Browsing by Author "Yuseff, Maria Isabel"

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    Antibody to AP1B adaptor blocks biosynthetic and recycling routes of basolateral proteins at recycling endosomes
    (AMER SOC CELL BIOLOGY, 2007) Cancino, Jorge; Torrealba, Carolina; Soza, Andrea; Yuseff, Maria Isabel; Gravotta, Diego; Henklein, Peter; Rodriguez Boulan, Enrique; Gonzalez, Alfonso
    The epithelial-specific adaptor AP1B sorts basolateral plasma membrane (PM) proteins in both biosynthetic and recycling routes, but the site where it carries out this function remains incompletely defined. Here, we have investigated this topic in Fischer rat thyroid (FRT) epithelial cells using an antibody against the medium subunit mu 1B. This antibody was suitable for immunofluorescence and blocked the function of AP1B in these cells. The antibody blocked the basolateral recycling of two basolateral PM markers, Transferrin receptor (TfR) and LDL receptor (LDLR), in a perinuclear compartment with marker and functional characteristics of recycling endosomes (RE). Live imaging experiments demonstrated that in the presence of the antibody two newly synthesized GFP-tagged basolateral proteins (vesicular stomatitis virus G [VSVG] protein and TfR) exited the trans-Golgi network (TGN) normally but became blocked at the RE within 3-5 min. By contrast, the antibody did not block trafficking of green fluorescent protein (GFP)-LDLR from the TGN to the PM but stopped its recycling after internalization into RE in similar to 45 min. Our experiments conclusively demonstrate that 1) AP1B functions exclusively at RE; 2) TGN-to-RE transport is very fast and selective and is mediated by adaptors different from AP1B; and 3) the TGN and AP1B-containing RE cooperate in biosynthetic basolateral sorting.
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    B cell mechanosensing regulates ER remodeling at the immune synapse
    (2024) Riobo, Isidora; Yuseff, Maria Isabel
    Introduction Engagement of the B-cell receptor with immobilized antigens triggers the formation of an immune synapse (IS), a complex cellular platform where B-cells recruit signaling molecules and reposition lysosomes to promote antigen uptake and processing. Calcium efflux from the endoplasmic reticulum (ER) released upon BCR stimulation is necessary to promote B-cell survival and differentiation. Whether the spatial organization of the ER within the B-cell synapse can tune IS function and B-cell activation remains unaddressed. Here, we characterized ER structure and interaction with the microtubule network during BCR activation and evaluated how mechanical cues arising from antigen presenting surfaces affect this process.Methods B-cells were cultured on surfaces of varying stiffness coated with BCR ligands, fixed, and stained for the ER and microtubule network. Imaging analysis was used to assess the distribution of the ER and microtubules at the IS.Results Upon BCR activation, the ER is redistributed towards the IS independently of peripheral microtubules and accumulates around the microtubule-organization center. Furthermore, this remodeling is also dependent on substrate stiffness, where greater stiffness triggers enhanced redistribution of the ER.Discussion Our results highlight how spatial reorganization of the ER is coupled to the context of antigen recognition and could tune B-cell responses. Additionally, we provide novel evidence that the structural maturation of the ER in plasma cells is initiated during early activation of B-cells.
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    Clinical and pulmonary function analysis in long-COVID revealed that long-term pulmonary dysfunction is associated with vascular inflammation pathways and metabolic syndrome
    (FRONTIERS MEDIA SA, 2023) Sanhueza, Sergio; Vidal, Mabel A.; Hernandez, Mauricio A.; Henriquez-Beltran, Mario E.; Cabrera, Camilo; Quiroga, Romina; Antilef, Barbara E.; Aguilar, Kevin P.; Castillo, Daniela A.; Llerena, Faryd J.; Figueroa, Marco Fraga; Nazal, Mauricio; Castro, Eritson; Lagos, Paola; Moreno, Alexa; Lastra, Jaime J.; Gajardo, Jorge; Garces, Pamela; Riffo, Benilde; Buchert, Jorge; Sanhueza, Rocio; Ormazaba, Valeska; Saldivia, Pablo; Vargas, Cristian; Nourdin, Guillermo; Koch, Elard; Zuniga, Felipe A.; Lamperti, Liliana; Bustos, Paula; Guzman-Gutierrez, Enrique; Tapia, Claudio A.; Ferrada, Luciano; Cerda, Gustavo; Woehlbier, Ute; Riquelme, Marcelo; Yuseff, Maria Isabel; Ramirez, Braulio A. Munoz; Lombardi, Giovanna; De Gonzalo-Calvo, David; Salomon, Carlos; Verdugo, Ricardo A.; Quinones, Luis A.; Colombo, Alicia; Barria, Maria I.; Labarca, Gonzalo; Nova-Lamperti, Estefania
    Introduction: Long-term pulmonary dysfunction (L-TPD) is one of the most critical manifestations of long-COVID. This lung affection has been associated with disease severity during the acute phase and the presence of previous comorbidities, however, the clinical manifestations, the concomitant consequences and the molecular pathways supporting this clinical condition remain unknown. The aim of this study was to identify and characterize L-TPD in patients with long-COVID and elucidate the main pathways and long-term consequences attributed to this condition by analyzing clinical parameters and functional tests supported by machine learning and serum proteome profiling. Methods: Patients with L-TPD were classified according to the results of their computer-tomography (CT) scan and diffusing capacity of the lungs for carbon monoxide adjusted for hemoglobin (DLCOc) tests at 4 and 12-months post-infection. Results: Regarding the acute phase, our data showed that L-TPD was favored in elderly patients with hypertension or insulin resistance, supported by pathways associated with vascular inflammation and chemotaxis of phagocytes, according to computer proteomics. Then, at 4-months post-infection, clinical and functional tests revealed that L-TPD patients exhibited a restrictive lung condition, impaired aerobic capacity and reduced muscular strength. At this time point, high circulating levels of platelets and CXCL9, and an inhibited FCgamma-receptor-mediated-phagocytosis due to reduced Fc gamma RIII (CD16) expression in CD14+ monocytes was observed in patients with L-TPD. Finally, 1-year post infection, patients with L-TPD worsened metabolic syndrome and augmented body mass index in comparison with other patient groups. Discussion: Overall, our data demonstrated that CT scan and DLCOc identified patients with L-TPD after COVID-19. This condition was associated with vascular inflammation and impair phagocytosis of virus-antibody immune complexes by reduced Fc gamma RIII expression. In addition, we conclude that COVID-19 survivors required a personalized follow-up and adequate intervention to reduce long-term sequelae and the appearance of further metabolic diseases.
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    Ecm29-Dependent Proteasome Localization Regulates Cytoskeleton Remodeling at the Immune Synapse
    (2021) Ibanez-Vega, Jorge; Del Valle, Felipe; Saez, Juan Jose; Guzman, Fanny; Diaz, Jheimmy; Soza, Andrea; Yuseff, Maria Isabel
    The formation of an immune synapse (IS) enables B cells to capture membrane-tethered antigens, where cortical actin cytoskeleton remodeling regulates cell spreading and depletion of F-actin at the centrosome promotes the recruitment of lysosomes to facilitate antigen extraction. How B cells regulate both pools of actin, remains poorly understood. We report here that decreased F-actin at the centrosome and IS relies on the distribution of the proteasome, regulated by Ecm29. Silencing Ecm29 decreases the proteasome pool associated to the centrosome of B cells and shifts its accumulation to the cell cortex and IS. Accordingly, Ecm29-silenced B cells display increased F-actin at the centrosome, impaired centrosome and lysosome repositioning to the IS and defective antigen extraction and presentation. Ecm29-silenced B cells, which accumulate higher levels of proteasome at the cell cortex, display decreased actin retrograde flow in lamellipodia and enhanced spreading responses. Our findings support a model where B the asymmetric distribution of the proteasome, mediated by Ecm29, coordinates actin dynamics at the centrosome and the IS, promoting lysosome recruitment and cell spreading.