LIFE SCIENCE

Seminars

Every year, the PhD school organizes or contribute to organize seminars on topics related to the PhD curricula.
The seminars are held face-to-face except for some dates which are highlighted.
In this page, we also alert students for seminars on related topics.

The seminars will be held in the room indicated in the single announce.

2024

Role of Extracellular microRNAs in the biology of the Tumor Microenvironment
16/12/2024 14.00 Room B (Building CU010) Prof. Muller Fabbri Children’s National Hospital and George Washington University (USA)
MicroRNAs (miRs) are small non-coding RNAs which regulate gene expression at the post-transcriptional level. Their dys-regulation has been observed in all types of cancer and, more broadly, in several human diseases. Extracellular Vesicles (EVs) are membrane-coated particles released by all cells, including cancer cells and are involved in inter-cellular communication. This seminar will highlight the role of miRs as cargo of EVs in shaping the tumor microenvironment, contributing to the growth and dissemination of cancer, as well as their potential therapeutic role, as powerful anticancer molecules released by Natural Killer (NK) cells.

The role of (lnc)RNA-protein interactions at the interface of chromatin and cancer biology
02/12/2024 Room 101 (RM 112-E01P01L005) – I° Floor, Building D - Viale Regina Elena 295a Dr. Giulia Guiducci Queen Mary University of London (UK)
Over the past two decades RNA molecules have emerged as active regulators across a variety of biological processes. Among them, the class of long non-coding RNAs (lncRNAs) is under the spotlight for their contribution to several cancer hallmarks. I will introduce a newly identified chromatin-enriched lncRNA, which we named Chromosome Segregation Regulating noncoding RNA (CHERR). CHERR is a cancerrelevant lncRNA, and its depletion in lung cancer cells leads to the non-genetic loss of a known tumor suppressor, ARID2, a key subunit of the SWI/SNF chromatin remodelling complex. Our findings indicate that CHERR binds to the SWI/SNF subunits, potentially promoting the stability of the complex. In line with this, loss of CHERR disrupts ARID2 association with chromatin. Additionally, I’ll present our ongoing efforts to dissect RNA-protein interactions across cell cycle stages and I’ll introduce TREX, an RNA-centric method to map endogenous RNAprotein interactions with region resolution.

Current and future consequences of the silent antimicrobial resistance pandemic and measures being adopted to combat AMR
24/05/2024 Stephen Hawser CEO IHMA Europe Sàrl Monthey Switzerland
Antimicrobial resistance (AMR) continues to evolve and spread beyond all boundaries. As a result, many infections have become more challenging or even impossible to treat, leading to an increase in morbidity and mortality. Despite the failure of conventional, traditional antimicrobial therapy, in the past two decades, no novel class of antibiotics has been introduced. While the COVID-19 pandemic was frequently mentioned daily worldwide, the AMR crisis is rarely talked about, hence it being labelled as the “Silent Pandemic”. Yet, there are more infections and deaths per year attributed to AMR compared with COVID-19 and indeed many COVID-19 patients died from antimicrobial resistant infections. Currently there are new therapeutic options, especially combination therapies though they are not completely novel. These therapies are helping to slow the silent pandemic but inevitably resistance is emerging. If no new therapies or technologies are developed for the future, AMR will be unstoppable and mortality rates will significantly increase. Fortunately, novel molecules amongst others are in early to mid-stage development though it remains to be seen if these can be developed quickly enough in order to be of clinical utility.

The pivotal role of functional biochemistry in Precision medicine.
17/05/2024 Prof. Maurizio Simmaco Dipartimento di Neuroscienze, Salute mentale ed Organi di senso, Università Sapienza
The number of people aged 65 years or more in the EU is expected to grow from around 84 million in 2008 to around 141 million by 2050 and they will account for 28.7% of the EU-28’s population by 2080. Italy is in second place in Europe by old age index, with strong consequences on health care system, due to the high number of chronically ill people. There is an urgent need for implementation in standard clinical practice of novel methods to ensure efficacy and safety of polypharmacy in the elderly. Precision Medicine (PM) represent a patient-centered model of health care, based on preemptive and monitoring strategies developed in the last decade thanks to the availability of novel cost-effective technologies to achieve deep molecular characterization of both patients and diseases. The striking impact of the proposed strategy derives by the system-level character of the project, intended to integrate a novel model of healthcare in an Italian National Health System (INHS) structure, that is the academic hospital Sant’Andrea of Rome. Such unique setting allow to integrate sophisticated preventive and monitoring strategies in the usual clinical workflow. The data constitute a proof-of-principle concerning the effectiveness of integration of PM in current clinical practice to improve adherence to therapies and clinical outcomes and is expected to provide evidence that translation of PM into the INHS is indeed actionable.

Measuring heat to understanding catalysis: isothermal titration
10/05/2024 Prof. Barbara Zambelli Dipartimento di Farmacia e Biotecnologie Università di Bologna
Isothermal titration calorimetry (ITC) is a widely used technique to characterize the thermodynamics of biochemical reactions, as it uses the heat released or absorbed upon virtually all reactions as an internal probe. Indeed, the measured heat is related to the amount of reacting molecules and to the rate of the reaction of interest. The ability of this technique to derive the thermodynamic parameters of the biochemical processes makes it an invaluable and powerful technique to study biomolecular binding equilibria. In addition, ITC has been demonstrated to be able of directly measuring kinetics and thermodynamic parameters (kcat, KM, ΔH) of enzymatic reactions. As heat changes spontaneously occur during enzymatic catalysis, ITC does not require any modification or labeling of the system under analysis and can be performed in solution. These properties render ITC a unique tool to study enzyme kinetics in applications such as drug discovery. These features will be described and discussed together with some applications to specific cases of enzyme catalysis and inhibition. A general guideline to choose the right procedure according to the system under analysis is given, together with some experimental tips on how to adjust the conditions for increased data quality. The method to analyze the obtained raw ITC curves and to derive the kinetic parameters and inhibition constants is described.

Unifying Concepts in Nitrogenase Catalysis
03/05/2024 Prof. Oliver Einsle University of Freiburg (Germany)
In the talk, the mechanism of nitrogenase will be discussed. Nitrogenases are responsible for the reduction of nitrogen (N2) to ammonia (NH3). Nitrogenases are the only family of enzymes known to catalyze this reaction, a step in the process of nitrogen fixation, in which the triple bond of the inert N2 molecule is broken. In the seminar, features of and problems of biological nitrogen fixation will also be introduced.

Chaperones Countering Protein Self-Assembly in Neurodegeneration
19/04/2024 Prof. Cláudio M. Gomes Faculdade de Ciências da Universidade de Lisboa (Portugal)
In this seminar I will provide an overview of how defective proteostasis underlies various neurodegenerative diseases characterized by protein deposition. I will unveil our recent discoveries demonstrating the neuroprotective functions of brain proteins from the S100 family. These proteins act as intra and extracellular chaperones, effectively halting the self-assembly and neurotoxicity of tau and amyloid β aggregates. I will delve into the underlying mechanisms of action, particularly focusing on S100B, and will conclude by proposing the hypothesis that S100 proteins collectively form a novel type of protective chaperone network in the nervous system. 1. Hagmeyer et al (2019) Frontiers in Neuroscience https://10.3389/fnins.2019.00640 2. Cristóvão et al (2018) Science Advances https://doi.org/10.1126/sciadv.aaq1702 3. Moreira et al (2021) Nature Communications https://10.1038/s41467-021-26584-2 4. Figueira et al (2022) Journal Molecular Biology https://doi.org/10.1016/j.jmb.2022.167791 5. Moreira and Gomes (2023) Journal Neurochemistry https://doi.org/10.1111/jnc.15756 6. Figueira et al (2023) S100B Front. Neuroscience https://doi.org/10.3389/fnins.2023.1162741

Rational design approaches to interfere with protein misfolding and aggregation through peptidomimetic foldamers
12/04/2024 Prof. Nicolo Tonali: FLUOPEPIT, BioCIS, CNRS, Université Paris Saclay,Châtenay-Malabry (France)
The aggregation of proteins into amyloid fibers is linked to more than 40 still incurable cellular and neurodegenerative diseases such as Alzheimer's disease (AD), Parkinson's disease and type 2 diabetes. The process of amyloid formation is a main feature in cell degeneration and disease pathogenesis. Many anti-amyloid molecules have been reported over the past 25 years and most of them belong to small molecules or antibodies. However, so far only one of the anti-amyloid drug candidates, Tafamidis, which inhibits transthyretin amyloidogenesis (TTR), and an antibody, Aducanumab, which targets aggregates of beta-amyloid peptide 1- 42 (Aβ1-42), reached the clinic. Peptides are an attractive alternative to small molecules and antibodies as anti-amyloid drugs, thanks to their improved efficacy, selectivity or specificity, and potency. However, very few of them have reached the (pre) clinical stages, and so far, none of them has reached the clinic. Peptidomimetic foldamers, bio-inspired by secondary structures of amyloid proteins, provide a promising alternative to peptides because they keep the specific side chains of a peptide sequence while having new and improved biological and pharmacokinetic properties and the possibility of adopting secondary structures frequently involved in protein- protein interactions. Rational design approaches to interfere with protein misfolding and aggregation through peptidomimetic foldamers (β-strand, PPII and α-helix) are here presented. The design has been based on both self-aggregation and cross- interaction processes. These foldamers have shown a good ability in vitro to interfere with the aggregation process of Aβ1-42 and tau, especially those with a stable secondary conformation. Circular dichroism analyses on the conformational change of the amyloid protein in the presence of these foldamers revealed their ability to stabilize intermediate conformations, which could be the reason for the reduced aggregation propensity and thus toxicity. These approaches represent a practical application of peptidomimetic foldamers in therapeutics, particularly in pathologies involving abnormal protein-protein interactions. 1. N. Tonali, L. Hericks, D.C. Schröder, O. Kracker, R. Krzemieniecki, J. Kaffy, V. Le Joncour, P. Laakkonen, A. Marion, S. Ongeri, V. I. Dodero, N. Sewald; ChemPlusChem 2021, 86(6), 840. 2. L. Ciccone, C. Shi, D. Di Lorenzo, A.C. Van Baelen, N. Tonali; Molecules 2020, 25(10), 2439. 3. Shi, C.; Kaffy, J.; Ha-Duong, T.; Gallard, J.-F.; Pruvost, A.; Mabondzo, A.; Ciccone, L.; Ongeri, S.; Tonali, N.; J. Med. Chem. 2023, 66 (17), 12005–12017.

Thermodynamic and kinetic approaches for drug discovery to target protein misfolding and aggregation
08/03/2024 Prof. Michele Vendruscolo: Yusuf Hamied Department of Chemistry University of Cambridge, UK
Protein misfolding diseases, including Alzheimer’s and Parkinson’s diseases, are characterised by the aberrant aggregation of proteins. These conditions are still largely untreatable, despite having a major impact on our healthcare systems and societies. To address this problem, I will describe drug discovery strategies to target protein misfolding and aggregation. More specifically, I will compare thermodynamic approaches based on the stabilization of the native states of proteins with kinetic approaches based on the slowing down of the aggregation process. This comparison will be carried out in terms of the current knowledge on the process of protein misfolding and aggregation, the mechanisms of disease and the therapeutic targets.

The role of cardiolipin in mitochondrial disorders
01/03/2024 Dr. Micol Falabella: Department of Neuromuscular Diseases University College London (UK)
Primary mitochondrial diseases (PMDs) are among the most common inherited neurological disorders. They are caused by pathogenic variants in mitochondrial or nuclear DNA that disrupt mitochondrial structure and/or function, leading to impaired oxidative phosphorylation. One emerging subcategory of PMDs involves defective phospholipid (PL) metabolism. Cardiolipin (CL), the signature PL of mitochondria, resides primarily in the inner mitochondrial membrane, where it is biosynthesised and remodelled via multiple enzymes and is fundamental to several aspects of mitochondrial biology. Disruption to genes involved in CL biosynthesis and remodelling has recently been associated with PMD. However, the pathophysiological mechanisms that underpin human CL-related disorders are not fully characterised. Here, we report six individuals, from three independent families, harbouring biallelic variants in PTPMT1 with a complex neurological and neurodevelopmental syndrome. Using patient-derived fibroblasts and skeletal muscle tissue, together with cellular rescue experiments, we characterise the molecular defects associated with mutant PTPMT1 and confirm the downstream pathogenic effects that loss of PTPMT1 function has on mitochondrial structure and function. To further characterise the functional role of PTPMT1 in CL homeostasis, we established a zebrafish ptpmt1 knockout model associated with abnormalities in body size, developmental alterations, decreased total CL levels, and oxidative phosphorylation deficiency. Together, these data indicate that loss of PTPMT1 function is associated with a new autosomal recessive PMD caused by impaired CL metabolism, emphasising the contribution of aberrant CL metabolism towards human disease and its importance in neurodevelopment.

2023

Riboregulation from bacteria to eukaryotes: mechanisms and challenges.
21/06/2023
The aim of this one-day workshop is to gather attention on the novel role of RNA as regulator of protein activity and its importance in the control of cellular metabolism. Understanding the molecular details of RNA-protein interactions and the effect on cellular physiology is a key step to exploit the impact of these discoveries to fundamental science and to translational applications in the field of RNA therapeutics

The axonal transport machinery and its dysfunctions in neurodegenerative diseases
19/05/2023 12:00 ONLINE Prof. Giampietro Schiavo Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology Dementia Research Institute, University College London, London
The molecular mechanisms causing neuronal death in many neurodegenerative diseases, such as amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD) and Charcot Marie Tooth (CMT) disease, are poorly understood. The key consequence of our incomplete understanding of disease pathogenesis is that there is a complete dearth of effective symptomatic treatments for these widespread global disorders, prompting the necessity for a step-change in treatment strategies to fight these pathologies. In this view, we are investigating ALS and CMT as disease paradigms to identify new, common targets for pharmacological intervention in these devastating pathologies. Recently, we uncovered alterations in axonal transport of several cytoplasmic organelles, such as mitochondria and signaling endosomes, at pre-symptomatic stages of ALS and CMT pathogenesis, suggesting that these impairments may play a causative role in disease onset and progression. Crucially, we have restored axonal transport to physiological levels at early symptomatic stages of disease, thus demonstrating that these pathological changes are fully reversible. In light of these results, our main goal is identifying novel signaling nodes that modulate axonal transport in healthy and diseased neurons. This will allow us to test the hypothesis that counteracting axonal transport deficits observed in neurodegenerative diseases, represents a novel, effective therapeutic strategy towards treating these pathologies.

The chaperonin GroEL nano-machine: allostery and function
12/05/2023 ore 13, Aula A CU010 Prof. Amnon Horowitz Department of Chemical and Structural Biology Weizmann Institute of Science Rehovot, Israel
Chaperonins are nano-machines that are built of two back-to-back stacked heptameric rings. They assist protein folding by undergoing large conformational changes that are controlled by ATP binding and hydrolysis. In the E. coli cell, only about 60 different proteins require GroEL for efficient folding. In the first part of the talk, I will describe work that was aimed at determining the properties that distinguish GroEL clients from non-clients. In the second part of the talk, I will describe the impact of encapsulation on the stability of protein substrates. In the third part of the talk, I will describe new approaches for elucidating allosteric mechanisms. Using these approaches, it has been possible to show that GroEL undergoes concerted intra-ring conformational changes. By contrast, the eukaryotic homologue CCT/TRiC undergoes sequential intra-ring conformational changes. The impact of these different allosteric mechanisms on the folding functions of GroEL and CCT/TRiC will be discussed.

Regulation of enzymatic activity mediated by liquid-liquid phase separation
28/04/2023 13:00 (Aula A CU010) Dott. Mirco DindoDepartment of Medicine and Surgery, Section of Physiology and Biochemistry University of Perugia, 06132 Perugia, Italy
A feature of biological evolution at the cellular level is the development of diverse organelle structures in the cytoplasmic environment. Cells contain membrane-bound organelles and membrane-less organelles. Liquid–liquid phase separation (LLPS) is gaining acceptance as a powerful mechanism to explain the formation of membrane-less organelles. Emerging evidence indicates that LLPS is important in many biological processes, such as transcription, signaling and metabolism playing a vital role in human health and disease. These discoveries describe the LLPS as a new fundamental physicochemical mechanism for organizing the biochemistry of the cells. LLPS is recognized as a mechanism for regulation of enzymatic activity. Biochemical mechanisms include concentrating reactants to enhance reaction rates or sequester enzymes and reactants from each other to reduce the reaction rate. On the other hand, LLPS might also regulate the diffusion of small molecules or important parameters for enzymatic activity (such as modulators, macromolecular crowding and changing the media physicochemical features) increasing or decreasing the reaction rate of the enzymes. Here, I will show how LLPS contributed to generate a new era for enzymic activity regulation as well as other subtle regulation mechanisms still unexplored and recent advances in the LLPS field.

Protecting our genome: mechanisms to maintain DNA repeats stability in human cells
21/04/2023 Aula A-CU10 ore 13.00 Prof. Simona Giunta Department of Biology and Biotechnology Charles Darwin, Sapienza University of Rome, Italy
Centromere alpha-satellite DNA repeats are amongst the most recalcitrant regions of our genome. If we envisage genome browsers as Google Maps, zooming into increasing amount of details, centromeres have historically been black boxes that yield no linear sequence information. Yet, they are especially fascinating for three reasons: (1) centromeres are cytologically distinctive parts of human chromosomes that are essential for their segregation; (2) unlike the rest of our genome, centromere do not undergo meiotic recombination, hence they are inherited as intact blocks of maternal and paternal DNA; (3) in spite of points 1 and 2, centromere alpha-satellite DNA repeats are intrinsically unstable, a behavior of centromeres that I first described in 2018 as akin to human fragile sites in our genome. Indeed, centromere alpha-satellite repeats witness the convergence of several scenarios that can trigger DNA instability. Unlike the flanking region of constitutive heterochromatin at the pericentromere, centromeres are transcribed into non-coding RNAs throughout the cell cycle which offers opportunities for the collision between the transcription and replication machineries. Ours and others work have shown that centromere DNA is late replicating, prone to undergo rearrangements which becomes exacerbated in cancer and during cellular senescence. We demonstrated that epigenetic disruption of centromeric chromatin triggers pathological generation of mutagenic R-loops due to transcription-replication collision. The ensuing DNA damage repair within highly homogeneous centromere repeats results in a specific form of aneuploidy distinct from the numerical aneuploidy triggered by centromere inactivation, that we defined as structural aneuploidy including whole arms events and translocations frequently observed in most solid tumors. Our recent work shows that centromeres suffer higher level of endogenous DNA damage in unperturbed conditions compared to the rest of the genome, and also display different DNA damage repair kinetics, both contributing to their intrinsic instability. My laboratory uses genomics, including the latest human genome assemblies now including linear sequences of alpha-satellites, cytogenetics and high-end imaging to study centromeres as vulnerable regions of our genome. Altogether, work from my lab aims to understand centromere mutagenesis at the incipit of degenerative human diseases.

Synthetic Biology: what, why and how
14/04/2023 13:00 Aula A CU010 Dott.ssa Velia Siciliano, Synthetic and Systems Biology for Biomedicine Istituto Italiano di Tecnologia-IIT
Synthetic Biology is a Bioengineering discipline that aims at reprogramming cell fate by designing genetic circuits that perform sophisticated information processing. Synthetic biology has the potential to revolutionize the treatment of hard-to-tackle diseases by reprogramming cells with synthetic devices. To obtain robust and specific activity, synthetic circuits must sense and respond to the intracellular or extracellular environment recognizing the unhealthy condition. Over the talk I will focus on the design of a platform that can be easily readapted to sense intracellular proteins of interest, and their application for engineering potential cell-based therapies. I will also show our work on RNA-encoded circuits that use RNA-binding proteins, siRNAs and proteases to engineer sensors, cascade and switches. Finally, I will present our recent research to address one of the standing bottlenecks of mammalian synthetic biology, namely the burden given by competition for intracellular resources that synthetic circuits impose to the cells.

Educating tutor-associate macrophages under metabolic stress
30/03/2023 ore 12:00 ONLINE Dr. Ping-Chih Ho Department of Fundamental Oncology & Ludwig Institute for Cancer Research University of Lausanne, Switzerland
Metabolic stress is believed to impede proper activation and differentiation of immune cells in the tumour microenvironment. Moreover, re-educating tumour-associated macrophages is believed to be a critical process to reprogram immune state in tumors and maximise host anti-tumor immunity. In my talk, I will cover our understandings on metabolic regulations in macrophage polarisation and education. Then, I will further discuss how we can achieve efficient re-education in tumour-associated macrophages by requiring their metabolic programs.

Insulin Signaling in Alzheimer’s Disease Brain and Models Thereof
Thursday 23/03/2023 Aula A CU010 13:00 Prof. Eugenio Barone Department of Biochemical Sciences -Sapienza University
Brain insulin signaling acts as a key regulator for gene expression and cellular metabolism, both of which sustain neuronal activity and synaptic plasticity mechanisms. Alterations in this pathway, known as brain insulin resistance, are associated with a higher risk of developing age-related cognitive decline and neurodegeneration. Studies from our group have uncovered the role of the enzyme biliverdin reductase A (BVR-A) in this process. BVR-A, beyond its activity in the degradation pathway of heme, is a novel regulator of insulin signaling. BVR-A is a direct target of the insulin receptor, similar to insulin receptor substrate-1 (IRS-1). The insulin receptor phosphorylates BVR-A on specific Tyr residues, activating it to function as a Ser/Thr/Tyr kinase. In addition, downstream from IRS-1, BVR-A works as a scaffold protein, promoting the translocation of GLUT4-containing vesicles to the plasma membrane (to increase glucose uptake in response to insulin), the AKT-mediated inhibition of GSK3β (which promotes cell survival), and the AMPK-mediated inhibition of MTOR (which is involved in autophagy). Our group's groundbreaking findings have revealed that oxidative stress-induced impairment of BVR-A is a key event driving brain insulin resistance development in Alzheimer's disease (AD). Conversely, rescuing BVR-A activity reduces oxidative stress levels and improves brain insulin signaling, both of which contribute to improved cognitive function in animal models of neurodegeneration. Overall, our data suggest that BVR-A represents a molecular link between oxidative stress and insulin signaling, and studies to further investigate its role in the development of neurodegenerative disorders are ongoing in our lab.

Deciphering self-renewal traits in epidermal stem cells
Friday 17/03/2023 12:00 Aula C CU010 Dott. Elena Enzo Centre for Regenerative Medicine ‘‘Stefano Ferrari’’, University of Modena and Reggio Emilia
Epithelial stem cells are endowed by impressive regeneration capacities, that sustain epithelia regeneration during homeostasis and wound healing in vivo. In vitro, they can proliferate for dozens of population doublings. Since the 1980s, cultured human keratinocytes have been extensively used to regenerate functional cornea and epidermis. Successful epithelia regenerations require an adequate number of epithelial stem cells, endowed by long lasting proliferative potential and self-renewing capacity. They progressively give rise to transient amplifying progenitors that proceed the onset of terminal differentiation. Despite their clinical success, we still are seeking a clear understanding of the molecular mechanisms that regulate self-renewal and regenerative potential self-renewal and regenerative potential in epithelial stem cells. Taking advantage of single-cell transcriptomic analysis of primary human epithelial cultures we identify the cluster of epithelial stem cells characterized by genes regulating DNA repair, chromosome segregation, spindle organization and telomerase activity. We identify FOXM1 as a YAP-dependent key regulator of epidermal stem cells. Moreover, we are focusing on how stem cell and transient amplifying progenitors respond to genotoxic insults, looking at specifics molecular mechanisms that allow to prevent accumulation of genetic lesions in epidermal stem cells, and we found that FOXM1 is specifically activated upon genotoxic insults to foster DNA repair process. A better understanding of the complex network acting to repair the genotoxic insults will be instrumental to improve error-free based gene editing techniques. Moreover, as many genotoxic insults assault skin epithelia eventually causing oncogenic growth, this knowledge will also add insights on skin cancer progression, to develop early diagnostic tools or personalized therapies.

Application of deep learning to the protein structure prediction: the tale of a “gigantic leap”
Thursday 09/03/2023 ore 13:00 Aula A CU010 Prof. Anna Marabotti, Department of Chemistry and Biology "A. Zambelli" University of Salerno
Abstract: Knowing the three-dimensional structure of a protein is of outmost importance for understanding its function and evolutionary relationships with other members of the same protein family. However, the determination of 3D protein structures by experimental methods is not straightforward. For this reason, several computational methods have been developed in the past decades to predict the structure of proteins from their sequences. This field has experienced tremendous development in the last 4 years, when deep learning algorithms have made it possible to obtain structural predictions at a level of accuracy approaching that of experimental studies. In this seminar, the evolution of protein structure prediction methods will be shown, with special reference to the development of artificial intelligence methods, and the implications of these new approaches for knowledge not only at the level of protein structure and function, but also in terms of knowledge of the interactions with other proteins and/or macromolecules inside and outside of cells, and, prospectively, for applying this knowledge in various fields, including rational drug design.

Engineering Proteins To Boost LTP And Memory
Friday 03/03/2023 ore 12.00 AULA C CU010 - Prof. Cristian Ripoli Department of Neuroscience of Università Cattolica del Sacro Cuore– Fondazione Policlinico Universitario A. Gemelli IRCCS, Rome, Italy
Synaptic plasticity (LTP) is considered the cellular correlate of learning and memory and its impairment has been linked to several neurological disorders, including Alzheimer's disease. Although the canonical pathways responsible for LTP involve dozens of proteins compartmentalized within dendritic spines, the LTP master controllers include key identified proteins such as CaMKII, LIMK1, PKC, BDNF and CREB. Using synthetic biology, we developed genetically encoded engineered proteins (GEEPs) to control their activity in living neurons boosting LTP. We employed artificial regulatory domains, whose conformations can be selectively and robustly controlled by the well-tolerated, clinically approved, blood-brain barrier permeant and procognitive drug rapamycin or its non-immunosuppressive analogs. Then, we combined biochemistry, electrophysiology, and two-photon imaging, demonstrating the viability of the proposed approach in controlling: •spine enlargement (GEE-LIMK1); •spine plasticity by either controlling BDNF maturation or engineering its receptor (GEE-BDNF/TrkB); •functional LTP (GEE-PKC) and •the clearance of misfolded proteins by engineering ADAM10. Using AAV infections and mouse behavioral analyses, we validated the potential of GEEPs to improve memory. Since several learning and memory disorders still lack efficacious treatments, our strategy will boost further research into next-generation bioengineering solutions as novel therapeutic strategies for in several brain disorders associated with cognitive dysfunctions.

2022

Protective / Preventive Role Of Natural Compounds In Human Health: Biochemical Approaches To Target Ageing And Neurodegeneration
01/06/2022 Silvana Hrelia and Cristina Angeloni University of Bologna
The modern “Nutrition 3.0” model will be introduced focusing on the growing interest in nutraceutical bioactive compounds. In particular, the role of biochemical research in identifying the molecular and cellular mechanisms underpinning the protective/preventive role of nutraceutical molecules against aging and neurodegeneration will be considered. Neurodegenerative diseases, such as Alzheimer's and Parkinson’s diseases, are multifactorial disorders characterised by the progressive loss of neurons in the central nervous system. Although each neurodegenerative disease exhibits specific pathological features, they also share some common molecular mechanisms, such as abnormal protein aggregation, mitochondrial disfunction, neuroinflammation and oxidative stress. The lack of effective treatments and the multi-factorial etiology of these disorders has developed a pressing need to identify preventive/therapeutic compounds with pleiotropic activity. Among the many risk factors for neurodegenerative diseases, the aging process has the strongest impact and the investigation on basic mechanisms of aging and their role in the onset and progression of neurodegenerative disease are fundamental to develop effective preventive/therapeutic strategies.

Mechanisms of ligand binding
27/05/2022 27/05/2022 at 5 pm Enrico Di Cera MD Edward A. Doisy Department of Biochemistry and Molecular Biology, Saint Louis University School of Medicine, St. Louis
Ligand binding is at the basis of all biological interactions and has long been subject to theoretical and experimental investigation. The presentation reviews basic mechanisms of ligand binding, from lock and key to induced fit and conformational selection (1). A case is made for the dominance of conformational selection in biology. 1. Di Cera E. Mechanisms of ligand binding. Biophys Rev (2020) 1, 011303.

NADPH Oxidase: Structure, Enzymology and Drug Design
20/05/2022 Andrea Mattevi Department of Biology and Biotechnology University of Pavia, Italy
NADPH oxidases (NOXs) are the only known human enzymes solely in charge of ROS production. In addition to their roles in the innate immunity and response to stressful conditions, NOXs are part of the redox signaling pathways that sustain cell proliferation, oncoprotein (e.g. RAS) driven cell transformation, and tumor microenvironment manipulation. NOXs are tightly controlled and understanding the molecular mechanisms underlying their isoform-specific regulation is an open issue with far-reaching implications for drug design. In 2017, our group in Pavia has solved and published the first crystal structures of the dehydrogenase and transmembrane domains of a bacterial NOX5, 40% identical to human NOX5 [1-3]. Computer modelling was then used to view the overall structure of the NOX catalytic core. This landmark result revealed the structural basis for flavin reduction, “across-the-membrane” electron transfer, and ROS generation on the outer side of the membrane. In parallel to the structural studies, we have thoroughly investigated the putative NOX inhibitors that have been reported in the literature. This painstaking project led us to realize that virtually all known NOX inhibitors (31 of them were evaluated) suffer from off-targets effects, such as ROS scavenging and assay interference. This issue is so overwhelming that it was impossible to discern between the non-specific effects exerted by these compounds and their specific binding to NOXs (if any [4]). We have therefore employed the NOX5 dehydrogenase domain as our initial platform to carry on a drug design campaign. To this end, together with our collaborators at Harvard Medical School and Dana Farber Cancer Institute, we have conducted an ultra large-library computational screening using our NOX5 dehydrogenase domain PDB structure. We have evaluated the chosen library using a robust and high throughput workflow comprising primary and orthogonal assays as well as control assays to probe assay interfering compounds and ROS scavengers. Protein crystallography studies of the protein with the best hits have yielded for the first-time a crystal structure of this class of enzymes in complex with our studied inhibitors. Accordingly, our current efforts rely on the study and development of new and effective isoform-specific NOX inhibitors and the understanding of their effect on cancer model cells in which NOXs have a key role. [1] Magnani, F., Nenci, S., Millana Fananas, E., Ceccon, M., Romero, E., Fraaije, M.W., Mattevi*, A. (2017) Crystal structures and atomic model of NADPH oxidase. Proc. Natl. Acad. Sci. USA 114, 6764-6769. [2] Oosterheert, W., Reis, J., Gros, P., Mattevi, A. (2020) An Elegant Four-Helical Fold in NOX and STEAP Enzymes Facilitates Electron Transport across Biomembranes-Similar Vehicle, Different Destination. Acc. Chem. Res. 53, 1969-1980. [3] Magnani, F., Mattevi*, A. (2019) Structure and mechanisms of ROS generation by NADPH oxidases. Curr. Opin. Struct. Biol.59, 91-97. [4] Reis J, Massari M, Marchese S, Ceccon M, Aalbers FS, Corana F, Valente S, Mai A, Magnani F, Mattevi, A. (2020) A closer look into NADPH oxidase inhibitors: Validation and insight into their mechanism of action. Redox Biol. 32, 101466.

When viral RNA met the cell: a story of protein-RNA interactions
12/05/2022 11 am Alfredo Castello MRC-University of Glasgow Centre for Virus Research (UK)
RNA is a central molecule for the RNA virus life cycle as it functions not only as messenger for the synthesis of proteins, but also as storage of genetic information as genome. Given the central role of viral RNAs in infection, I hypothesised that it must be a hub for critical host-virus interactions. To test this, my laboratory has developed new approaches to elucidate the composition of viral ribonucleoproteins using Sindbis virus, SARS-CoV-2 and human immunodeficiency virus (HIV) as discovery models. With them, we have discovered a new universe of host-virus interactions with central regulatory roles in infection. Interestingly, these viruses, despite having different sequences and infection cycles, engage with a largely shared pool of cellular RNA-binding proteins. The efforts of my laboratory are now focused on understanding the molecular mechanisms underpinning these master regulators of virus infection, as we envision that they are promising targets for broad-spectrum antiviral therapies.

RNA-based therapeutic approaches for neurodegenerative tauopathies
06/05/2022 Michela A. Denti University of Trento
There are currently no disease-modifying therapies for the treatment of tauopathies, a group of progressive neurodegenerative disorders that are pathologically defined by the presence of tau protein aggregates in the brain. In Frontotemporal Lobar Degeneration (FTLD), a familial tauopathy, 57 different mutations in the microtubule-associated protein tau (MAPT) gene have been identified. About half of these mutations perturb the finely regulated balance in the splicing of MAPT exon 10, inducing the increase of exon 10-containing splicing isoforms. In turn, this affects the number of microtubule-binding domains present in the mature tau protein, ultimately causing neurodegeneration and tau aggregates. RNA Therapies are recently gaining their momentum and 12 approved RNA-based drugs have been approved as of today. We have designed three different RNA-based therapeutical approaches for FTLD, which might be beneficial also for other tauopathies: exon-skipping antisense oligonucleotides (AONs), AAV-vectored antisense chimeric U1 or U7 snRNAs and isoform-specific short interfering RNAs (siRNAs). Via reporter minigenes we have screened several of these molecules. We have validated the efficacy of the best hits for each approach in cultured cells. We are presently performing Proof-of-Concept studies in two orthogonal models of FTLD disease: human iPSc-derived neurons bearing the IVS10+16 mutation and a mouse model bearing the human MAPT gene with the same mutation and recapitulating the disease’s molecular, histopathological and behavioral aspects.

Metabolic control of T cell immunity
29/04/2022 Dott. Mauro Corrado, CECAD center, University of Cologne (Germany)
Tissue homeostasis is maintained via a fine balance between pro- and anti-inflammatory signals, a balance that is lost when mitochondrial metabolism is compromised in T cells, resulting in impaired immunity, multimorbidity and inflammation. The dynamic metabolism of T cells during an infection and upon nutrient stress depends indeed on the ability of T cells to fine tune their mitochondrial lipid composition (and in particular their content of the specific mitochondrial lipid cardiolipin). Nevertheless, how different tissue specific microenvironments and immune cells crosstalk and determine the damage to different tissues as well as the systemic outcome of a metabolic dysfunction in T cell is poorly understood. Our lab combines in vivo mouse phenotyping, classic immunology and biochemistry with quantitative proteomics and metabolomics to study the immune response in mouse models and patients affected by mitochondrial diseases.

Architecture of the human erythrocyte ankyrin-1 complex
22/04/2022 ORE 17.00 Francesca Vallese Dept. of Anesthesiology Columbia University, New York, USA
The ankyrin-1 complex tethers the spectrin-actin cytoskeleton to the red blood cell (RBC) membrane, and acts as a metabolic hub, connecting membrane proteins that are involved in gas exchange, pH control, and regulation of cellular volume and deformability. Mutations in components of the complex lead to inherited defects in erythrocyte shape and stability, such as hereditary spherocytosis. Ankyrins are also broadly expressed adaptors functioning as master-organizers of membrane-associated protein complexes in neurons and other cell types. However, the precise composition of the ankyrin-1 complex, and the structural basis for membrane association and recruitment of target membrane proteins remains unknown in any context. We solve the single particle cryo-EM structures of the human ankyrin-1 complex. Our structures reveal the architecture of this 1.2 MDa supercomplex, which includes the Rhesus polypeptides RhCE & RhAG, ankyrin 1, protein 4.2 and three copies of the dimeric band 3 anion exchanger bound to glycophorin A, assembled into a 1.2 MDa supercomplex. Additional complexes carrying one aquaporin-1 (AQP1) tetramer are also identified. The structure of membrane-bound ankyrin shows that the first five repeats adopt an unexpected T-shaped configuration whereby the inner groove is oriented parallel to the membrane, facilitating recognition of integral membrane binding partners such as RhCE and AQP1. Both the inner groove and the convex outer surface of ankyrin participate in specific interactions with protein 4.2 and band 3. Our structures also reveal the architecture and stoichiometry of the heterotrimeric Rh channel, highlighting the role of ankyrin in mediating clustering of multiple copies of structurally diverse membrane proteins.

Evolution of Klebsiella pneumoniae high-risk clones to pan-resistance
08/04/2022 Alessandra Carattoli Dept. Molecular Medicine Sapienza University of Rome
Infections caused by antibiotic-resistant Klebsiella pneumoniae are considered a major emerging problem in hospitals. K. pneumoniae behaves like an opportunistic bacterium, causing serious infections especially in the most fragile and long-term patients. In the nosocomial field, the greater burden of infections is due to the global dispersion of some high-risk clones of K. pneumoniae that spread successfully in Asia, the United States, Europe. Genomics made possible to analyze and understand the evolutionary strategies of these successful clones and their adaptation to antimicrobial use, leading to the emergence of clinically relevant mechanisms of resistance to antibiotics. Since the third generation cephalosporin resistance in the early 2000s, K. pneumoniae has progressively evolved acquiring resistance to carbapenems, colistin, tigecycline and more recently fosfomycin and new drug-inhibitor combinations such as ceftazidime-avibactam in the last decade. Pan-resistant clones (defined as cells resistant to all classes of antibiotics) and treatable only with the newest formulation drugs have been reported in the world and could quickly represent a microbiological problem of strategic relevance in clinical settings.

Mechanism of a cell wall transporter involved in lipoteichoic acid synthesis and bacterial adaptation
01/04/2022 Camilo Perez. Biozentrum, Basel, CH
Transport of lipids across membranes is fundamental for diverse biological pathways in cells. Multiple ion-coupled transporters take part in lipid translocation, but their mechanisms remain largely unknown. Major facilitator superfamily (MFS) lipid transporters play central roles in cell wall synthesis, brain development and function, lipids recycling, and cell signalling. Recent structures of MFS lipid transporters revealed overlapping architectural features pointing towards a common mechanism. We elucidated the structure of LtaA, and showed that it functions as a proton-dependent lipid antiporter, which contributes to the adaptation of the pathogen Staphylococcus aureus to acidic conditions, common in the skin and nasopharynx of the human host. LtaA takes part in the assembly of lipoteichoic acid, important for protection of S. aureus from environmental stress, host cell adhesion, antibiotic resistance, and immune evasion. The essential role of LtaA in adjusting the pool of glycolipids available for lipoteichoic acid assembly makes it a potential target for drugs aiming to counteract antimicrobial-resistant S. aureus strains e.g., methicillin-resistance S. aureus (MRSA) and vancomycin-resistant S. aureus (VRSA). Recently, we used cysteine disulfide trapping, molecular dynamics simulations, mutagenesis analysis, and transport assays in vitro and in vivo, to investigate the mechanism of LtaA. We revealed that LtaA displays asymmetric lateral openings with distinct functional relevance and that cycling through outward- and inward-facing conformations is essential for transport activity. We demonstrate that while the entire amphipathic central cavity of LtaA contributes to lipid binding, its hydrophilic pocket dictates substrate specificity. We propose that LtaA catalyzes lipid translocation by a ‘trap-and-flip’ mechanism that might be shared among MFS lipid transporters.

Amino acid metabolism: involvement in tumor progression and new therapeutic approaches
25/03/2022 Dott. Alessio Paone (Dept Biochemical Sciences, Sapienza University of Rome)
Metabolic adaptation is necessary for malignant tumor cells to survive the extreme conditions to which they are exposed within the mass of the primary tumor as well as throughout the metastatic process. In recent years it has been described that, despite having all the biosynthetic pathways intact, cancer cells of different origins are dependent on extracellular non-essential amino acids to support fundamental processes such as proliferation and survival. Here I will present data showing the centrality of amino acid metabolism, in particular of one-carbon metabolism, in the survival and metastatic ability of lung cancer cells. We studied serine hydroxymethyl transferases (SHMT), proteins fundamental in serine/glycine metabolism and revealed peculiar new properties such as their RNA binding ability. Taking advantage of these results, we are developing new approaches based on small RNA molecules to inhibit SHMTs, an enzyme for which effective small molecule inhibitors do not yet exist. We are also studying how the availability of selected amino acids (serine/glycine/glutamate) affect the ability of highly metastatic lung and breast cancer cells to select the brain as a target organ for metastasis formation. We clarified that these cells exploit extracellular amino acids to increase their extravasation ability, and that interfering with the import of these amino acids, which are abundant in the brain parenchyma, limits the formation of metastases in this organ, suggesting a novel therapeutic strategy to limit brain metastasis formation, a major cause of death in cancer patients.

Functional and Pathological Interactions of α synuclein
18/03/2022 Alfonso De Simone Department of Pharmacy, University of Naples Federico II
The aggregation of α-synuclein (αS), a neuronal protein that is abundant at the pre-synaptic terminals, is associated with a range of highly debilitating neurodegenerative conditions including Parkinson’s Disease (PD). Fibrillar aggregates of αS are the major constituents of proteinacious inclusions known as Lewy bodies that form in dopaminergic neurons of patients suffering from these conditions. The function of αS, however, is currently unknown, with evidences suggesting a role in the regulation of the trafficking of synaptic vesicles. We study the structure and interactions of αS in its functional state and in the form of pathological aggregates by means of biophysics and biomolecular NMR [1]. Our research has identified the nature of the physiological membrane interaction of αS and elucidated how this transient binding is involved functional processes such as the clustering of synaptic vesicles [2] or their docking onto the plasma membrane [3]. In the context of αS aggregation, we focus on the properties of elusive intermediate oligomers and how they impair neuronal function in the context of PD [4-5]. References 1. Fusco G, et al, (2014) Nat Commun 5:3827. 2. Man WK, et al, (2021) Nat Commun 12:927. 3. Fusco G, et al, (2016) Nat Commun 7:12563. 4. Fusco G, et al, (2017) Science 358:1440-3. 5. Cascella, et al, (2019) ACS chem bio 14:1352-1362.

Emerging role for m6A RNA modification in cancer: learning from leukemia
11/03/2022 Alessandro Fatica Dept. Biology and Biotechnology Sapienza
We are currently assisting at the explosion of the epitranscriptomics, which studies the functional role of chemical modifications into RNA molecules. Among more than 100 RNA modifications, the N6-methyladenosine (m6A) has attracted the interest of researchers all around the world. m6A is the most abundant internal chemical modification in mRNA and it can control any aspect of mRNA post-transcriptional regulation. m6A is installed by “writers”, removed by “erasers”, and recognized by “readers”, thus, it can be compared to the reversible and dynamic epigenetic modifications in histones and DNA. Given its fundamental role in determining the fate of mRNAs, it comes as no surprise that alterations to m6A modifications have a deep impact in cell differentiation, normal development and human diseases. Here, I will present the important role of m6A modification in gene expression and its contribution to cancer development. In particular, I will focus on myeloid leukaemia that, among first, has indicated how alteration in m6A modification can disrupt normal cellular proliferation and lead to cancer.

2021

Search for efficient diagnosis and therapy of advanced melanoma using biophysical methods
28/10/2021 at noon Prof. Tomasz Kobiela Warsaw University of Technology, Head of Laboratory of Biomolecular Interactions Studies
Melanoma, which originates from melanocytes, after entering the metastatic stage causes the highest mortality among skin malignant tumors resulting from the lack of effective therapy due to common resistance to the applied drugs [1]. In the view of increasing worldwide prevalence the intensive search for successful therapy pattern is undertaken with the use of combined strategy. This includes the combination of recognized anti-cancer drugs with compounds acting on various vital cellular signaling pathways, particularly on the energy metabolism. In melanoma, the mutation of the proto-oncogene B-Raf cytoplasmic serine–threonine kinase (BRAF) gene is the most common (over 50% of patients), which is why most of the applied drugs are directed at inhibiting that signaling pathway. For the optimization of the choice of possible compounds, we performed the modified SynGeNet drug combination prediction study [2]. Experimental validation of effective combinations was done by monitoring the interaction of specific lectins with cellular surface glycans typical for various stages of melanoma progression in real-time experiments using quartz crystal microbalance with the dissipation monitoring and atomic force microscopy [3]. Evaluation of the modification of the glycosylation process of metastatic melanoma cells as result of the applied combination of compounds could reveal potential usability in malignant melanoma treatment. [1] Sobiepanek, A. et al. (2021). European Biophysics Journal, 50, 523. [2] Regan-Fendt, K. E. et al. (2019). Npj Systems Biology and Applications, 5(1), 6. [3] Sobiepanek et al. (2017) Biosensors and Bioelectronics, 93:274-281

Structural Basis for SARS-CoV-2 Neutralization by Human Antibodies
18/06/2021 15.00 Dr. Gabriele Cerutti, Zuckerman Institute Columbia University, New York, USA
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent for COVID-19, emerged in late 2019, rapidly establishing an ongoing worldwide pandemic with more than a hundred million infected and over three million dead. In response, an unprecedented global effort to develop vaccines and therapeutics is well underway. One promising approach is the identification and structural characterization of SARS-CoV-2-neutralizing antibodies, which could be used as therapeutic or prophylactic agents. Potent neutralizing antibodies directed against SARS-CoV-2 spike, isolated from infected patients, target two main regions: the receptor binding domain (RBD) and the N-terminal domain (NTD). RBD-directed neutralizing antibodies target different epitopes on the domain and neutralize the virus by blocking receptor binding. NTD-directed neutralizing antibodies target a single supersite and their mechanism of action is less clear. Circulating SARS-CoV-2 variants seem to arise in response to human antibody pressure and the molecular basis for immune evasion or accommodation of mutants can be explained from a structural perspective.

Transcription factor EB in vascular and heart biology
04/06/2021, Prof. Federico Bussolino, Department of Oncology, University of Torino Candiolo Cancer Institute-IRCCS-FPO
TFEB (Transcription factor EB) represents an emerging player in the biology of cardiovascular system. TFEB was originally described to be translocated in a juvenile subset of paediatric renal cell carcinoma but whole genome sequencing reported somatic mutations sporadically found in many different cancers. Besides its oncogenic activity, TFEB controls the autophagy-lysosomal pathway by recognizing a recurrent motif present in the promoter regions of a set of genes that participate to lysosome biogenesis and its dysregulation is instrumental in the pathogenesis of lysosomal storage diseases. Emerging findings suggest that TFEB exerts wider regulatory activities in response to stress encompassing immunity, longevity and metabolism. In this seminar I’ll summarize the data of my Lab obtained by the specific deletion of Tfeb in mouse endothelial and epicardial cells. Our data demonstrate that TFEB activates specific genetic programs respectively regulating cell-cycle in endothelial cells and epithelial-mesenchymal transition in epicardial cells. Recognizing vascular and epicardial TFEB as a hub of a network of signals between tissues and bloodstream provides a fresh perspective on the molecular principles regulating organogenesis and tissue functions in physiology and pathology.

Molecular nature and regulation of the mitochondrial permeability transition pore
28/05/2021 Paolo Bernardi, Department of Biomedical Sciences and CNR Neuroscience Institute, University of Padova, Italy
Molecular nature and regulation of the mitochondrial permeability transition pore Paolo Bernardi* Department of Biomedical Sciences and CNR Neuroscience Institute, University of Padova, Italy The molecular nature of the mitochondrial permeability transition (PT) is a long-standing mystery of mitochondrial biology. Occurrence of the PT strictly requires matrix Ca2+ accumulation and is favored by matrix cyclophilin D, which mediates the inhibitory effects of cyclosporin A. Several hypotheses about the molecular nature of the PT have been put forward over the years. Today the prevailing view is that permeabilization of the inner membrane follows opening of a high-conductance channel, the PT pore, which is also called mitochondrial megachannel or multiconductance channel. I will provide an overview of the field with specific emphasis on the potential role of F-ATP synthase in channel formation.

The evolving spectrum of vitamin B6 responsive disorders
21/05/2021 Philippa Mills Great Ormond Street Institute of Child Health, University College London, London, UK.
Vitamin B6 is a micronutrient essential for normal physiological functioning in humans, serving as a cofactor for >70 human enzymes. Unlike lower organisms, humans cannot synthesise this vitamin and rely on dietary forms and those synthesised by the microbiome. These dietary forms include pyridoxine, pyridoxamine and pyridoxal and their 5’-phosphorylated derivatives. The intestine can only absorb the non-phosphorylated B6 vitamers, therefore phosphorylated forms must first be hydrolysed. Specific enzymes are then involved in the conversion of pyridoxine, pyridoxamine and pyridoxal to pyridoxal 5’-phosphate (PLP), the only active form of vitamin B6. Inborn errors of metabolism which affect the interconversion and availability of PLP, lead to a deficiency of this vitamer which manifests as an epilepsy that responds to treatment with supraphysiological doses of vitamin B6. This is not surprising given the vital role that PLP plays in neurotransmitter metabolism. Disorders include pyridox(am)ine phosphate oxidase deficiency (a disorder affecting PLP synthesis and recycling), disorders where metabolites accumulate that inactivate PLP, for example, ALDH7A1 deficiency and hyperprolinaemia type II, disorders which affect PLP import into the brain (hypophosphatasia and glycosylphosphatidylinositol anchor synthesis defects), and the recently described disorder in which mutations in an intracellular PLP-binding protein result in abnormal B6 homeostasis (PLPHP deficiency). We have shown recently however, by using next generation sequencing for the investigation of patients that have remained undiagnosed for many years, that: i) disorders of vitamin B6 deficiency do not always manifest as an epilepsy ii) supraphysiological doses of B6 can not only be used to treat the vitamin B6-dependent epilepsy disorders but also other seizure disorders that are not associated with vitamin B6 metabolism, such as the KCNQ2-related epileptic encephalopathies.

Diet and Metabolic Therapeutics in Cancer
14/05/2021 Jason LoCasale (Duke University School of Medicine, USA).
This presentation will discuss efforts to understand glucose and amino acid metabolism in cancer biology using metabolomics approaches. First, I will discuss new work on our understanding of central carbon metabolism. This part of the talk will focus on efforts to target cancer metabolism by disrupting the processing of macronutrient sources. Several examples of biological consequences of this pathway will then be presented. Next, I will focus on methionine metabolism. I will discuss work on dietary influences on the activity of the pathway and its relation to the regulation of one carbon metabolism in health. How methionine restricted diets may allow for interventions in cancer treatment will also be discussed. This concept also provides a link between nutrient status and chromatin biology which I will briefly touch upon.

“Identification of novel mechanisms in innate antiviral immunity”
07/05/2021 Søren Riis Paludan (Arhus Univ., DK)
The innate immune system is essential for host defense against virus infections. Type I interferon (IFN) is particularly important for early antiviral defense, and understanding of how IFN is induced is therefore of central importance. However, IFN also possesses inflammatory activities, and can contribute to disease development if produced in high amounts or over long time. Therefore, immune mechanisms that control virus infections without inducing IFN expression may contribute to a “silent” layer of the immune system. In this presentation, I will present our recent research on how herpes simplex virus induces expression of type I IFN, and also on novel constitutive immune mechanisms that control the virus.

Rett Syndrome as a model to study neurobiology and candidate therapeutics for brain disorders
30/04/2021 Daniela Tropea Trinity College (Dublin, Ireland)
Rett Syndrome (RTT) is a neurodevelopmental disorder associated to mutations in the X-linked gene MECP2, which codes for the protein Methyl-CpG binding protein 2. MeCP2 acts mainly a chromatin-binding protein, and determines activation or inhibition of gene expression, depending on its binding co-factors. Mecp2 is also involved in RNA splicing. Although mostly associated to RTT, MECP2 is involved in several other neuropsychiatric and neurological conditions, and its dysregulation (both upregulation and downregulation) has functional consequences. In addition, several cellular phenotypes identified in RTT are present in other brain disorders and treatment that benefit patients with RTT are also effective in other neurodevelopmental disorders. In this seminar I will present the evidence that Mecp2 controls many molecular mechanisms across different brain pathologies and that RTT can be used as a model to uncover the pathophysiology of several disorders.

The stressed synapses
23/04/2021 Tiziana Borsello, Università degli Studi di Milano
Synaptopathy define key features of many of neurodegenerative and psychiatric disorders. The synaptic degeneration, which undergo to a first reversible phase of “spine dysfuncion” leading to second irreversible phase of synaptic loss/death, which will lead to dendrite retraction and progress with neuronal death. Because synaptic injury precedes neuronal death and dysfunctional synapse possess a remarkable capacity for repair and functional recovery, we focus our efforts to develop a strategy to protect synapses during this early phase of the brain pathology. However, the intracellular mechanisms regulating synaptic dysfunction in AD are currently not fully understood. We identify the c-Jun N-terminal kinase (JNK) as a key player in AD synaptopathy. We characterized the JNK role in A oligomer-induced synaptopathy in vitro and in vivo in post-synaptic element describing the PSD alteration mediated by JNK. Our results demonstrated that the specific JNK inhibition (D-JNKI1) protected synapsed degeneration. Importantly JNK role in the post synaptic element is double: phophorylation of PSD-95 and Tau. JNK action on PSD-95 induced it down-relulation and PSD marker alterations, while JNK hyper-phosphorylation of Tau, altered Tau and Drebrin interaction, this leads to spine dysmorphogenesis. We then focused on JNK presynaptic localization and its role at this site. We proved that presynaptic fractions contained significant amount of JNK protein and its activated form. With biochemical approaches we demonstrated the interaction between JNK Synataxin-1,2 and Snap25. We defined JNK action on the SNARE complex formation and its role in modulation of vesicle release. These data showed JNK important functional role in post and pre-synaptic element. In conclusion, all together these results set the basis to develop a JNK-base therapeutic strategy to tackle synapse degeneration.

Understanding how RNA-based mechanisms control genome stability in cancer
16/04/2021 Lovorka Stojic - Barts Cancer Institute, Queen Mary University of London, London, UK
Genome stability is paramount to cellular homeostasis throughout the human lifespan. Cells have developed several surveillance mechanisms to protect the genome from mutations and ensure faithful duplication and transmission of the genetic material. Defects in any of these mechanisms leads to genome instability, which drives cancer evolution and contributes to tumour heterogeneity, drug resistance and poor prognosis. Protein-mediated mechanisms controlling genome stability are well described, however, the biological and regulatory function of RNA-based mechanisms in this context, and in particular the contribution of long noncoding RNAs (lncRNAs), is largely unknown. We have recently identified novel lncRNAs linked to chromosome mis-segregation, a process common to different types of cancer. I will focus on two nuclear localised lncRNAs whose expression is altered in cancer, and highlight mechanisms through which these lncRNAs safeguard genome integrity and their relevance to cancer.

Metabolic rewiring driving metastasis formation
09/04/2021 Sarah Maria Fendt (VIB-KU Leuven Center Cancer Biology)
Metabolic rewiring is a hallmark of cancer cells. However, how nutrients drive the ability of cancer cells to rewire their metabolism is poorly defined. We are investigating the in vivo nutrient metabolism during metastasis formation to mechanistically understand how nutrients from the microenvironment enable cancers to progress from a local to a systemic disease. Using 13C tracer infusions in mouse models we find that nutrient availability shapes the metabolism and phenotype of cells and subsequently promotes the progression of cancer. Consequently, interfering with nutrient metabolism emerges as a promising therapeutic strategy against cancer. Taken together, our research highlights that nutrient metabolism is an important driver of cancer progression.

RNA regulates Glycolysis and Embryonic Stem Cell Differentiation via Enolase 1
26/03/2021 Ina Huppertz, European Molecular Biology Laboratory (EMBL)
Metabolic pathways are transpiring to be significant regulatory sites that participate in controlling stem cell fates. One common feature in stem cell differentiation is the metabolic remodelling from aerobic glycolysis to respiration during the exit from pluripotency, with distinct paths taken by different germ layers. Since glycolytic enzymes have been reproducibly found to associate with RNA, we studied the conserved interaction of enolase 1 (ENO1) with RNA in vitro, in human cells, and during mouse embryonic stem cell differentiation. We show that ENO1 specifically binds RNA targets in human and murine cells. RNA ligands inhibit ENO1’s enzymatic activity in vitro, and ENO1’s enzymatic substrates specifically compete with its RNA binding. Increasing the concentration of RNA ligands in cultured cells inhibits glycolysis. We demonstrate that the differentiation of embryonic stem cells to specific germ layers involves changes in ENO1’s RNA binding. Importantly, pluripotent stem cells expressing an ENO1 mutant that is hyper-inhibited by RNA are severely impaired in their glycolytic capacity and in endodermal differentiation, whereas cells with an RNA binding-deficient ENO1 mutant display disproportionately high endodermal marker expression. The data uncover ENO1 riboregulation as a novel form of metabolic control. They also describe an unprecedented mechanism underlying the regulation of stem cell differentiation.

“Single particle cryo-electron microscopy, a quantum leap in structural biology”
19/03/2021 Martino Bolognesi (Dipartimento di Bioscienze e Centro di Ricerca Pediatrica Invernizzi Università di Milano)
The development of direct electron detectors, coupled to cryo-vitrification methods and developments of computational approaches, brought to the explosion of the “structural revolution” just a few years ago. Based on the above developments it is now possible to solve the 3D structure of macromolecular aggregates, protein:protein and protein:nucleic acid complexes in an almost direct approach, often reaching near atomic resolution. A real explosion of new structures and studies on macromolecular complexes is currently witnessed throughout the literature. In particular, it must be noted that specific macromolecular samples that could not be faced by previous techniques (e.g amyloid fibrils) are now within reach, shedding first light and entirely new studies on crucial research lines. I will present an overview of the single particle cryoEM method, explaining the basic principles behind, on one hand, and the practical lab aspects involved in sample/grid preparation, on the other. Following the theoretical aspects, I will briefly present the results of three research lines from our lab, focusing on the experimental approaches followed and on the biological implications of the results achieved.

“Analysis of protein-protein complexes and scoring of docking models”
12/03/2021 Romina Oliva (Dipartimento di Scienze e Tecnologie, Università di Napoli “Parthenope”Università Parthenope, Napoli)
Most proteins fulfill their functions through interaction with other proteins. A detailed structural analysis of the interaction between proteins in functional complexes is fundamental for understanding the mechanisms underlying biological processes and for possible biomedical and biotechnological applications. However, a dramatic disproportion still exists between the number of experimental structures solved for protein complexes and the number of structures available for single proteins. In this scenario, molecular docking, i.e. predicting the structure of a protein complex starting from the two separate components, is the method of choice for investigating the molecular basis of recognition in many functional biological systems. Reliably predicting the three-dimensional structure of protein-protein complexes by molecular docking is nonetheless an open challenge, with one of the critical steps being the scoring, i.e. the ability to discriminate between correct and incorrect solutions within a wide pool of generated models. In the last 10 years we have developed tools both for the analysis of the interface in biomolecular complexes and for the scoring of protein-protein docking models. Since 2013, our scoring algorithms have been blindly tested in CAPRI (Critical Assessment of PRedicted Interactions) experiments, docking challenges launched worldwide, where they proved to perform competitively with the state-of-the-art methods in the field.

“Templated Folding of Intrinsically Disordered Proteins”
05/03/2021 Angelo Toto (Department of Biochemical Sciences (Sapienza University)
Our understanding of protein biochemistry is mostly based on the structure-function relationship. This approach was revolutionized by the discovery that proteins can lack a well defined three dimensional structure, while being able to exert their functions in the cellular environment. These proteins are defined as Intrinsically Disordered Proteins, or IDPs. Since the discovery of protein disorder, IDPs have been extensively studied, and for many of them a disorder-to-order transition has been reported upon binding with their natural ligands. By focusing on the interaction between the transactivation domain of the cMyb protein, a prototypical IDP, and the globular KIX domain of the CBP protein, the fundamentals of the methodology used to study binding induced folding reactions will be recapitulated, together with the biophysical approach to interpret data. In particular it will be shown that the folding pathway of cMyb is malleable and dictated by the binding partner KIX. This mechanism is called “templated folding” and we propose it to be a general folding mechanism for IDPs.