboxyl exposed to the aqueous phase. The presence of aldehyde function bearing oxidized phospholipid was observed to enhance the intercalation of four antimicrobial peptides into phospholipid monolayers and liposomal bilayers. Partitioning of the peptides to monolayers was markedly attenuated by the aldehyde scavenger methoxyamine, revealing it to be mediated by the carbonyl moiety possibly through efficient hydrogen bonding or, alternatively, formation of covalent adduct in form of a Schiff base between the lipid aldehydes and primary amine groups of the peptide molecules. Lastly, both oxidized phospholipid species were observed to bind with high affinity three small membrane-partitioning therapeutic agents, viz. chlorpromazine, haloperidol, and doxorubicin. In conclusion, the results of studies conducted using biomimetic model systems support the notion that oxidative damage influences the molecular architecture as well as the bulk physicochemical properties of phospholipid membranes. Further, common polar functional groups carried by phospholipids subjected to oxidation were observed to act as molecular binding sites at the lipid-water interface. It is thus plausible that oxidized phospholipid species may elicit cellular level effects by modulating integration of various membrane-embedded and surface-associated proteins and peptides, whose conformational state, oligomerization, and functionality is known to be controlled by highly specific lipid-protein interactions and proper physical state of the membrane environment.Solujen kalvorakenteet koostuvat lipidi- ja proteiinimolekyyleistä sekä osaan näistä kovalenttisesti kiinnittyneistä hiilihydraattiosista. Kalvot reunustavat solulimaa sekä solun sisäisiä organelleja ja ylläpitävät näiden kemiallista koostumusta säätelemällä molekyylien kulkeutumista sisä- ja ulkopuolisten nestetilojen välillä. Lisäksi ne toimivat erityyppisten soluviestintään osallistuvien molekyylien kohteina ja varastoina sekä tarjoavat alustan useille keskeisille biokemiallisille prosesseille. Kalvojen rakenteellinen runko muodostuu seoksesta amfipaattisia fosfolipidimolekyylejä, jotka organisoituvat vesiliuoksissa spontaanisti kaksoiskerrokseksi. Fosfolipidien tiedetään jakaantuvan muodostamissaan kalvoissa koostumukseltaan erilaistuneisiin alueisiin, ja on osoitettu, että tämä heterogeeninen järjestäytyminen on vahvasti kytkeytynyt kalvorakenteiden toiminnallisiin ominaisuuksiin. Valtaosa aitotumallisten solujen kalvojen sadoista erityyppisistä fosfolipideistä kantaa tyydyttymättömiä rasvahappoketjuja. Nämä ovat alttiita solumetabolian sivutuotteina syntyvien reaktiivisten happiyhdisteiden välittämille hapetusreaktioille, joissa ketjut lyhenevät ja niihin muodostuu erityyppisiä polaarisia funktionaalisia ryhmiä. Tämän tutkimuksen päämääränä oli kartoittaa hapettuneiden fosfolipidien ominaisuuksia biofysikaalisia metodeja ja biomimeettisiä mallikalvosysteemejä hyödyntäen sekä selvittää hapetusreaktioiden mukanaan tuomia muutoksia biologisten kalvojen rakenteessa ja toiminnassa. Tutkittaessa kahden hapettuneen fosfolipidin käyttäytymistä veden ja ilman väliselle rajapinnalle muodostetuissa ns. Langmuir-kalvoissa havaittiin, että molekyylien aldehydi- tai karboksyyliryhmän sisältävillä hapettuneilla rasvahappoketjuilla on taipumus siirtyä kalvon hydrofobisesta ytimestä ympäröivään vesifaasiin, mahdollistaen ketjujen toimimisen kiinnittymiskohtina kalvorakenteisiin assosioituville molekyyleille. Pintakemiallisia ja fluoresenssispektroskooppisia tekniikoita hyödyntäen saadut tulokset osoittivat, että karboksyyliryhmää kantava lipidi sitoo voimakkaasti periferaalista kalvoproteiini sytokromi c:tä elektrostaattisten interaktioiden seurauksena. Lisäksi havaittiin, että tällä fosfolipidillä on jopa 100-kertainen affiniteetti eräisiin amfipaattisiin lääkeaineisiin (klooripromatsiini, haloperidoli ja doksorubisiini) ei-hapettuneisiin molekyyleihin verrattuna. Aldehydiryhmän sisältämä hapettunut fosfolipidi puolestaan lisäsi merkittävästi neljän antimikrobiaalisen peptidin sitoutumista Langmuir-kalvoihin sekä kalvovesikkeleihin. Väitöskirjatyön tulokset valottavat sitä, kuinka biologisten kalvojen fosfolipidimolekyylien hapettuminen voi johtaa muutoksiin kalvorakenteiden fysikokemiallisissa ominaisuuksissa, joilla nykytiedon valossa on keskeinen merkitys kalvoproteiinien lokalisaation, kolmiulotteisen rakenteen sekä aktiivisuuden säätelyssä. Hapettuneiden fosfolipidien lisääntynyt muodostus on yhdistetty solujen ikääntymiseen sekä eräiden kertymäsairauksien, kuten Alzheimerin taudin ja ateroskleroosin patogeneesiin, joskin taustalla vaikuttavat molekyylitason mekanismit tunnetaan toistaiseksi huonosti. Esitettyjen tulosten perusteella voidaan olettaa, että molekyylien lisääntyneestä hapettumisesta seuraavat muutokset fosfolipidimatriisin kemiallisessa koostumuksessa heijastuvat kokonaisvaltaisesti kalvorakenteiden pinnoilla tapahtuviin fysiologisiin prosesseihin ja saattavat siten olla yhteydessä näiden sekä koko solun toiminnan häiriöihin

Characterization of the main transition of dinervonoylphosphocholine liposomes by fluorescence spectroscopy

AbstractThe structural dynamics of the main phase transition of large unilamellar dinervonoylphosphocholine (DNPC) vesicles was investigated by steady state and time-resolved fluorescence spectroscopy of the membrane incorporated fluorescent lipid analog, 1-palmitoyl-2[10-(pyren-1-yl)]decanoyl-sn-glycero-3-phosphocholine (PPDPC). These data were supplemented by differential scanning calorimetry (DSC) and fluorescence anisotropy measured for 1-palmitoyl-2-(3-(diphenylhexatrienyl) propanoyl)-sn-glycero-3-phosphocholine (DPHPC). The collected data displayed several discontinuities in the course of the main transition and the pretransition. The discontinuities seen in the fluorescence properties may require modification of the existing models for phospholipid main transition as a first order process. From our previous study on dipalmitoylphosphocholine (DPPC), we concluded the transition to involve a first-order process resulting in the formation of an intermediate phase, which then converts into the liquid crystalline state by a second order process. Changes in the physical properties of the DNPC matrix influencing probe behavior were similar to those reported previously for PPDPC in DPPC. In gel state DNPC [(T−Tm)<−10] the high values for excimer/monomer emission ratio (Ie/Im) suggest enrichment of the probe in clusters. In this temperature range, excimer fluorescence for PPDPC (mole fraction XPPDPC=0.02) is described by two formation times up to (T−Tm)≈−10, with a gradual disappearance of the fractional intensity (IR1) of the shorter formation time (τR1) with increasing temperature up to (T–Tm)≈−10. This would be consistent with the initiation of the bilayer melting at the PPDPC clusters and the subsequent dispersion of the one population of PPDPC domains. A pronounced decrement in Ie starts at (T–Tm)=−10, continuing until Tm is reached. No decrease was observed in fluorescence quantum yield in contrast to our previous study on DPPC/PPDPC large unilamellar vesicles (LUVs) [J. Phys. Chem., B 107 (2003) 1251], suggesting that a lack of proper hydrophobic mismatch may prevent the formation of the previously reported PPDPC superlattice. With further increase in temperature and starting at (T−Tm)≈−1, Ie, τR2, and excimer decay times (τD) reach plateaus while increment in trans→gauche isomerization continues. This behavior is in keeping with an intermediate phase existing in the temperature range −1<(T−Tm)<4 and transforming into the liquid disordered phase as a second order process, the latter being completed when (T−Tm)→4 and corresponding to ≈50% of the total transition enthalpy

Missing-in-metastasis and IRSp53 deform PI(4,5)P2-rich membranes by an inverse BAR domain–like mechanism

The actin cytoskeleton plays a fundamental role in various motile and morphogenetic processes involving membrane dynamics. We show that actin-binding proteins MIM (missing-in-metastasis) and IRSp53 directly bind PI(4,5)P2-rich membranes and deform them into tubular structures. This activity resides in the N-terminal IRSp53/MIM domain (IMD) of these proteins, which is structurally related to membrane-tubulating BAR (Bin/amphiphysin/Rvs) domains. We found that because of a difference in the geometry of the PI(4,5)P2-binding site, IMDs induce a membrane curvature opposite that of BAR domains and deform membranes by binding to the interior of the tubule. This explains why IMD proteins induce plasma membrane protrusions rather than invaginations. We also provide evidence that the membrane-deforming activity of IMDs, instead of the previously proposed F-actin–bundling or GTPase-binding activities, is critical for the induction of the filopodia/microspikes in cultured mammalian cells. Together, these data reveal that interplay between actin dynamics and a novel membrane-deformation activity promotes cell motility and morphogenesis

Molecular Mechanisms of Membrane Deformation by I-BAR Domain Proteins

SummaryBackgroundGeneration of membrane curvature is critical for the formation of plasma membrane protrusions and invaginations and for shaping intracellular organelles. Among the central regulators of membrane dynamics are the BAR superfamily domains, which deform membranes into tubular structures. In contrast to the relatively well characterized BAR and F-BAR domains that promote the formation of plasma membrane invaginations, I-BAR domains induce plasma membrane protrusions through a poorly understood mechanism.ResultsWe show that I-BAR domains induce strong PI(4,5)P2 clustering upon membrane binding, bend the membrane through electrostatic interactions, and remain dynamically associated with the inner leaflet of membrane tubules. Thus, I-BAR domains induce the formation of dynamic membrane protrusions to the opposite direction than do BAR and F-BAR domains. Strikingly, comparison of different I-BAR domains revealed that they deform PI(4,5)P2-rich membranes through distinct mechanisms. IRSp53 and IRTKS I-BARs bind membranes mainly through electrostatic interactions, whereas MIM and ABBA I-BARs additionally insert an amphipathic helix into the membrane bilayer, resulting in larger tubule diameter in vitro and more efficient filopodia formation in vivo. Furthermore, FRAP analysis revealed that whereas the mammalian I-BAR domains display dynamic association with filopodia, the C. elegans I-BAR domain forms relatively stable structures inside the plasma membrane protrusions.ConclusionsThese data define I-BAR domain as a functional member of the BAR domain superfamily and unravel the mechanisms by which I-BAR domains deform membranes to induce filopodia in cells. Furthermore, our work reveals unexpected divergence in the mechanisms by which evolutionarily distinct groups of I-BAR domains interact with PI(4,5)P2-rich membranes

Unexpectedly High Prevalence of Common Variable Immunodeficiency in Finland

Background: Common variable immunodeficiency (CVID) is the most common primary immunodeficiency. Prevalence varies greatly between countries and studies. Most diagnostic criteria include hypogammaglobulinemia and impaired vaccine response. Aim: To evaluate the minimum prevalence as well as the clinical and immunological phenotypes of CVID in Southern Finland. Methods: We performed a cross-sectional study to assess all adult CVID patients followed up in three hospital districts in Southern and South-Eastern Finland between April 2007 and August 2015. CVID diagnosis was based, with a minor modification, on the ESID/PAGID criteria for primary CVID. Antipolysaccharide responses to Pneumovax (R) were defined as impaired only if 50% or more of the serotypes did not reach a level of 0.35 mu g/mL after vaccination. We further characterized the patients' B cell phenotypes and complications associated with CVID. Results: In total, 9 patients were excluded due to potential secondary causes before diagnosis. ESID/PAGID criteria were met by 132 patients (males 52%), of whom, 106 had "probable" and 26 "possible CVID." Based on the population statistics in the three hospital districts, the minimum adult prevalence per 100,000 inhabitants in Finland for all CVID ("probable CVID," respectively) patients was 6.9 (5.5). In the highest prevalence district (Helsinki and Uusimaa), the prevalence was 7.7 (6.1). CVID patients suffer from frequent complications. Ten patients died during follow-up. Of probable CVID patients, 73% had more than one clinical phenotype. Intriguingly, gradual B cell loss from peripheral blood during follow-up was seen in as many as 16% of "Xprobable CVID" patients. Patients with possible CVID displayed somewhat milder clinical and laboratory phenotypes than probable CVID patients. We also confirm that large granular lymphocyte lymphoproliferation is a CVID-associated complication. Conclusion: The prevalence of CVID in Finland appears the highest recorded, likely reflecting the genetic isolation and potential founder effects in the Finnish population. Studies to discover potential gene variants responsible for the high prevalence in Finland thus seem warranted. Increased awareness of CVID among physicians would likely lead to earlier diagnosis and improved quality of care.Peer reviewe

Best food - good life

Visio tulevaisuuden ruokajärjestelmästä vastaa niin globaaleihin kuin paikallisiinkin haasteisiin, jotka johtuvat ilmastonmuutoksesta, ekosysteemipalvelujen heikkenemisestä, resurssien niukkenemisesta, ruoan ja energian lisääntyvästä tarpeesta, nopeasta sosiaalisesta eriarvoistumisesta, maahanmuutosta ja kansalaisten etääntymisestä ruoan alkuperästä. Visio ottaa kantaa ruoan huoltovarmuuteen kansallisesti ja ruokaan globaalina ihmisoikeutena. Visiossa hahmotellaan tulevaisuuden ruokajärjestelmä ja keinot sen toteuttamiseksi viiden seuraavan kokonaisuuden avulla. 1. Tavoitteena ruokaturva Kotimainen ruokaturva perustuu kannattavaan ja ympäristöystävälliseen, paikalliseen ja verkottuneeseen tuotantojärjestelmään, joka on tärkeimpien tuotannontekijöitten suhteen kansallisesti omavarainen. Järjestelmän perusominaisuuksiin kuuluu kierrätys ja vähäiset hävikit. Ruokajärjestelmä pystyy takaamaan kansalaisille kaikissa olosuhteissa ravitsemuksellisesti riittävän ruoan, samalla kun se tuottaa muita biotuotannon raaka-aineita ja energiaa. Ruokajärjestelmälle on annettu yhteisöjä vahvistava tehtävä, ja sen sosiaalipoliittinen merkitys on huomattu. 2. Kuluttamisesta osallistumiseen Osallistava ruokajärjestelmä kytkee jokaisen ihmisen ruoantuotantoon ja lisää näin tietoisuutta sekä arvostusta ruoan tuottamisesta ja ympäristöstä. Ruokakansalainen on tietoinen ruoan laadun eri ulottuvuuksista. Osallistuminen mm. lähiruokayhteisöihin luo merkityksellisyyttä ja hyvinvointia. 3. Ihmisten yhteisöjä ja luonnon resursseja vaalien Tehokkain ekologisin menetelmin viljelty maa tuottaa ravitsemuksellisesti korkeita ja varmoja satoja. Viljelymaat sitovat hiiltä maaperään ja hidastavat ilmastonmuutosta. Tasapainoinen tuotantorakenne on mahdollistanut kierrätysmaatalouden: rehu- ja ravinnevirrat ovat paikallisia. Maatalousmaan väheneminen on estetty maankäytön kokonaisvaltaisella suunnittelulla, ja maanvuokrausta on kehitetty pitkäjänteiseksi mahdollistamaan investoinnit maan laatuun. Maatalouden ja elintarvikejalostuksen paikalliset, integroidut järjestelmät ovat tuoneet ruoan huoltovarmuuden kannalta merkityksellistä sitkeyttä ja vakautta, ne mahdollistavat tehokkaan uusiutuvan energian tuotannon sekä ravinteiden kierrätyksen. Maaseudun yhteisöt ovat vahvistuneet ja niiden yhteydet kaupunkialueisiin konkretisoituneet. Maaseutu on jälleen paikka työskennellä, harrastaa, asua ja elää. 4. Ruoantuotanto turvaa ja tuottaa ekosysteemipalveluja Uudet tuotantotavat nojaavat ekosysteemipalveluihin. Ruoka ymmärretään ekosysteemipalveluna, ja viljely-ympäristöjen monimuotoinen eliöstö ymmärretään ekosysteemipalvelujen perustaksi. Ekosysteeminen ajattelu huomioidaan läpäisevästi niin perus- ja ammattiopetuksessa kuin maankäytön suunnittelussakin. Ruoan lisäksi viljelijät luonnon- ja maisemanhoitajina ansaitsevat muiden ekosysteemipalvelujen tuottamisesta sekä monimuotoisuuden vaalimisesta. 5. Maatalouspolitiikasta ruokapolitiikkaan Uusi ruokapolitiikka vähentää maataloustuotannon tukiriippuvuutta ja kannustaa ekologisesti tehokkaaseen korkeatuottoiseen, korkea-arvoisen ruoan tuotantoon. Politiikkatoimin on ohjattu ruokajärjestelmä toteuttamaan paitsi ravitsemuksellisia ja terveydellisiä, myös ympäristöllisiä, sosiaalisia ja taloudellisia tavoitteita. Ruokaturva, oikeus hyvään ravitsemukseen on nostettu ruokapolitiikan perustaksi, ja Suomi edistää tätä näkemystä kansainvälisesti

A molecular-based identification resource for the arthropods of Finland

Publisher Copyright: © 2021 The Authors. Molecular Ecology Resources published by John Wiley & Sons Ltd.To associate specimens identified by molecular characters to other biological knowledge, we need reference sequences annotated by Linnaean taxonomy. In this study, we (1) report the creation of a comprehensive reference library of DNA barcodes for the arthropods of an entire country (Finland), (2) publish this library, and (3) deliver a new identification tool for insects and spiders, as based on this resource. The reference library contains mtDNA COI barcodes for 11,275 (43%) of 26,437 arthropod species known from Finland, including 10,811 (45%) of 23,956 insect species. To quantify the improvement in identification accuracy enabled by the current reference library, we ran 1000 Finnish insect and spider species through the Barcode of Life Data system (BOLD) identification engine. Of these, 91% were correctly assigned to a unique species when compared to the new reference library alone, 85% were correctly identified when compared to BOLD with the new material included, and 75% with the new material excluded. To capitalize on this resource, we used the new reference material to train a probabilistic taxonomic assignment tool, FinPROTAX, scoring high success. For the full-length barcode region, the accuracy of taxonomic assignments at the level of classes, orders, families, subfamilies, tribes, genera, and species reached 99.9%, 99.9%, 99.8%, 99.7%, 99.4%, 96.8%, and 88.5%, respectively. The FinBOL arthropod reference library and FinPROTAX are available through the Finnish Biodiversity Information Facility (www.laji.fi) at https://laji.fi/en/theme/protax. Overall, the FinBOL investment represents a massive capacity-transfer from the taxonomic community of Finland to all sectors of society.Peer reviewe

Genetic variation in English oak (Quercus robur) in Finland.

Genetic variation in 5 natural stands of Quercus robur in Finland was analyzed electrophoretically for 13 isozyme loci. Stands were on average polymorphic at 49.2% of the loci, with 2.1 alleles per locus. Observed heterozygosities, ranging from 13.6% to 16.9%, were slightly lower than estimates reported for German stands. The majority of the species’ genetic variation was found within each studied stand, and only 5.5% was between stands. Mean genetic differentiation was the same as that found in the primary range of the species, but the differentiation estimates for single Finnish populations were more variable