Molecular mechanisms of selective autophagy in innate immunity

Selective autophagy is a catabolic route that engulfs cellular material on specialized vesicles called “autophagosomes” before turning them over to lysosomes for degradation. Such degradation substrates often interact and are anchored by the autophagosome-localized core autophagy protein LC3 (Atg8a in Drosophila). Autophagy is a pivotal process that helps in maintaining a homeostatic intracellular environment by contributing, among other functions, to the attenuation of innate immune signalling for several pathways, such as the IMD response in Drosophila. The IMD pathway, which this work focused on, is among the innate immune cascades that become increasingly harder to terminate with age; correlating with a concomitant waning of autophagy’s functions. The exact regulatory interactions between IMD pathway components and the autophagy machinery remain largely unknown. This work identified that the apical kinase of the Drosophila IMD pathway, dTAK1, as well as its co-activator dTAB2, interact with the autophagy protein Atg8a as examined by GSTpulldown experiments and confocal microscopy colocalization studies. Furthermore, the interaction between dTAK1 and Atg8a seems to rely on the functional LIR motif harboured in the C-terminal of dTAK1, that binds its cognate LDS region on Atg8a. Observations from qPCR assays that were employed to measure mRNA expression levels of the IMD-regulated genes AttA, DptB and Dro in conventionally reared young and old adult flies, suggest that the dTAK1 LIR motif prevents at least partly, the chronic overactivation of the IMD pathway. This study also further delineates the role of the Atg8a-interacting protein SH3PX1 in downregulating the IMD pathway, by characterizing the interaction between dTAB2 and SH3PX1, assessed by mass-spectroscopy and GST pulldown experiments. Genetic ablation of Sh3px1 correlates with both, increased levels of AttA, DptB and Dro that suggest overactivation of the IMD pathway in young and old Sh3px1-deficient flies, as well as their overall markedly reduced lifespan compared to controls in survival assays. Based on the insight gleaned by this study, I propose a mechanistic model for the dTAK1/dTAB2/SH3PX1 interactions with Atg8a, which may collectively mediate the selective autophagic degradation of the dTAK1/dTAB2 complex, and prevent in turn the constitutive activation of the IMD pathway in Drosophila. The conservation of all components in mammals provides encouraging evidence for potential similarities with the human condition as well

Caspase involvement in autophagy

Caspases are a family of cysteine proteases widely known as the principal mediators of the apoptotic cell death response, but considerably less so as the contributors to the regulation of pathways outside cellular demise. In regards to autophagy, the modulatory roles of caspases have only recently begun to be adequately described. In contrast to apoptosis, autophagy promotes cell survival by providing energy and nutrients through the lysosomal degradation of cytoplasmic constituents. Under basal conditions autophagy and apoptosis cross-regulate each other through an elaborate network of interconnections which also includes the interplay between autophagyrelated proteins (ATGs) and caspases. In this review we focus on the effects of this crosstalk at the cellular level, as we aim to concentrate the main observations from research conducted so far on the fine-tuning of autophagy by caspases. Several members of this protease-family have been found to directly interact with key ATGs involved in different tiers across the autophagic cascade. Therefore, we firstly outline the core mechanism of macroautophagy in brief. In an effort to emphasize the importance of the intricate cross-regulation of ATGs and caspases, we also present examples drawn from Drosophila and plant models regarding the contribution of autophagy to apoptotic cell death during normal development

Η επίδραση της σωματικής άσκησης στην ποιότητα ζωής ασθενών με πολλαπλή σκλήρυνση

Η πολλαπλή σκλήρυνση αποτελεί μια χρόνια, φλεγμονώδη ασθένεια του κεντρικού νευρικού συστήματος. Πρόκειται για ένα νόσημα το οποίο έχει σημαντική επίδραση στην καθημερινότητα των νοσούντων. Ο επιπολασμός της ασθένειας από τη δεκαετία του 1990 αυξάνεται με όλο και γοργότερους ρυθμούς και το 2020 υπολογίζεται ότι παγκοσμίως οι ασθενείς έφταναν τα 2,8 εκατομμύρια. Ενώ παλαιότερα η άσκηση δεν προτείνονταν στα άτομα αυτά, τα τελευταία χρόνια οι έρευνες δείχνουν ότι η φυσική άσκηση είναι μια πολλά υποσχόμενη μέθοδος διαχείρισης πολλών επιπτώσεων της ασθένειας. Στη παρούσα μελέτη, επιλέχθηκε η μεθοδολογία της βιβλιογραφικής ανασκόπησης προκειμένου να διερευνηθεί η επίδραση της σωματικής άσκησης στην ποιότητα ζωής των ατόμων με πολλαπλή σκλήρυνση. Τα τριάντα άρθρα που συμπεριλαμβάνονται στην έρευνα είναι πρόσφατα και καλύπτουν την περίοδο 2008 έως και 2022, με 19 από αυτά να έχουν δημοσιευθεί τα τελευταία έξι έτη. Σύμφωνα με τα αποτελέσματα της βιβλιογραφικής ανασκόπησης, οι τομείς όπου επιδρά θετικά η σωματική άσκηση στους ασθενείς με πολλαπλή σκλήρυνση, με σειρά συχνότητας εμφάνισης στα υπό μελέτη άρθρα είναι οι ακόλουθοι: μυϊκή δύναμη, κινητικότητα και ισορροπία, καρδιοαναπνευστική και καρδιαγγειακή λειτουργία, κόπωση, γενικό επίπεδο υγείας και ποιότητα ζωής, κατάθλιψη, διανοητική λειτουργία (συγκέντρωση, μνήμη, προσοχή), μυϊκή μάζα, ενεργοποίηση νεύρων και θετική επίδραση στο μικρό-περιβάλλον του κεντρικού νευρικού συστήματος, αυτοπεποίθηση και αίσθημα αυτοπραγμάτωσης, παχυσαρκία, σπαστικότητα, υποτροπής της ασθένειας, ποιότητα ύπνου, πόνος, άγχος και σεξουαλική λειτουργία. Η άσκηση ενδυνάμωσης αποτελεί το περισσότερο ενδεδειγμένο είδος άσκησης, στην περίπτωση των ατόμων με πολλαπλή σκλήρυνση, αφού συμβάλλει στη βελτίωση σε έντεκα υπό διερεύνηση τομείς. Η αεροβική άσκηση αποτελεί τη δεύτερη πιο ενδεδειγμένη μέθοδο άσκησης για τους ασθενείς με πολλαπλή σκλήρυνση. Επίσης, προτείνεται η άσκηση ισορροπίας και η άσκηση αντοχής. Η μελέτη μπορεί να αποδειχθεί ένα χρήσιμο εργαλείο για τους επαγγελματίες γυμναστές οι οποίοι επιθυμούν να ασχοληθούν – ή ασχολούνται ήδη - με την αποκατάσταση ασθενών με Πολλαπλή Σκλήρυνση.Multiple Sclerosis represents a chronicle, inflammatory disease of the Central Neural System. It is a disease with important impact on the daily life of patients. The prevalence of the disease during since the 1990s is more and more intense and it is estimated that, in 2020, patients were about 2.8 million, at a global level. While during the previous years, exercise was not recommended in patents with multiple sclerosis, during the last years, research reveals that exercise represents a promising method for handling a lot of negative consequences of the disease. For the purposes of the present study, the methodology of the literature review was chosen, so as to investigate the impact of physical exercise in the quality of life of people with multiple sclerosis. The 30 articles that are included in the research are very recent and cover the period 2008-2022, with 19 among them being published during the last six years. According to the research results, the areas where physical exercise has positive impact for patients suffering from multiple sclerosis include the following (in order of frequency): muscle strength, mobility and balance, cardiorespiratory and cardiovascular operation, fatigue, general health status and quality of life, depression, cognitive ability (concentration, memory, attention), muscle mass, neural activity and positive impact on the micro environment of the central neural system, self-confidence and self-realization, obesity, spasticity, relapse of the disease, sleep quality, pain, anxiety and sexual function. Empowerment exercise represents the most suitable type of exercise in the case of people with multiple sclerosis, since it contributes to the improvement of eleven different areas of symptoms. Aerobic exercise is the second most preferable type of exercise. Also, balance exercise and endurance exercise are recommended. The present study can be proven as a useful tool for professional trainers who wish to wish to work – or are already working – with the rehabilitation of patients suffering from multiple sclerosis

A yeast -two-hybrid screening identifies novel Atg8a-interacting proteins in Drosophila

Macroautophagy/autophagy-related protein Atg8/LC3 is important for autophagosome biogenesis and required for selective degradation of various substrates. In our recent study, we performed a yeast two-hybrid screening to identify proteins that interact with Atg8a, the Drosophila homolog of Atg8/LC3. The screening identified several Atg8a-interacting proteins. These proteins include: i) proteins which have already been experimentally verified to bind Atg8a, such as Atg1, DOR, ref(2)P and key (Kenny); ii) proteins for which their mammalian homologs interact with Atg8-family members, like Ank2, Atg4, and Nedd4; and iii) several novel Atg8a-interacting proteins, such as trc/STK38 and Tak1. We showed that Tak1, as well as its co-activator, Tab2, both interact with Atg8a and are substrates for selective autophagic clearance. We also determined that SH3PX1 interacts with Tab2 and is necessary for the effective regulation of the immune-deficiency (IMD) pathway. Our findings suggest a mechanism for the regulatory interactions between Tak1-Tab2-SH3PX1 and Atg8a, which contribute to the fine-tuning of the IMD pathway

Selective autophagy controls innate immune response through a TAK1/TAB2/SH3PX1 axis

Selective autophagy is a catabolic route which turns over specific cellular material for degradation by lysosomes, and whose role in the regulation of innate immunity is largely unexplored. Here, we show that the apical kinase of the Drosophila immune deficiency (IMD) pathway Tak1, as well as its co-activator Tab2, are both selective autophagy substrates that interact with the autophagy protein Atg8a. We also present a role for the Atg8a-interacting protein Sh3px1 in the downregulation of the IMD pathway, by facilitating targeting of the Tak1/Tab2 complex to the autophagy platform through its interaction with Tab2. Our findings show the Tak1/Tab2/Sh3px1 interactions with Atg8a mediate the removal of the Tak1/Tab2 signaling complex by selective autophagy. This in turn prevents constitutive activation of the IMD pathway in Drosophila. This study provides mechanistic insight on the regulation of innate immune responses by selective autophagy

Antibacterial activity of vB_AbaM_PhT2 phage hydrophobic amino acid fusion endolysin, combined with colistin against Acinetobacter baumannii

Abstract Phage lytic enzymes are promising antimicrobial agents. In this study, an endolysin derived from vB_AbaM_PhT2 (vPhT2), was identified. This endolysin represented the conserved lysozyme domain. Recombinant endolysin (lysAB- vT2) and hydrophobic fusion endolysin (lysAB-vT2-fusion) were expressed and purified. Both endolysins showed lytic activity against bacterial crude cell wall of Gram-negative bacteria. The MIC of lysAB-vT2-fusion was 2 mg/ml corresponding to 100 µM, while the MIC of lysAB-vT2 was more than 10 mg/ml (400 µM). Combination of lysAB-vT2-fusion with colistin, polymyxin B or copper was synergistic against A. baumannii (FICI value as 0.25). Antibacterial activity of lysAB-vT2-fusion plus colistin at the fractional inhibitory concentrations (FICs) revealed that it can inhibit Escherichia coli, Klebsiella pneumoniae and various strains of extremely drug-resistant A. baumannii (XDRAB) and phage resistant A. baumannii. The lysAB- vT2-fusion still retained its antibacterial activity after incubating the enzyme at 4, 20, 40 and 60 °C for 30 min. The lysAB-vT2-fusion could inhibit the mature biofilm, and incubation of lysAB-vT2-fusion with T24 human cells infected with A. baumannii led to a partial reduction of LDH release from T24 cells. In summary, our study highlights the antimicrobial ability of engineered lysAB-vT2-fusion endolysin, which can be applied for the control of A. baumannii infection

Atg8 family proteins, LIR/AIM motifs and other interaction modes

The Atg8 family of ubiquitin-like proteins play pivotal roles in autophagy and other processes involving vesicle fusion and transport where the lysosome/vacuole is the end station. Nuclear roles of Atg8 proteins are also emerging. Here, we review the structural and functional features of Atg8 family proteins and their protein-protein interaction modes in model organisms such as yeast, Arabidopsis, C. elegans and Drosophila to humans. Although varying in number of homologs, from one in yeast to seven in humans, and more than ten in some plants, there is a strong evolutionary conservation of structural features and interaction modes. The most prominent interaction mode is between the LC3 interacting region (LIR), also called Atg8 interacting motif (AIM), binding to the LIR docking site (LDS) in Atg8 homologs. There are variants of these motifs like “half-LIRs” and helical LIRs. We discuss details of the binding modes and how selectivity is achieved as well as the role of multivalent LIR-LDS interactions in selective autophagy. A number of LIR-LDS interactions are known to be regulated by phosphorylation. New methods to predict LIR motifs in proteins have emerged that will aid in discovery and analyses. There are also other interaction surfaces than the LDS becoming known where we presently lack detailed structural information, like the N-terminal arm region and the UIM-docking site (UDS). More interaction modes are likely to be discovered in future studies