Open Access
RTP801 interacts with the tRNA ligase complex and dysregulates its RNA ligase activity in Alzheimer's disease
(Oxford University Press, 2024-09-12) Campoy-Campos, Genís; Solana-Balaguer, Júlia; Guisado-Corcoll, Anna; Chicote-González, Almudena; García-Segura, Pol; Pérez-Sisqués, Leticia; Gabriel Torres, Adrián; Canal, Mercè; Molina-Porcel, Laura; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Pouplana, Lluís Ribas de; Alberch, Jordi; Martí, Eulàlia; Giralt, Albert; Pérez-Navarro, Esther; Malagelada, Cristina; Ciencias de la Salud; Osasun Zientziak
RTP801/REDD1 is a stress-responsive protein overexpressed in neurodegenerative diseases such as Alzheimer's disease (AD) that contributes to cognitive deficits and neuroinflammation. Here, we found that RTP801 interacts with HSPC117, DDX1 and CGI-99, three members of the tRNA ligase complex (tRNA-LC), which ligates the excised exons of intron-containing tRNAs and the mRNA exons of the transcription factor XBP1 during the unfolded protein response (UPR). We also found that RTP801 modulates the mRNA ligase activity of the complex in vitro since RTP801 knockdown promoted XBP1 splicing and the expression of its transcriptional target, SEC24D. Conversely, RTP801 overexpression inhibited the splicing of XBP1. Similarly, in human AD postmortem hippocampal samples, where RTP801 is upregulated, we found that XBP1 splicing was dramatically decreased. In the 5xFAD mouse model of AD, silencing RTP801 expression in hippocampal neurons promoted Xbp1 splicing and prevented the accumulation of intron-containing pre-tRNAs. Finally, the tRNA-enriched fraction obtained from 5xFAD mice promoted abnormal dendritic arborization in cultured hippocampal neurons, and RTP801 silencing in the source neurons prevented this phenotype. Altogether, these results show that elevated RTP801 impairs RNA processing in vitro and in vivo in the context of AD and suggest that RTP801 inhibition could be a promising therapeutic approach.
Open Access
Signature-driven repurposing of Midostaurin for combination with MEK1/2 and KRASG12C inhibitors in lung cancer
(Springer Nature, 2023) Macaya, Irati; Roman, Marta; Welch, Connor; Entrialgo-Cadierno, Rodrigo; Salmon, Marina; Santos, Alba; Feliu, Iker; Kovalski, Joanna; López Erdozain, Inés; Rodríguez-Remírez, María; Palomino Echeverría, Sara; Lonfgren, Shane M.; Ferrero, Macarena; Calabuig, Silvia; Ludwig, Iziar A.; Lara-Astiaso, David; Jantus-Lewintre, Eloisa; Guruceaga, Elizabeth; Narayanan, Shruthi; Ponz Sarvisé, Mariano; Pineda Lucena, Antonio; Lecanda, Fernando; Ruggero, Davide; Khatri, Purvesh; Santamaría Martínez, Enrique; Fernández Irigoyen, Joaquín; Ferrer, Irene; Paz-Ares, Luis; Drosten, Matthias; Barbacid, Mariano; Gil-Bazo, Ignacio; Vicent, Silvestre; Ciencias de la Salud; Osasun Zientziak
Drug combinations are key to circumvent resistance mechanisms compromising response to single anti-cancer targeted therapies. The implementation of combinatorial approaches involving MEK1/2 or KRASG12C inhibitors in the context of KRAS-mutated lung cancers focuses fundamentally on targeting KRAS proximal activators or effectors. However, the antitumor effect is highly determined by compensatory mechanisms arising in defined cell types or tumor subgroups. A potential strategy to find drug combinations targeting a larger fraction of KRAS-mutated lung cancers may capitalize on the common, distal gene expression output elicited by oncogenic KRAS. By integrating a signature-driven drug repurposing approach with a pairwise pharmacological screen, here we show synergistic drug combinations consisting of multi-tyrosine kinase PKC inhibitors together with MEK1/2 or KRASG12C inhibitors. Such combinations elicit a cytotoxic response in both in vitro and in vivo models, which in part involves inhibition of the PKC inhibitor target AURKB. Proteome profiling links dysregulation of MYC expression to the effect of both PKC inhibitor-based drug combinations. Furthermore, MYC overexpression appears as a resistance mechanism to MEK1/2 and KRASG12C inhibitors. Our study provides a rational framework for selecting drugs entering combinatorial strategies and unveils MEK1/2- and KRASG12C-based therapies for lung cancer.
Open Access
Amyotrophic lateral sclerosis is accompanied by protein derangements in the olfactory bulb-tract axis
(MDPI, 2020) Lachén Montes, Mercedes; Mendizuri, Naroa; Ausín, Karina; Andrés Benito, Pol; Ferrer, Isidro; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua, Ref. 0011-1411-2020-000028
Amyotrophic lateral sclerosis (ALS) is a fatal disease characterized by progressive muscle paralysis due to the degeneration of upper and lower motor neurons. Recent studies point out an involvement of the non-motor axis during disease progression. Despite smell impairment being considered a potential non-motor finding in ALS, the pathobiochemistry at the olfactory level remains unknown. Here, we applied an olfactory quantitative proteotyping approach to analyze the magnitude of the olfactory bulb (OB) proteostatic imbalance in ALS subjects (n = 12) with respect to controls (n = 8). Around 3% of the quantified OB proteome was differentially expressed, pinpointing aberrant protein expression involved in vesicle-mediated transport, macroautophagy, axon development and gliogenesis in ALS subjects. The overproduction of olfactory marker protein (OMP) points out an imbalance in the olfactory signal transduction in ALS. Accompanying the specific overexpression of glial fibrillary acidic protein (GFAP) and Bcl-xL in the olfactory tract (OT), a tangled disruption of signaling routes was evidenced across the OB–OT axis in ALS. In particular, the OB survival signaling dynamics clearly differ between ALS and frontotemporal lobar degeneration (FTLD), two faces of TDP-43 proteinopathy. To the best of our knowledge, this is the first report on high-throughput molecular characterization of the olfactory proteostasis in ALS.
Open Access
Influence of short-term training on functional capacity and (anti-)inflammatory immune signalling in acute hospitalization
(Wiley, 2020) Ramírez Vélez, Robinson; Martínez Velilla, Nicolás; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Izquierdo Redín, Mikel; Palomino Echeverría, Sara; Ciencias de la Salud; Osasun Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa; Gobierno de Navarra / Nafarroako Gobernua
To investigate the infuence of exercise on inflammatory signalling, it was performed cytokine array profiling in human serum to identify inflammatory cytokines produced after a 3 day in-hospital intervention including individualized moderate-intensity resistance, balance, and walking exercises vs. medical usual-care for acute hospitalization in very elderly patients.
Open Access
Docosahexaenoic acid ameliorates contextual fear memory deficits in the Tg2576 Alzheimer´s disease mouse model: cellular and molecular correlates
(MDPI, 2023) Badesso, Sara; Cartas Cejudo, Paz; Espelosín, María; Santamaría Martínez, Enrique; Cuadrado-Tejedor, Mar; García-Osta, Ana; Ciencias de la Salud; Osasun Zientziak
Docosahexaenoic acid (DHA), the most abundant polyunsaturated fatty acid in the brain, is essential for successful aging. In fact, epidemiological studies have demonstrated that increased intake of DHA might lower the risk for developing Alzheimer’s disease (AD). These observations are supported by studies in animal models showing that DHA reduces synaptic pathology and memory deficits. Different mechanisms to explain these beneficial effects have been proposed; however, the molecular pathways involved are still unknown. In this study, to unravel the main underlying molecular mechanisms activated upon DHA treatment, the effect of a high dose of DHA on cognitive function and AD pathology was analyzed in aged Tg2576 mice and their wild-type littermates. Transcriptomic analysis of mice hippocampi using RNA sequencing was subsequently performed. Our results revealed that, through an amyloid-independent mechanism, DHA enhanced memory function and increased synapse formation only in the Tg2576 mice. Likewise, the IPA analysis demonstrated that essential neuronal functions related to synaptogenesis, neuritogenesis, the branching of neurites, the density of dendritic spines and the outgrowth of axons were upregulated upon-DHA treatment in Tg2576 mice. Our results suggest that memory function in APP mice is influenced by DHA intake; therefore, a high dose of daily DHA should be tested as a dietary supplement for AD dementia prevention.
Open Access
Metschnikowia pulcherrima as an efficient biocontrol agent of Botrytis cinerea infection in apples: unraveling protection mechanisms through yeast proteomics
(Elsevier, 2023) Fernández San Millán, Alicia; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Larraya Reta, Luis María; Farrán Blanch, Inmaculada; Veramendi Charola, Jon; Ciencias de la Salud; Osasun Zientziak; Institute for Multidisciplinary Research in Applied Biology - IMAB; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
The results obtained in this study show that the Mp-30 strain of Metschnikowia pulcherrima is able to completely prevent Botrytis cinerea infection in apples, which is a major postharvest disease of fruits throughout the world. We have observed that although Mp-30 is able to rapidly colonize wounds, sequestrate iron and secrete antifungal compounds, other unknown mechanisms that occur in the early phase of the yeast-fungal interaction must be implicated in the biocontrol response. The main objective of this study was to identify the pathways involved in the mechanism of action of Mp-30 against B. cinerea in apples. Therefore, differentially accumulated yeast proteins in the presence/absence of B. cinerea on wounded apples were studied to elucidate Mp-30 biocontrol mechanisms and regulation at the protein level. A comparative proteomic analysis showed that 114 yeast proteins were increased and 61 were decreased. The Mp-30 antagonistic response mainly showed the increase of (1) gene expression and protein translation related proteins, (2) trafficking and vesicle-mediated transport related proteins, (3) pyruvate metabolism and mitochondrial proteins related to energy and amino acid production, (4) fatty acid synthesis, and (5) cell envelope related proteins. On the other hand, redox homeostasis, and amino acid and carbon metabolism were downregulated. Since there is no yeast growth enhancement associated with the presence of B. cinerea, such regulation mechanisms may be related to the reprogramming of metabolism, synthesis of new compounds and reorganization of yeast cell structure. Indeed, the results show that several pathways cooperate in restructuring the plasma membrane and cell wall composition, highlighting their major role in the antagonistic interactions for apple protection against gray mold proliferation. These results are of great interest since they provide a clear insight into the yeast mechanisms involved in B. cinerea inactivation during the first hours of contact in the wounded fruit. They shed light on the unknown yeast molecular biocontrol mechanisms.
Open Access
Motor skill learning modulates striatal extracellular vesicles' content in a mouse model of Huntington's disease
(BMC, 2024-06-11) Solana-Balaguer, Júlia; García-Segura, Pol; Campoy-Campos, Genís; Chicote-González, Almudena; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Pérez-Navarro, Esther; Masana, Mercè; Alberch, Jordi; Malagelada, Cristina; Ciencias de la Salud; Osasun Zientziak
Huntington's disease (HD) is a neurological disorder caused by a CAG expansion in the Huntingtin gene (HTT). HD pathology mostly affects striatal medium-sized spiny neurons and results in an altered cortico-striatal function. Recent studies report that motor skill learning, and cortico-striatal stimulation attenuate the neuropathology in HD, resulting in an amelioration of some motor and cognitive functions. During physical training, extracellular vesicles (EVs) are released in many tissues, including the brain, as a potential means for inter-tissue communication. To investigate how motor skill learning, involving acute physical training, modulates EVs crosstalk between cells in the striatum, we trained wild-type (WT) and R6/1 mice, the latter with motor and cognitive deficits, on the accelerating rotarod test, and we isolated their striatal EVs. EVs from R6/1 mice presented alterations in the small exosome population when compared to WT. Proteomic analyses revealed that striatal R6/1 EVs recapitulated signaling and energy deficiencies present in HD. Motor skill learning in R6/1 mice restored the amount of EVs and their protein content in comparison to naïve R6/1 mice. Furthermore, motor skill learning modulated crucial pathways in metabolism and neurodegeneration. All these data provide new insights into the pathogenesis of HD and put striatal EVs in the spotlight to understand the signaling and metabolic alterations in neurodegenerative diseases. Moreover, our results suggest that motor learning is a crucial modulator of cell-to-cell communication in the striatum.
Open Access
Sex-specific role of galectin-3 in aortic stenosis
(BMC, 2023) Matilla Cuenca, Lara; Martín Núñez, Ernesto; Garaikoetxea Zubillaga, Mattie; Navarro, Adela; Tamayo Rodríguez, Ibai; Fernández Celis, Amaya; Gaínza Calleja, Alicia; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Muntendam, Pieter; Álvarez, Virginia; Sádaba Sagredo, Rafael; Jover, Eva; López Andrés, Natalia; Ciencias de la Salud; Osasun Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
Background: Aortic stenosis (AS) is characterized by infammation, fbrosis, osteogenesis and angiogenesis. Men and women develop these mechanisms diferently. Galectin-3 (Gal-3) is a pro-infammatory and pro-osteogenic lectin in AS. In this work, we aim to analyse a potential sex-diferential role of Gal-3 in AS. Methods: 226 patients (61.50% men) with severe AS undergoing surgical aortic valve (AV) replacement were recruited. In AVs, Gal-3 expression and its relationship with infammatory, osteogenic and angiogenic markers was assessed. Valve interstitial cells (VICs) were primary cultured to perform in vitro experiments. Results: Proteomic analysis revealed that intracellular Gal-3 was over-expressed in VICs of male AS patients. Gal-3 secretion was also higher in men’s VICs as compared to women’s. In human AVs, Gal-3 protein levels were signifcantly higher in men, with stronger immunostaining in VICs with myofbroblastic phenotype and valve endothelial cells. Gal-3 levels in AVs were positively correlated with infammatory markers in both sexes. Gal-3 expression was also posi tively correlated with osteogenic markers mainly in men AVs, and with angiogenic molecules only in this sex. In vitro, Gal-3 treatment induced expression of infammatory, osteogenic and angiogenic markers in male’s VICs, while it only upregulated infammatory and osteogenic molecules in women-derived cells. Gal-3 blockade with pharma cological inhibitors (modifed citrus pectin and G3P-01) prevented the upregulation of infammatory, osteogenic and angiogenic molecules. Conclusions: Gal-3 plays a sex-diferential role in the setting of AS, and it could be a new sex-specifc therapeutic target controlling pathological features of AS in VICs.
Open Access
Multi-laboratory experiment PME11 for the standardization of phosphoproteome analysis
(Elsevier, 2022) Colomé, Núria; Abian, Joaquín; Aloria, Kerman; Arizmendi, Jesús M.; Barceló-Batllori, Silvia; Braga-Lagache, Sophie; Burlet-Schiltz, Odile; Carrascal, Montse; Casal, Ignacio J.; Chicano-Gálvez, Eduard; Chiva, Cristina; Clemente, Luis F.; Elortza, Félix; Estanyol, Josep M.; Fernández Irigoyen, Joaquín; Fernández-Puente, Patricia; Fidalgo, María J.; Froment, Carine; Fuentes, Manuel; Fuentes-Almagro, Carlos; Gay, Marina; Hainard, Alexandre; Heller, Manfred; Hernández, María Luisa; Ibarrola, Nieves; Iloro, Ibon; Kieselbach, Thomas; Lario, Antonio; Locard-Paulet, Marie; Marina-Ramírez, Anabel; Martín, Luna; Morato-López, Esperanza; Muñoz, Javier; Navajas, Rosana; Odena, Antonia M.; Odriozola, Leticia; Oliveira, Eliandre de; Paradela, Alberto; Pasquarello, Carla; Rios, Vivian de los; Ruiz-Romero, Cristina; Sabidó, Eduard; Sánchez del Pino, Manuel; Sancho, Jaime; Santamaría Martínez, Enrique; Schaeffer-Reiss, Christine; Schneider, Justine; Torre, Carolina de la; Valero, Luz M.; Vilaseca, Marta; Wu, Shuai; Wu, Linfeng; Ximénez de Embún, Pilar; Canals, Francesc; Corrales, Fernando J.; ProteoRed-ISCIII; EuPA; Ciencias de la Salud; Osasun Zientziak
Global analysis of protein phosphorylation by mass spectrometry proteomic techniques has emerged in the last decades as a powerful tool in biological and biomedical research. However, there are several factors that make the global study of the phosphoproteome more challenging than measuring non-modified proteins. The low stoichiometry of the phosphorylated species and the need to retrieve residue specific information require particular attention on sample preparation, data acquisition and processing to ensure reproducibility, qualitative and quantitative robustness and ample phosphoproteome coverage in phosphoproteomic workflows. Aiming to investigate the effect of different variables in the performance of proteome wide phosphoprotein analysis protocols, ProteoRed-ISCIII and EuPA launched the Proteomics Multicentric Experiment 11 (PME11). A reference sample consisting of a yeast protein extract spiked in with different amounts of a phosphomix standard (Sigma/Merck) was distributed to 31 laboratories around the globe. Thirty-six datasets from 23 laboratories were analyzed. Our results indicate the suitability of the PME11 reference sample to benchmark and optimize phosphoproteomics strategies, weighing the influence of different factors, as well as to rank intra and inter laboratory performance.
Open Access
Improvement of cognitive function in wild-type and Alzheimer's disease mouse models by the immunomodulatory properties of menthol inhalation or by depletion of T regulatory cells
(Frontiers Media, 2023) Casares, Noelia; Alfaro Larraya, María; Cuadrado-Tejedor, Mar; Lasarte-Cía, Aritz; Navarro Negredo, Flor; Vivas, Isabel; Espelosín, María; Cartas Cejudo, Paz; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; García-Osta, Ana; Lasarte, Juan José; Ciencias de la Salud; Osasun Zientziak
A complex network of interactions exists between the olfactory, immune and central nervous systems. In this work we intend to investigate this connection through the use of an immunostimulatory odorant like menthol, analyzing its impact on the immune system and the cognitive capacity in healthy and Alzheimer’s Disease Mouse Models. We first found that repeated short exposures to menthol odor enhanced the immune response against ovalbumin immunization. Menthol inhalation also improved the cognitive capacity of immunocompetent mice but not in immunodeficient NSG mice, which exhibited very poor fear-conditioning. This improvement was associated with a downregulation of IL-1β and IL-6 mRNA in the brain´s prefrontal cortex, and it was impaired by anosmia induction with methimazole. Exposure to menthol for 6 months (1 week per month) prevented the cognitive impairment observed in the APP/PS1 mouse model of Alzheimer. Besides, this improvement was also observed by the depletion or inhibition of T regulatory cells. Treg depletion also improved the cognitive capacity of the APPNL-G-F/NL-G-F Alzheimer´s mouse model. In all cases, the improvement in learning capacity was associated with a downregulation of IL-1β mRNA. Blockade of the IL-1 receptor with anakinra resulted in a significant increase in cognitive capacity in healthy mice as well as in the APP/PS1 model of Alzheimer´s disease. These data suggest an association between the immunomodulatory capacity of smells and their impact on the cognitive functions of the animals, highlighting the potential of odors and immune modulators as therapeutic agents for CNS-related diseases.