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
Alteration in the cerebrospinal fluid lipidome in Parkinson’s disease: a post-mortem pilot study
(MDPI, 2021) Fernández Irigoyen, Joaquín; Cartas Cejudo, Paz; Iruarrizaga-Lejarreta, Marta; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak
Lipid metabolism is clearly associated to Parkinson’s disease (PD). Although lipid homeostasis has been widely studied in multiple animal and cellular models, as well as in blood derived from PD individuals, the cerebrospinal fluid (CSF) lipidomic profile in PD remains largely unexplored. In this study, we characterized the post-mortem CSF lipidomic imbalance between neurologically intact controls (n = 10) and PD subjects (n = 20). The combination of dual extraction with ultra-performance liquid chromatography-electrospray ionization quadrupole-time-of-flight mass spectrometry (UPLC-ESI-qToF-MS/MS) allowed for the monitoring of 257 lipid species across all samples. Complementary multivariate and univariate data analysis identified that glycerolipids (mono-, di-, and triacylglycerides), saturated and mono/polyunsaturated fatty acids, primary fatty amides, glycerophospholipids (phosphatidylcholines, phosphatidylethanolamines), sphingolipids (ceramides, sphingomyelins), N-acylethanolamines and sterol lipids (cholesteryl esters, steroids) were significantly increased in the CSF of PD compared to the control group. Interestingly, CSF lipid dyshomeostasis differed depending on neuropathological staging and disease duration. These results, despite the limitation of being obtained in a small population, suggest extensive CSF lipid remodeling in PD, shedding new light on the deployment of CSF lipidomics as a promising tool to identify potential lipid markers as well as discriminatory lipid species between PD and other atypical parkinsonisms.
Open Access
Network-driven proteogenomics unveils an aging-related imbalance in the olfactory IκBα-NFκB p65 complex functionality in Tg2576 Alzheimer’s disease mouse model
(MDPI, 2017) Palomino Alonso, Maialen; Lachén Montes, Mercedes; González Morales, Andrea; Ausín, Karina; Pérez Mediavilla, Alberto; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua, PC023-24; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
Olfaction is often deregulated in Alzheimer’s disease (AD) patients, and is also impaired in transgenic Tg2576 AD mice, which overexpress the Swedish mutated form of human amyloid precursor protein (APP). However, little is known about the molecular mechanisms that accompany the neurodegeneration of olfactory structures in aged Tg2576 mice. For that, we have applied proteome- and transcriptome-wide approaches to probe molecular disturbances in the olfactory bulb (OB) dissected from aged Tg2576 mice (18 months of age) as compared to those of age matched wild-type (WT) littermates. Some over-represented biological functions were directly relevant to neuronal homeostasis and processes of learning, cognition, and behavior. In addition to the modulation of CAMP responsive element binding protein 1 (CREB1) and APP interactomes, an imbalance in the functionality of the IκBα-NFκB p65 complex was observed during the aging process in the OB of Tg2576 mice. At two months of age, the phosphorylated isoforms of olfactory IκBα and NFκB p65 were inversely regulated in transgenic mice. However, both phosphorylated proteins were increased at 6 months of age, while a specific drop in IκBα levels was detected in 18-month-old Tg2576 mice, suggesting a transient activation of NFκB in the OB of Tg2576 mice. Taken together, our data provide a metabolic map of olfactory alterations in aged Tg2576 mice, reflecting the progressive effect of APP overproduction and β-amyloid (Aβ) accumulation on the OB homeostasis in aged stages.

Santamaría Martínez, Enrique

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