Biology subjects

Vitorino, R.

Publications and source records attributed to Vitorino, R..

6 recordsLinked to original sources

bioRxiv2024

BCL2L13 Influences Autophagy and Ceramide Metabolism without Affecting Temozolomide Resistance in Glioblastoma

Temozolomide (TMZ) resistance in glioblastoma (GBM) arises through metabolic rewiring that links mitochondrial function, autophagy balance, and sphingolipid metabolism. TMZ resistant (R) U251 cells exhibited suppressed apoptosis and complete blockade of autophagy flux, evidenced by LC3II and p62 accumulation and insensitivity to Bafilomycin A1. BCL2L13, strongly upregulated in R cells, emerged as a dual regulator of mitophagy and ceramide metabolism. BCL2L13 knockdown (KD) produced opposite effects in TMZ sensitive (NR) and resistant cells: in NR cells, KD elevated LC3II, reduced respiratory reserve, and triggered compensatory lipid synthesis; in R cells, KD decreased LC3II without restoring flux or TMZ sensitivity. Lipidomic profiling revealed that BCL2L13 loss reactivated CerS6 in NR cells, increasing C16:0 and mid-chain ceramides, while relieving CerS2 inhibition in R cells, elevating very long chain (C22 to C24) and glycosylated ceramides. These distinct sphingolipid signatures were confirmed by PLS-DA and KEGG enrichment, which highlighted steroid hormone, arachidonic, and linoleic acid metabolism in NR KD cells versus neuroactive ligand-receptor and signaling pathways in R KD cells. Together, these findings position BCL2L13 as a molecular integrator of mitochondrial respiration, autophagy flux, and CerS-dependent lipid remodeling, unveiling a context-specific metabolic mechanism that supports GBM cell survival under chemotherapeutic stress.

cancer biology↗

bioRxiv2024

A Lipidomics Approach to Determine the Role of Lipids and its Crosstalk with Autophagy in Lung Cancer Metastasis

Summary/AbstractNon-small cell lung cancer (NSCLC) is among the most malignant tumors with high propensity for metastasis and is the leading cause of cancer-related death globally. Most patients present with regional and distant metastasis, associated with poor prognosis. Lipids may play an essential role in either activating or inhibiting detachment-induced apoptosis (anoikis), where the latter is a crucial mechanism to prevent metastasis, and it may have a cross-talk with autophagy. Autophagy has been shown to be induced in various human cancer metastasis, modulating tumor cell motility and invasion, cancer cell differentiation, resistance to anoikis, and epithelial to mesenchymal transition. Hence, it may play a crucial role in the transition of benign to malignant phenotypes, the core of metastasis initiation. Here, we provide a method we have established in our laboratory for detecting of lipids in attached and detached non-small lung cancer cells and show how to analyze lipidomics data to find its correlation with autophagy-related pathways. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=95 SRC="FIGDIR/small/573842v2_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@1127909org.highwire.dtl.DTLVardef@1c86359org.highwire.dtl.DTLVardef@f29722org.highwire.dtl.DTLVardef@a05a7d_HPS_FORMAT_FIGEXP M_FIG C_FIG Workflow of A549 attached and detached cell culture. Cells are collected, lipids are extracted and subjected to LC/MS processing. Next, lipidomics resulting data is analyzed through various steps involving utilization of Metaboanlyst, HDMB, Swiss Target Prediction, and String.

cancer biology↗

bioRxiv2023

A proteogenomic pipeline for the analysis of protein biosynthesis errors in the human pathogen Candida albicans.

Candida albicans is a diploid pathogen known for its ability to live as a commensal fungus in healthy individuals, but causing both superficial infections and disseminated candidiasis in immunocompromised patients where it is associated with high morbidity and mortality. Its success in colonizing the human host is attributed to a wide range of virulence traits that modulate interactions between the host and the pathogen, such as optimal growth rate at 37{degrees}C, the ability to switch between yeast and hyphal forms and a remarkable genomic and phenotypic plasticity. A fascinating aspect of its biology is a prominent heterogeneous proteome that arises from frequent genomic rearrangements, high allelic variation, and high levels of amino acid misincorporations in proteins. The latter leads to increased morphological and physiological phenotypic diversity of high adaptive potential, but the scope of such protein mistranslation is poorly understood due to technical difficulties in detecting and quantifying amino acid misincorporation events in complex proteomic samples. To address this question, we have developed and optimized mass spectrometry and bioinformatics pipelines capable of identifying low-level amino acid misincorporation events at the proteome level. We have also analysed the proteomic profile of an engineered C. albicans strain that exhibits high level of leucine misincorporation at protein CUG sites and employed an in vivo quantitative gain-of-function fluorescence reporter system to validate our MS/MS data. The data show that C. albicans misincorporates amino acids above the background level at protein sites of diverse codons, particularly at CUG sites, confirming our previous data on the quantification of leucine incorporation at single CUG sites of recombinant reporter proteins. The study also demonstrates that increasing misincorporation of Leucine at CUG sites does not alter the translational fidelity of the other codons. These findings advance existing knowledge on amino acid misincorporations in C. albicans and add a new dimension to the remarkable capacity of this fungus to diversify its proteome.

microbiology↗

bioRxiv2023

Influenza A virus activates the unfolded protein response and induces the accumulation of insoluble protein aggregates that are essential for efficient viral propagation

Influenza A virus (IAV) is one of the main causes of annual respiratory epidemics in humans. IAV employs multiple strategies to evade host immunity and hijack cellular mechanisms to support proper virion formation and propagation. Some of these strategies encompass the manipulation of pathways involved in protein homeostasis, leading to changes in the host proteome and protein distribution within the cell. In this study, we performed a detailed analysis of the interplay between IAV and the host cells proteostasis mechanisms throughout the entire infectious cycle. We reveal that IAV infection induces the activation of the inositol requiring enzyme 1 (IRE1) branch of the unfolded protein response (UPR), at an infection stage that coincides with high rates of viral protein translation. This activation is particularly important for infection, as attenuation of virus production was observed upon IRE1 inhibition. Concomitantly to UPR activation, we observed the accumulation of virus-induced insoluble protein aggregates, which contain both viral and host proteins and are associated with a dysregulation of the host cell RNA metabolism. We demonstrate that this accumulation is important for IAV propagation, as its prevention using a quinoline-steroid hybrid compound significantly reduces the number of produced infectious virus particles. Our data suggests that the formation of these insoluble protein aggregates favors the final steps of the infection cycle, more specifically the virion assembly. Our findings reveal additional mechanisms by which IAV disrupts the host cell proteostasis to favor infection and uncover new cellular targets that can be explored for the development of host-directed antiviral strategies.

microbiology↗

bioRxiv2023

Prefrontal cortex proteomic alteration after social instability stress in adolescents rats

Early life stress can have significant effects on the developing brain and lead to changes in the structure and function of brain regions involved in stress regulation, emotion and cognitive control. Here, we used the social instability stress (SIS) protocol to understand the impact of social stress during mild (PND30) and late (PND45) adolescence. Our results revealed that SIS can compromise the dominance-subordination coping strategy but does not affect social recognition and motivation in rats. Moreover, SIS can lead to subtle modifications at the molecular level that hamper normal development of the prefrontal cortex in a sex- and age-dependent manner. Understanding the impact of early life stress on brain organization is crucial for developing effective prevention and intervention strategies. By identifying those who are most vulnerable to the effects of stress and providing targeted support and resources, it may be possible to mitigate the negative consequences of early adversity and promote healthy brain development.

neuroscience↗

bioRxiv2022

A multi-omics analysis of glioma chemoresistance using a hybrid microphysiological model of glioblastoma

Chemoresistance is a major clinical challenge in the management of glioblastoma (GBM) Temozolomide (TMZ) is the chemotherapeutic drug of choice for GBM; however, the therapeutic effect of TMZ is limited due to the development of resistance. Recapitulating GBM chemoresistance in a controlled environment is thus essential in understanding the mechanism of chemoresistance. Herein, we present a hybrid microphysiological model of chemoresistant GBM-on-a-chip (HGoC) by directly co-culturing TMZ-resistant GBM spheroids with healthy neurons to mimic the microenvironment of both the tumor and the surrounding healthy tissue. We characterized the model with proteomics, lipidomics, and secretome assays. The results showed that our artificial model recapitulated the molecular signatures of recurrent GBM in humans. Both showed alterations in vesicular transport and cholesterol pathways, mitotic quiescence, and a switch in metabolism to oxidative phosphorylation associated with a transition from mesenchymal to amoeboid. This is the first report to unravel the interplay of all these molecular changes as a mechanism of chemoresistance in glioblastoma. Moreover, we have shown that the acquisition of resistance increases invasiveness and the presence of neurons decreases this property.

bioengineering↗