Associative Conditioning Is a Robust Systemic Behavior in Unicellular Organisms: An Interspecies Comparison

The capacity to learn new efficient systemic behavior is a fundamental issue of contemporary biology. We have recently observed, in a preliminary analysis, the emergence of conditioned behavior in some individual amoebae cells. In these experiments, cells were able to acquire new migratory patterns and remember them for long periods of their cellular cycle, forgetting them later on. Here, following a similar conceptual framework of Pavlov’s experiments, we have exhaustively studied the migration trajectories of more than 2000 individual cells belonging to three different species: Amoeba proteus, Metamoeba leningradensis, and Amoeba borokensis. Fundamentally, we have analyzed several relevant properties of conditioned cells, such as the intensity of the responses, the directionality persistence, the total distance traveled, the directionality ratio, the average speed, and the persistence times. We have observed that cells belonging to these three species can modify the systemic response to a specific stimulus by associative conditioning. Our main analysis shows that such new behavior is very robust and presents a similar structure of migration patterns in the three species, which was characterized by the presence of conditioning for long periods, remarkable straightness in their trajectories and strong directional persistence. Our experimental and quantitative results, compared with other studies on complex cellular responses in bacteria, protozoa, fungus-like organisms and metazoans that we discus here, allow us to conclude that cellular associative conditioning might be a widespread characteristic of unicellular organisms. This new systemic behavior could be essential to understand some key principles involved in increasing the cellular adaptive fitness to microenvironments.This work was supported by a grant of the University of Basque Country (UPV/EHU), GIU17/066, the Basque Government grant IT974-16, the UPV/EHU and Basque Center of Applied Mathematics, grant US18/21, and the Israel Science Foundation (536/19)Peer reviewe

ICAM-1 nanoclusters regulate hepatic epithelial cell polarity by leukocyte adhesion-independent control of apical actomyosin

Epithelial intercellular adhesion molecule (ICAM)-1 is apically polarized, interacts with, and guides leukocytes across epithelial barriers. Polarized hepatic epithelia organize their apical membrane domain into bile canaliculi and ducts, which are not accessible to circulating immune cells but that nevertheless confine most of ICAM-1. Here, by analyzing ICAM-1_KO human hepatic cells, liver organoids from ICAM-1_KO mice and rescue-of-function experiments, we show that ICAM-1 regulates epithelial apicobasal polarity in a leukocyte adhesion-independent manner. ICAM-1 signals to an actomyosin network at the base of canalicular microvilli, thereby controlling the dynamics and size of bile canalicular-like structures. We identified the scaffolding protein EBP50/NHERF1/ SLC9A3R1, which connects membrane proteins with the underlying actin cytoskeleton, in the proximity interactome of ICAM-1. EBP50 and ICAM-1 form nano-scale domains that overlap in microvilli, from which ICAM-1 regulates EBP50 nano-organization. Indeed, EBP50 expression is required for ICAM-1-mediated control of BC morphogenesis and actomyosin. Our findings indicate that ICAM-1 regulates the dynamics of epithelial apical membrane domains beyond its role as a heterotypic cell– cell adhesion molecule and reveal potential therapeutic strategies for preserving epithelial architecture during inflammatory stress

Genomic transformation and social organization during the Copper Age-Bronze Age transition in southern Iberia

The emerging Bronze Age (BA) of southeastern Iberia saw marked social changes. Late Copper Age (CA) settlements were abandoned in favor of hilltop sites, and collective graves were largely replaced by single or double burials with often distinctive grave goods indirectly reflecting a hierarchical social organization, as exemplified by the BA El Argar group. We explored this transition from a genomic viewpoint by tripling the amount of data available for this period. Concomitant with the rise of El Argar starting ~2200 cal BCE, we observe a complete turnover of Y-chromosome lineages along with the arrival of steppe-related ancestry. This pattern is consistent with a founder effect in male lineages, supported by our finding that males shared more relatives at sites than females. However, simple two-source models do not find support in some El Argar groups, suggesting additional genetic contributions from the Mediterranean that could predate the BA

ICAM-1 nanoclusters regulate hepatic epithelial cell polarity by leukocyte adhesion-independent control of apical actomyosin

Epithelial intercellular adhesion molecule (ICAM)-1 is apically polarized, interacts with, and guides leukocytes across epithelial barriers. Polarized hepatic epithelia organize their apical membrane domain into bile canaliculi and ducts, which are not accessible to circulating immune cells but that nevertheless confine most of ICAM-1. Here, by analyzing ICAM-1_KO human hepatic cells, liver organoids from ICAM-1_KO mice and rescue-of-function experiments, we show that ICAM-1 regulates epithelial apicobasal polarity in a leukocyte adhesion-independent manner. ICAM-1 signals to an actomyosin network at the base of canalicular microvilli, thereby controlling the dynamics and size of bile canalicular-like structures. We identified the scaffolding protein EBP50/NHERF1/SLC9A3R1, which connects membrane proteins with the underlying actin cytoskeleton, in the proximity interactome of ICAM-1. EBP50 and ICAM-1 form nano-scale domains that overlap in microvilli, from which ICAM-1 regulates EBP50 nano-organization. Indeed, EBP50 expression is required for ICAM-1-mediated control of BC morphogenesis and actomyosin. Our findings indicate that ICAM-1 regulates the dynamics of epithelial apical membrane domains beyond its role as a heterotypic cell–cell adhesion molecule and reveal potential therapeutic strategies for preserving epithelial architecture during inflammatory stress

Stability of SARS-CoV-2 spike antigens against mutations

Modern health care needs preventive vaccines and therapeutic treatments with stability against pathogen mutations to cope with current and future viral infections. At the beginning of the COVID-19 pandemic, our analytic and predictive tool identified a set of eight short SARS-CoV-2 S-spike protein epitopes that had the potential to persistently avoid mutation. Here a combination of genetic, Systems Biology and protein structure analyses confirm the stability of our identified epitopes against viral mutations. Remarkably, this research spans the whole period of the pandemic, during which 93.9% of the eight peptides remained invariable in the globally predominant 43 circulating variants, including Omicron. Likewise, the selected epitopes are conserved in 97% of all 1,514 known SARS-CoV-2 lineages. Finally, experimental analyses performed with these short peptides showed their specific immunoreactivity. This work opens a new perspective on the design of next-generation vaccines and antibody therapies that will remain reliable against future pathogen mutations.Dr. Lozano-Perez acknowledges the European Commission ERDF/FEDER Operational Program 'Murcia' CCI No. 2007ES161PO001 (Project No. 14-20/20). Miodrag Grbic acknowledges support from the NSERC Discovery grant (Canada). This work also has received funding from the Department of Education of the Basque Government via the Consolidated Research Group MATH MODE (IT1456-22). Besides, Ildefonso Martinez De la Fuente and Iker Malaina were supported by the UPV/EHU and Basque Center of Applied Mathematics, grant US21/27N

La biosíntesis de trehalosa como potencial diana antifúngica en candida albicans

Introducción: En las últimas décadas se ha producido un enorme incremento de las infecciones causadas por hongos, siendo el género Candida el agente etiológico con mayor prevalencia, afectando en mayor medida a la población humana de mayor edad o aquejada por algún tipo de inmunodeficiencia. De hecho, Candida albicans es la cuarta causa de infecciones nosocomiales. La quimioterapia antifúngica necesita el desarrollo de compuestos más potentes, seguros y con menor toxicidad. En este contexto, el disacárido no reductor trehalosa ha sido propuesto como una interesante diana terapéutica para el diseño de nuevos antifúngicos. Objetivos: Con el fin de profundizar en esta hipótesis, hemos examinado en el patógeno oportunista Candida albicans, el nivel de susceptibilidad que presentan los mutantes homocigóticos deficientes en los dos genes que codifican las etapas secuenciales en la biosíntesis de trehalose, denominados respectivamente como trehalosa sintasa (TPS1) y trehalosa fosfatasa (TPS2), frente a los siguientes antifúngicos de uso clínico: Anfotericina B (AmB), Micafungina (MF) y Fluconazol (Flz). Métodos: En el transcurso del trabajo experimental, se ha llevado a cabo la determinación de varios parámetros fisiológicos y bioquímicos. En concreto, podemos destacar el recuento de células viables y la sensibilidad en placa, la medida de la producción intracelular de ROS, el potencial de membrana mitocondrial, el contenido de trehalosa y la formación de biofilms. Mencionar igualmente, el ensayo de enzimas antioxidantes, junto con la extracción y metilación de extractos lipídicos. Resultados: Mientras el mutante tps1Δ se comportó como muy sensible frente a la exposición con AmB, mostró un significativo nivel de resistencia a MF. Curiosamente, el fenotipo opuesto se registró en el mutante tps2Δ. A su vez, la MF promovió un cierto incremento en la producción endógena de ROS en la cepa parental SC5314 y en células tps2Δ, mientras la formación de ROS en el mutante tps1Δ fue virtualmente indetectable. En este mismo mutante tps1Δ el tratamiento con Flz mostró una ligera susceptibilidad que no puede considerarse relevante. El nivel endógeno de ROS se correlaciona positivamente con la actividad mitocondrial. La aplicación de Flz no causó variaciones de importancia en la generación de estrés oxidativo en ninguna de las levaduras analizadas. Tan sólo AmB fue capaz de promover la síntesis intracelular neta de trehalosa en la cepa parental SC5314, mientras el disacárido estaba ausente en células tps1Δ y mostró valores residuales en las tps2Δ, confirmando que otras fosfatasas citosólicas distintas a la enzima tps2p pueden llevar a cabo la defosforilación de trehalosa-6P in C. albicans. Además, la capacidad de ambos mutantes tps1Δ y tps2Δ para formar biofilms experimentó una drástica disminución tras el tratamiento con AmB; si bien experimentó un ligero aumento en células tps1Δ después de añadir MF. Los análisis de los respectivos perfiles lipídicos no fueron concluyentes, sin que ninguna evidencia pueda ser aportada, por el momento, que permita explicar la diferente sensibilidad a los antifúngicos de las cepas de C. albicans bajo estudio. Conclusión: Nuestros datos aportan consistencia a la idea de utilizar trehalosa como una interesante diana para nuevas terapias antifúngicas.   Abstract Introduction: The fungal infections have dramatically increased in the last decades, being the genus Candida the most prevalent etiological agent, which mainly affect to the immnunocompromised and aging human population. In fact, Candida albicans being the fourth etiological agent of nosocomial infections. The antifungal chemotherapy needs the development of safer, more potent and less toxic compounds. In this context, the non-reducing disaccharide trehalose has been proposed as a promising target for the design of new antifungals. Objectives: To further explore this hypothesis, we have examined in the opportunistic pathogen Candida albicans the degree of susceptibility shown by null mutants disrupted in the two genes coding for the sequential steps of trehalose biosynthesis, namely trehalose synthase (TPS1) and trehalose phosphatase (TPS2), to Amphotericin B (AmB), Micafungin (MF) and Fluconazol (Flz). Methods: We have carried the determination of several physiological and biochemical parameters. We remark the cell viability and sensitivity in plate, the measurement of intracellular ROS, mitochondrial membrane potential and the trehalose content and formation of biofilms. Furthermore, the assays of antioxidant enzymes and the fat extraction and methylation of the lipid extract. Results: While tps1Δ mutant was highly sensitive to AmB exposure, it displayed a significant level of resistance to MF. Notably, the opposite phenotype was recorded in the tps2Δ mutant. In turn, MF induced a significant level of endogenous ROS production in the parental SC5314 strain and tps2Δ cells, whereas ROS formation in the tps1Δ mutant was virtually undetectable. In turn, the tps1Δ cells after Flz treatment showed a slight susceptibility, which was not relevant.The level of endogenous ROS positively correlated with the mitochondrial activity. The application of Flz did not cause variations in the generation of oxidative stress in the yeasts analyzed. Only AmB was able to promote the net intracellular synthesis of trehalose in the parental strain SC5314, although it was absent from tps1Δ cells and showed only low levels in tps2Δ, confirming that other phosphatases distinct of tps2p can bring about the dephosphorylation of trehalose-6P in C. albicans. Furthermore, the capacity of both tps1Δ and tps2Δ mutants to form biofilms was drastically reduced after treatment with AmB, although it increased in tps1Δ cells after the addition of MF. The analyses of the lipid profile were not conclusive. No evidence could be found that can explain the different susceptibility to the antifungals of the C. albicans strains used in this study. Conclusion: Our data lend weight to the idea of using trehalose as a target for antifungal therapy