Last Three Days of the Liposomes, Exosomes and Virosomes Meeting … a sunny afternoon off, a couple of thunderstorms and a late night dancing party

ascona2Over the last three days of the liposomes, exosomes and virosomes meeting in the lovely Monte Verita’, Ascona, we had the opportunity to attend a series of very interesting talks and to have just a little bit of fun too. Here is a “brief” summary of the notes Marina and I were taking on the talks (disclaimer: you might find some inaccuracy; spoiler alert: this post is unbelievably very long):

Wolfgang Meier showed how biomimetic polymer and peptide membranes made of amphiphilic block copolymers form nanotubes, vesicle containers, cubic gels and lamellar gels. In particular block copolymer vesicles with a maximum diameter of 100 nm and a thickness of 10.9 nm are thicker and more stable than lipid vesicles. They can be used as a nanoparticle platform for the drug delivery of the hydrophobic anticancer drug doxorubicin and the serine hydroxymethil transferase inhibitor to treat malaria. Interestingly, the insertion of membrane proteins in block copolymer membranes does not affect their activity and function. The lateral mobility of proteins inside the more viscous block copolymer membranes is comparable to the lateral mobility of polymers, which is ten times slower than that of lipids in lipid bilayers. Block copolymers membranes are thicker and more stable than lipid membranes. Mixed block copolymer:DPPC monolayers and bilayers can also be produced with the formation of lipid domains due to the big thickness mismatch between the block copolymer (50 nm) and lipid (3 nm) domains. The insertion of a transmembrane protein in mixed systems occurs in different domains depending on the composition of the system: in systems with DPPC:DPPE, POPC and DOPC in the block copolymer domain, in both domains and in the lipid domain, respectively.

Andreas Kuhn showed different pathways for the membrane insertion of single proteins in bacteria, in particular the stepwise insertion of the Pf3 coat protein involving binding, transmembrane conformation and release. He proposed a two step insertion mechanism for the insertion of a Pf3 coat protein into a lipid bilayer, involving the binding to a groove and a greasy slide of the protein followed by bilayer insertion. He also talked about the purification and reconstitution of YidC protein in liposomes to form proteoliposomes with the cytoplasmisc domain of the protein in the outside or inside of the liposome. Fluorescent labelled Pf3 at the N- and C-terminii added to liposomes and proteoliposomes can be used to investigate protein translocation of single molecules with confocal microscope and fluorescence: fluorescent labelled Pf3 inserts into YidC proteoliposomes but not in liposomes and its C-terminal region is not translocated if labelled.

Sonia Troeira Henriques talked about the interaction of cyclic peptides Kalata B1, which are expressed in plants, with POPC and POPE model membranes. The presence of PE enhances the amount of peptides bound to the lipid bilayer because of a specific binding with PE lipids. She also showed that single lysine mutants have stronger haemolytic and insecticidal activity and the internalisation of the peptide labelled with Alexa and pHRodo fluorescent labels at different temperatures in endosomes. At 4 oC the internalisation is lost because the membrane is more rigid. The presence of the dynamin inhibitor peptide enhances the interaction with PE and internalisation. The insertion of the peptide into the membrane of GUVs is shown by solid state NMR.

Burkhard Bechinger showed how the amphipathic peptide Htt17, the membrane anchor of Huntingtin, which is a more than 3500 aa long protein involved in Huntington’s disease, is associated with POPC and other lipid bilayers through 15N and 2H NMR. He also showed that the extension of this peptide with Q17 forms aggregate only in the presence of vesicles and that the presence of more glutamines induces a faster aggregation leading to the formation of fibers observed in the disease.

Daniel Müller talked about the application of single molecule atomic force microscopy (AFM) to generate high resolution AFM images in biology showing how force distance curve based AFM imaging of biological membranes topography, adhesion and other processes, such as ligand-receptor binding, can be studied. In particular he showed that the folding of the membrane protein LacY, which is present in Escherichia coli inner and outer membranes, can be investigated by its mechanical unfolding from the C-terminus. LacY unfolds via structural segments as shown by force distance spectra, in which the probability of misfolding, unfolding, and folding events can be estimated, the folding probability being higher in the presence of YidC. Free energy profiles show that the misfolding process is associated with a high free energy barrier. Unfolded and extracted LacY cannot self insert into the membrane, while they can in the presence of YidC. Moreover, PE depletion leads to a topological change in LacY. ascona7

Christian Eggeling described how super resolution optical microscopy can be used to study lipid rafts/nanodomains, which are transient and smaller than 200 nm, using fluorescence labelled lipids and fluorescence correlation spectroscopy (FCS) to determine transit times for different areas. STED-FCS can be also employed to measure apparent diffusion coefficients for the lipids and highlight molecular interactions. He also talked about live cell model systems, such as giant plasma membrane vesicles (GPMVs), in which diffusion is not hindered as in cells, and the hopping of lipids proposed by Kusumi.

Phyllis Hanson gave a beautiful overview of the role of ESCRT III in the formation of membrane tubules and in the exocytosis of viral particles from infected cells. She showed us results using electron microscopy, mutational analysis, and simulation of the physical principles of coils to understand the coiling and three dimensional growth of this complex. ESCRT III creates ‘fences’ in the membrane. The diameter and the handedness of the coil varies depending on the proteins forming the complex.

On Wednesday afternoon the organizers let us take a well deserved break 🙂 I spent it going to Ascona via the steep stairs of the picture, enjoying as many views of Lake Maggiore as possible and walking around the historical centre for a couple of hours. Then, on my way back my heart realised how steep the stairs were and I met Robert Vácha, a colleague I met at the Biophysical Society Annual Meeting held in Philadelphia three years ago, who was starting his steepest decent, here is an inside joke for computational chemists (sorry I couldn’t resist).. I tried to convince him that I needed to be rescued, but he said that it was good for my health and he was right. Other attendees were more adventurous than me, for example my roommate, Marek Cebecauer, managed to go to Ascona and to hike a nearby mountain in the same day. At dinner I had the pleasure to talk to Karin Norling, who actually worked on the afternoon, and my roommate. After dinner I had a lovely chat with Helen Saibil, who gave a very interesting talk on toxins and pore forming proteins a couple of days before, about her love for Sardinia, its prehistoric nuragic age, an EMBO conference she organized in a beautiful place in my region three years ago and other things … and while looking for the website of that conference I’ve just found out that a thematically related EMBO conference was being held in Sardinia in the same week of the meeting we’ve just attended.

Anne Kenworthy showed how the cooperation of lipids and proteins is involved in the formation of liquid ordered (Lo) membrane domains. In particular she talked about the preferential interaction of cholera AB5 bacterial toxin, CTxB, with Lo domains in a more efficient manner when it is in its pentameric form rather than in its monomeric form. Pentameric CTxB reorganises membrane domain structure in model membranes and plasma membrane blebs affecting their phase behaviour, enhancing raft association and stabilising raft-like domains. She also introduced another interesting property of toxins, such as shiga toxin, which is the ability to induce their own uptake by bending membranes forming tubulations. CTxB tubules have a similar orientation of microtubules in cells. Microtubules and motors like dynein bend the plasma membrane during endocytosis.ascona6

Horst Vogel talked about the structure and function of the pentameric ligand-gated ion channel (pLGIC) 5HT3R, which has the neurotransmitter serotonin, showing its purification, ligand binding and CD spectroscopy studies at different temperatures. The crystal structure of 5HT3R in complex with VHH15 has the neurotransmitter serotonin in its binding site and a 0.45 nm hydrophobic gate at the centre of the transmembrane domain. The structures of 5HT3R in lipid vesicles and bilayers revealed by cryo EM are similar. Molecular dynamics simulations of the ligand associated in the binding site show that hydrophobic residues, such as tyrosine and phenylalanine, undergo a conformational change inducing a cascade of conformational changes that allow the of the transmembrane helical domain letting the water flow into the channel.

Stavroula Sofou explained how the pH can affect the phase separation of lipid bilayers and liposomes (or vesicles) and the roles played by electrostatic repulsion and hydrogen bonding in systems with domains of negatively charged lipids (PS) at neutral pH 7.2-7.4 (in normal cells) and acidic pH 6-6.5 (in cancer cells). She also showed the binding reactivity of uniformly functionalized nanoparticles, how receptor targeting nanoparticles deliver measurable amounts of the antitumoral drug doxorubicin and the pathway of the interaction of liposomes with cancer cells. Drug-loaded liposomes are sticky/active and selective only at the acidic pH of 6, they are not toxic to normal cells expressing the same receptor and their internalisation doesn’t depend on the type of ligand.

Karin Norling talked about the properties and cellular uptake characteristics of liposomes delivering vaccines, showing that liposomes with antigens inside and outside without and with PEG have different activity, the non PEGylated liposomes (150 nm) being more active that the PEGylated ones (200 nm). The in vivo study was performed in a mouse model and the antigen uptake was monitored on dendritic cells surface by means of TIRF microscopy and topographical micropattern to image liposomes. TIRF-based single particle tracking was also used to study interactions between cells and liposomes, showing that the liposome bound antigen is more present on the cell surface.

Anne Spang showed that the communication between organelles happens through transport vesicles via contact sites, allowing the exchange of lipids and ions in the plasma membrane (PM). Processing bodies (P-bodies) full of mRNA generated under different stress conditions might have other subcompartments. Csh3p, which is colocalized in the PM, and Pin2p are cargo proteins that can undergo prion formation. Pin2p is rapidly internalised upon osmotic stress and the mutation of the prion-like domain reduces its retention in the trans Golgi network (TGN), in which a phase separation occurs due to prion-like domain interactions.ascona3

Lukas Tamm talked about the role of cholesterol in the entry of Influenza and Ebola viruses into endosomes through pH mechanisms. He also showed the structure of influenza hemagglutinin monomer, it’s different domains, receptor binding and antigenic sites. He described how structural intermediates in HIV gp41 mediate membrane fusion, the post fusion crystal structure class 1 viral fusion protein having a 6 helix bundle. He also presented the NMR structure of Ebola virus fusion loop at pH 7 (inactive) and pH 5.5 (active) forming a fist structure, in which the double mutation to alanine residues removes completely the activity of the virus. HIV gp41 fusion domain contains alpha helix and beta sheet secondary structure elements, which mediate membrane fusion. The role of CHOL rich domains in HIV entry was also described via TIRF of SM:PS:CHOL and PC:PS 3:1 systems, showing that 50% of the particles bind to Lo/Ld boundaries, 35% to the Lo domains and the remaining to the Ld domains of supported lipid bilayers. Moreover, virosomes with Lo/Ld phases exhibit more fusion, the increase of the domain height mismatch from DLPC to DSPC increases also the fusion due to the reduction of the line tension and the energy gain due to reduction of line tension depends on the size of vesicles and lipid domains.

Sarah Veatch gave a very clear and interesting talk on the study of the phase behaviour of lipid bilayers and vesicle having Lo and Ld phases by means of 2H NMR spectroscopy. She showed what kind of composition fluctuations occur near critical points and how critical fluctuations can be studied with the 2D Ising model in GPMVs. She pointed out that raft domains are small dynamic regions of the lipid membrane and protein-protein interactions occur because proteins like to share the same local lipids. She also showed that the protein CTxB is clustered in certain domains of the PM depending on the type of probe: if a ordered probe is used the Lo domain is enriched in CTxB, while using a disordered probe the protein partitions more in the Ld phase. This analytical approach is going to be used to study the PM heterogeneity of the PM in intact cells. She also presented really interesting results on the anesthetic properties of small compounds, which depend on their structure and how they partition. Specific ligand-gated ion channels can be affected by anesthetics. Ethanol lowers the critical temperature Tc in GPMVs in a quantitative way: the more alcohol the lower Tc. Different alcohols have also the same effect on Tc of GMPVs. These properties of anesthetics are correlated with hydrophobicity and non anesthetics do not lower Tc. A shift in Tc can switch the channel from an inactive to an active state. Ligand-gated ion channels acquire the slow dynamics of membrane fluctuations. She also showed how tadpoles placed in ethanol and put to sleep become less sleepy with the addition of hexadecanol because it raises Tc.

Susan Daniel talked about membrane protein mobility and orientation are conserved in supported lipid bilayers (SLBs) created directly from cell PM blebs. These mammalian cells and GPMVs blebs can be used to study coronavirus, ebola virus and E. coli outer membrane vesicles. She also showed adsorbed blebs on the PM and their fusion and that lipid bilayers containing protein have a fluidity lower than protein free lipid bilayers. The protein mobility can be conserved with PEGylated lipid vesicles to form a cushion, in which protein can diffuse faster. An enzymatic orientation assay can be performed to see how the proteins are oriented. A mixed population of adsorbed blebs with a diameter size under 500 nm and PEG liposomes is used to generate SLBs.

Kelly Lee gave an excellent talk on the visualisation and sequencing of membrane remodelling leading to influenza virus fusion through cryo electron tomography (cryo-ET) tomography. He gave some background on endocytosis and pH triggered membrane fusion by influenza A virus, which contains 350-500 copies of the protein hemagglutinin (HA), and showed how the attachment via HA bridges leads to a significant deformation of the attached liposome. HA coordinates also the formation of dimples and can form extended interfaces with membranes tightly docked consistent with partially dehydrated proximal leaflets co-mingling. When HA is in the periphery of the liposome a trilayer structure, structurally similar to the one reported by the molecular dynamics study by Marrink cited in Marina’s post, is observed. In the presence of fusogenic lipidsNat pH 5.5 intermediates accumulate and if the pH is lowered to 5.25-5 the fusion progresses and more HAs are activated. He also pointed out that hemifusion is a very rare event.

Max Piffoux showed how the monitoring of extracellular vesicles dynamics at the nanoscale can be performed with liquid cell TEM of EVs coated with gold nanoparticles having a 100 nm diameter. The morphological analysis highlights the presence of different sized and shaped EVs in different media/buffers. Dynamics of EVs depends on nanoparticles coating not on their size and the aggregation of gold nanoparticles might be due to PS flip-flop.ascona5

Suzanne Eaton talked about the tissue size, shape, and collective cell behaviour in Drosophila (fruit flies), whose feeding occurs on rotting fruit allowing the uptake of yeast (fungal sterols) and plant (phitosterols) sterols. Yeast and plants have different lipid membrane compositions. She showed that animals fed with yeast lipid extracts have short lives, while plant fed animals live longer because they release less protein Dilp2 and have low systemic insulin signalling. Lipophorin ( Lpp) and Lipid transfer particles (LTP) are proteins that accumulate in the drosophila brain, but LTP is present on specific neurons. In particular two specific neurons recruit LTP only when larvae feed on yeast food, which enhances calcium release in glial cells. Since yeasts are a summer resource, the effect of the winter can be reproduced by performing experiments at 12 C during which drosophila eat more plant food. In another experiment she showed tha yeast-fed flies become uncoordinated at 12 C, while plant fed flies survive at lower temperatures. Moreover, larvae fed with plants survive outside in Dresden in the fall and adults can survive in the winter too. High levels of insulin are good for survival at high T, while flies fed on plants at 12 C have more fluid membranes, which prevent the formation of gel phases.

Matthew Wood gave a very interesting evening lecture on the development of treatments for muscular distrophy disease using RNA splicing. In his introductory slides he showed how the secretion of dystrophin, which is the protein product of the duchenne muscular distrophy locus, can be reduced by modulating splicing with small oligonucleotides to repair genetic defects. He explained that the absence of exons 49 and 50 leads to muscular distrophy and the removal of exon 51and the linking of exons 48 and 52 does not ead to the disease. He also pointed out that oligonucleotides tested in patients have had marginal benefits so far because RNA drugs are large macromolecules very difficult to be delivered and to be bioavailable in the tissue. Then, he showed the next generation of oligonucleotides and a peptide platform technology to synthesise cell penetrating peptides with a solid phase synthesis. In particular he talked about the use of Pip6a, a 22 amino acids arginine rich peptide, to treat spinal muscular atrophy, which is a childhood motoneuron disease due to the missing of exon 7 and whose current invasive treatment involves the drug to be delivered directly to the brain with possible infection risks. The delivery of Pip6a-PMO corrects sMN expression and phenotype in SMA mice, increases the lifetime with a single dose and with a double dose mice live a normal life. The shortening to 12-14 amino acid long peptides has generated other 17 peptides that can be used as drugs. He explained that there are different brain barriers, no barrier, a blood CSF barrier and a blood brain barrier, which form a complex multicellular barrier. Then, he showed how EVs can be used as possible delivery methods, a hypothesis supported by the ability of exosomes to transport mRNA between cells. Their EV engineering strategy involved the presence of the integral exososomal protein Lamp2b and dendritic cells to have rabies virus glycopeptide (RVG) and RNA encapsulation into EVs, in particular single stranded RNA (siRNA) and microRNA. Targeting peptides enhance brain penetration leading to the siRNA drug delivery to mouse cortical neurons. He also talked about the treatment of Parkinson’s disease Lewy bodies and alpha synuclein accumulation with RVG-exosomes. Finally, he described the EV delivery mechanism, intercellular vehicles for RNA delivery and the retro engineering into lipid nanoparticles (LNPs), which are extremely inefficient to deliver siRNA. He showed that in the EV cell uptake most LNPs aggregate on the cell surface with an uptake time of 5 minutes. Exosomes are taken up as single vesicles in all recorded events andore than 80% of EVs are internalised at equilibrium conditions. EV cell uptake is highly polarised and associated with filopodia active regions, which are cortical actin bundles rich domains. EV uptake is facilitated by filopodia showing surfing along and filopodia mediated exosome grabbing; more than 98% uptake events happen at the base of the filopodia. These 80-100 nm Evs make stop-and-go movements along the ER and after kiss and run, dynamic and sampling events are finally sorted to the lysosomes.

On a rainy Thursday night, after dinner, we also had the chance to dance and I ended up dancing (or at least that’s what I was trying to do) very shyly with a playlist that included songs by Michael Jackson, David Bowie and other artists, and we kept drinking and socialising..

Sarah Keller gave the most energetic, enthusiastic and lively talk of the meeting to wake up an audience dealing with sleep deprivation, hangover and other more or less personal problems. At the beginning she showed clearly how at high temperature GUVs display an uniform disordered domain and the appearance of vesicle liquid ordered domains lowering the temperature. The mixing of high and low melting temperatures lipids leads to domains in both membrane leaflets. She also explained how to read a ternary phase diagram with lipids in ternary and binary mixtures phase diagrams and to quantify the strength of the transbilayer coupling between domains in each leaflet. The energy of the transbilayer coupling ranges from 10-8 kT to 0.15-0.5 kT/nm2, the latter values being reported from MD simulations. Lipid bilayers on a solid subtstrate move like tanks, in order to avoid this process GUVs are burst on a solid substrate, which will make a lipid bilayer after a flow of water. Small domains are stuck big domains move hydrodynamic with the shear stress being inversely proportional to the size of domains. Four forces are involved in the process: shear stress, dragging, friction and registration forces. The measured interleaflet coupling parameter of 0.016 makes possible the registration of bigger than 10 nm domains. Finally, she made some pretty reasonable wild speculations on the origins of life by reporting a study of the fatty acids interacting with different RNA bases. Adenine and guanine stick the best to vesicles because these bases bind better to membranes.

Petra Dittrich talked about microfluidic devices to study lipid membrane properties by creating of nL and pL volumes, allowing the fast exchange of fluids and the immobilisation/trapping of cells and vesicles through the fabrication of different sized channels chambers with multilayer soft lithography. A single vesicle can be trapped without its rupture and fluids can be exchanged very quickly (100 ms). She showed the permeation partitioning across membranes containing membrane pore alpha hemolysin, studies of cell penetrating peptides (CPPs) and their permeation across a POPC membrane, and a study of the fusion of membranes by trapping two vesicles and using a modified fusogenic amphipathic peptide derived from influenza virus. She also presented a study of the fusion of membranes by electrical fields, how forces acting on membranes generate deformation making flat vesicles, how small domains fuse to larger domains and Ld domain budding. Finally, she gave an overview of the different applications of microfluidic devices, including cell/vesicle analysis, vesicles in droplets a model to study intracellular compartments, liposome screening platforms used for permeation studies, ligand-receptor binding and hormone-receptor binding.

Wye Khay Fong talked about different lipid phases, in particular bicontinuos cubic and reversed hexagonal phases, can be used as substrates biosensor and have applications to drug delivery. She showed how the cubosome nanostructure controls the drug release in vitro and in vivo with a temperature switch, which can be monitored with kinetics synchrotron SAXS measurements to study external and internal triggered changes in the nanostructure. She also described the lipid digestion on the nanoscale, in particular the milk digestion involving fats that can form different mesophases, and introduced a pharmaceutical milk shake in which invertase catalyses the digestion of the sugar glucose. She also mentioned that hydrolysis leads to phase transitions and drug release.ascona1

Roy Ziblat talked about the building-up of a membrane library for protein-lipid match with a very nice movie and a funny soundtrack. He pointed out that membrane receptors interacting with viruses are not only proteins but are also lipid based ones. He showed affinity profiles of Dengue, Ebola and vesicular stomatite virus for different lipids. He highlighted that liposomes treatment with infected cells is biocompatible, FDA approved and only few micrograms are effective.

Chen-Yu Zhang talked about exosomes sufficiently delivering secreted small RNA to recipient tissues. Initially, he explained some biological functions of microRNA, the use of circulating microRNA as a novel class of biomarker for diseases and the employment of Microvesicles MVs, which are identical to exosomes secreted in the cells, as carriers for microRNA. He also showed how human blood cells and cultured THP-1cells selectively package miRNAs in exosomes and MVs and the biological function of exogenous miRNA-150 in circulation. Secreted microRNA acts as a signalling molecule in mediating intercellular communication and in MVs enhances angiogenesis in cancer cells (enriched in MVs) and hepatic insulin resistance. Plant microRNAs absorbed by mammalians can be packaged into MVs leading to cross kingdom regulation. He pointed out that honeysuckle microRNA, which is not destructed by the boiling process, shuts down the viral activity because of its specific sequence and it can be used to treat viral infection in the fetus.

Gisou van dear Groot gave a very interesting talk on the function and dynamics of protein palmitoylation. Initially, she explained how S-palmitoylation adds hydrophobicity to a protein containing cysteine residues, phosphorylation adds a charge changing the conformation of a protein, affecting its folding and function, and ubiquitination puts a tag on a protein. Then, she talked about the DHHC family of palmitoyltransferases, which are 23 in human genome with 18 out of 23 in the ER and are involved in brain and cancer diseases. She also promoted the website in which different properties of these enzymes are described. She highlighted that when a protein acquire a lipid within the transmembrane region a conformational change of the transmembrane domain occurs, especially those proteins associated with lipid rafts are affected by palmitoylation and specific lipids are required to modulate the hydrophobic mismatch. The formation of protein complexes and interplay with other post translational modifications can be investigated with tritium labelled or clickable palmitate only the non palmitoylated proteins are labelled.

She also stressed that palmitoylation contributes to the integration of diverse ER functions and it is also involved in intraorganelles contacts. Then, she described calnexin palmitoylation (protein-SH –> proteinS-palmitoyl),cal-SH –> calS-palmitoyl, the presence of two palmitoylation sites and the long time (45 hours) of the dual palmitoylation. The frequency of palmitoylation has a median time very slow of 6 hours due to phosphorylation and the palmitoylation of the second site is cooperative. DHHC6 spans the membrane three times with three cysteines that can be palmitoylated by DHHC16 and undergoes rapid depalmitoylation. Palmitoylation destabilises the enzyme, affects localisation and oligomerization and it is required for activity. The stability of DHHC6 is site dependent and most of the protein is not palmitoylated, only one of the site is active, the presence of three sites protects it from degradation and preserves its activity. PEGylation can be used to check palmitoylation and site occupancy. Finally, she presented the results of a molecular dynamics simulation showing that the palmitoylated cysteines of calnexin are inside the lipid bilayer and they need to be exposed in order to be palmitoylated by the enzyme.

At the end of the meeting Kalina Hristova announced the recipients of two student and two postdoctoral poster awards, which were assigned to Erik Henrich, Georg Krainer, Kirstin Hobiger and Radhakrishnan Panatala.

If you made it to the end of this very long post you are about to read a flashback with my personal comments on the meeting, which actually started for me at Locarno railway station, where I was supposed to meet Marina Ramirez Alvarado. I eventually ended up meeting Kellen Brunaldi, Carmen Domene, Kirstin Hobiger and Marek Cebecauer before having the pleasure of meeting Marina. Initially, she introduced me to Rosalba Kampman, the executive director of the Biophysical Society. Then, I also started seeing the familiar faces of Robert Vácha, Luca Monticelli and Giulia Rossi. I met Luca for the first time at the Biophysical Society Annual Meeting held in Salt Lake City in 2006 and less than two years later we were working in Ilpo Vattulainen’s group, in which Giulia was also working at that time. I had the pleasure to see Giulia giving a very interesting talk at an International Workshop on Biomembranes held in Espoo, Helsinki, a couple of years ago. It was kind of interesting and funny at the same time that we were all clustered in the first poster session. I enjoyed talking to them about different, not always specifically scientific, things, it was really nice seeing both of them again. Then, during my poster presentation and the other days of the meeting at breakfast, lunch and dinner I had the chance to talk to many nice and extremely talented people. Here is a brief list of the interactions we had in a more or less chronological order.ascona4

Marie-Eve Aubin-Tam was the first person interested in the research shown in my poster (maybe she was judging me :)) and in seeing the movies of coarse grained MD simulations reported in it. We also had very interesting more or less scientific conversations over lunch and dinner. Marina, Steven Boxer (the official judge), Carmen, Giulia and Alexander Karabadzhak asked excellent questions about my poster too. The same day I had the pleasure to have lunch with Donald Engelman, who told me he had an Italian postdoc from Sardinia working in his lab in the past, his wife, Daniel Müller, Stefania Mari, Botond Roska, who is also in love with Sardinia, Giulia and Luca. Stefania, Johannes Thoma and Estefania Mulvihill were also interested in molecular dynamics simulations and it was nice talking to them about other things too. I also had very nice conversations with Xiaojun Shi, Kirstin Hobiger, Nikhil Gandasi, Hudson Pace, Jenny Isaksson, Vanessa Carvalho, Radhakrishnan Panatala, Marisa Sarria, Wye Khay Fong, Sonia Troeira Henriques and Binjong Liang. I really hope to see all of them again in the near future.

It was a very intensive, extremely well organised and loaded with so many excellent talks kind of meeting, and I hope there will the possibility to attend similar meetings in the future, not near because my brain needs some time to recover a bit now… 🙂


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