Proteins in human brain cortex are modified by oxidation, glycoxidation, and lipoxidation. Effects of alzheimer disease and identification of lipoxidation targets

Diverse oxidative pathways, such as direct oxidation of amino acids, glycoxidation, and lipoxidation could contribute to Alzheimer disease pathogenesis. A global survey for the amount of structurally characterized probes for these reactions is lacking and could overcome the lack of specificity derived from measurement of 2,4-dinitrophenylhydrazine reactive carbonyls. Consequently we analyzed (i) the presence and concentrations of glutamic and aminoadipic semialdehydes, N(epsilon)-(carboxymethyl)-lysine, N(epsilon)-(carboxyethyl)-lysine, and N(epsilon)-(malondialdehyde)-lysine by means of gas chromatography/mass spectrometry, (ii) the biological response through expression of the receptor for advanced glycation end products, (iii) the fatty acid composition in brain samples from Alzheimer disease patients and age-matched controls, and (iv) the targets of N(epsilon)-(malondialdehyde)-lysine formation in brain cortex by proteomic techniques. Alzheimer disease was associated with significant, although heterogeneous, increases in the concentrations of all evaluated markers. Alzheimer disease samples presented increases in expression of the receptor for advanced glycation end products with high molecular heterogeneity. Samples from Alzheimer disease patients also showed content of docosahexaenoic acid, which increased lipid peroxidizability. In accordance, N(epsilon)-(malondialdehyde)-lysine formation targeted important proteins for both glial and neuronal homeostasis such as neurofilament L, alpha-tubulin, glial fibrillary acidic protein, ubiquinol-cytochrome c reductase complex protein I, and the beta chain of ATP synthase. These data support an important role for lipid peroxidation-derived protein modifications in Alzheimer disease pathogenesis

Whole exome sequencing of Rett syndrome-like patients reveals the mutational diversity of the clinical phenotype

Classical Rett syndrome (RTT) is a neurodevelopmental disorder where most of cases carry MECP2 mutations. Atypical RTT variants involve mutations in CDKL5 and FOXG1. However, a subset of RTT patients remains that do not carry any mutation in the described genes. Whole exome sequencing was carried out in a cohort of 21 female probands with clinical features overlapping with those of RTT, but without mutations in the customarily studied genes. Candidates were functionally validated by assessing the appearance of a neurological phenotype in Caenorhabditis elegans upon disruption of the corresponding ortholog gene. We detected pathogenic variants that accounted for the RTT-like phenotype in 14 (66.6 %) patients. Five patients were carriers of mutations in genes already known to be associated with other syndromic neurodevelopmental disorders. We determined that the other patients harbored mutations in genes that have not previously been linked to RTT or other neurodevelopmental syndromes, such as the ankyrin repeat containing protein ANKRD31 or the neuronal acetylcholine receptor subunit alpha-5 (CHRNA5). Furthermore, worm assays demonstrated that mutations in the studied candidate genes caused locomotion defects. Our findings indicate that mutations in a variety of genes contribute to the development of RTT-like phenotypes

PDR-1/hParkin negatively regulates the phagocyosis of apoptotic cell corpses in Caenorhabditis elegans

Apoptotic cell death is an integral part of cell turnover in many tissues, and proper corpse clearance is vital to maintaining tissue homeostasis in all multicellular organisms. Even in tissues with high cellular turnover, apoptotic cells are rarely seen because of efficient clearance mechanisms in healthy individuals. In Caenorhabditis elegans, two parallel and partly redundant conserved pathways act in cell corpse engulfment. The pathway for cytoskeletal rearrangement requires the small GTPase CED-10 Rac1 acting for an efficient surround of the dead cell. The CED-10 Rac pathway is also required for the proper migration of the distal tip cells (DTCs) during the development of the C. elegans gonad. Parkin, the mammalian homolog of the C. elegans PDR-1, interacts with Rac1 in aged human brain and it is also implicated with actin dynamics and cytoskeletal rearrangements in Parkinsons's disease, suggesting that it might act on engulfment. Our genetic and biochemical studies indicate that PDR-1 inhibits apoptotic cell engulfment and DTC migration by ubiquitylating CED-10 for degradation

Estudi de les sinucleïnopaties: L'alfa-sinucleïna a la sinapsi i efectes de l'estrès oxidatiu.

[cat] L'objectiu general d'aquest treball és el d'aprofundir en l'estudi de les sinucleïnopaties, principalment a nivell bioquímic, en les interaccions de l'alfa-sinucleïna amb altres proteïnes del cervell humà per tal d'estudiar l'efecte de la unió en aquestes patologies. També hem volgut comprovar la similitud de les sinucleïnopaties humanes amb un model animal transgènic per a la forma humana mutada de l'alfa-sinucleïna. D'altra banda i també relacionat amb aquestes neurodegenarions, s'ha estudiat de quina manera resultaven afectats diferents marcadors de l'estrès oxidatiu, que se sap juga un important paper en el desenvolupament d'aquestes patologies. Mitjançant estudis d'immunoprecipitació i de pull-down en cervell humà s'ha trobat interacció de l'alfa-sinucleïna amb les proteïnes sinàptiques rab3a i rabfilina a córtex de la sinucleïnopatia anomenada Demència amb cossos de Lewy (DLBD), i no en cervells control. Donada la interacció anòmala amb aquestes proteïnes sinàptiques podem afirmar que a DLBDs la transmissió sinàptica es troba alterada, i una de les causes és la interacció anòmala de l'alfa-sinucleïna amb rab3a i rabfilina. Aquest tipus d'interacció també s'ha estudiat a l'MSA o Atròfia Sistèmica Múltiple, una sinucleïnopatia que té afectat sobretot el cerebel, i no el córtex, com les DLBDs. Doncs, tot i estar classificada com a sinucleïnopatia a part, hem observat la mateixa interacció anòmala tan a cerebel com escorça de pacients d'MSA, suggerint-se un mecanisme comú de neurodegeneració per a totes les sinucleïnopaties. Paral.lelament també hem estudiat la via de transducció glutamatèrgica a pacients amb DLBDs, i s'ha trobat que en condicions normals exiteix interacció de l'alfa-sinucleïna amb un efector del receptors metabotròpics del glutmat tipus I, com és la PLC-beta-1. Al mateix temps, i a través d'estudis de solubilitat, s'han observat canvis en la solubilitat de PLC-beta-1 en aquestes patologies, tal com succeeix amb l'alfa-sinucleïna. Quan s'han estudiat marcadors d'estrès oxidatiu per una forma primerenca de la malaltia de Parkinson s'han trobat alterats a córtex d'aquest pacients, quan la patología, en els primers estadis només es detecta fins ara a substància negra. Per tant, l'estrès oxidatiu a córtex d'una forma pre-clínica de la malaltia de Parkinson, ja té un paper en el desenvolupament d'estadis posterior de la malaltia, com podrien ser les DLBDs.[eng] The aim of this work is to study alpha-synuclein interactions in human brain to deep on the knowledge of a group of neurodegenerative diseases called synucleinopathies in which alpha-synuclein accumulates in the form of Lewy bodies. In human cortex we found alpha-synuclein interactions with the synaptic proteins rab3a and rabphilin in Dementia with Lewy Bodies (DLBDs) patients, and not in control brains. This interaction is corroborated in transgenic mice expressing human A30P human mutated alpha-synuclein. When a different synucleipathy , like Multiple Systemic Atrophy (MSA), is studied we found the same anomalous interaction even different areas were affected, cerebellum vs cortex in the case of MSA and DLBDs, indicating a common neurodegenerative mechanism for the synucleinopathies group. Also in DLBDs we studied the glutamatergic signal transduction pathway immunoprecipitating alpha-synuclein from brain cortex and we found that PLC-beta-1, an effector of the metabotropic glutamate receptors (mGLuR1) binds to alpha-synuclein only in control conditions and not in DLBDs. At the same time PLC-beta-1 showed changes in solubility like alpha-synuclein in DLBDs. So, the results presented here indicate a kind of relation between alpha-synuclein and PLC-beta-1 in DLBDs which damage can affect the mGLuR1 transduction pathway which is altered in cortex from DLBDs. The other factor studied in this work is the oxidative stress effect in brain cortex from early stages Parkinson's disease (PD) patients. We found an increase of the oxidative stress markers used in this work indicating that oxidative stress in cortex from PD patients starts at early stages in human cortex, even though in early stages only the substantia nigra seems to be affected. This markers of oxidative stress were also studied in Alzheimer's disease (AD) obtaining similar results. Finally another point of this work is the study of the Amyloid Precursor Protein (A-beta-PP) mRNA isoforms in human cortex from DLBDs. We found a relative increase of the isoforms contained the Kunitz Protease Inhibitor, implicating A-beta-PP the isoforms processing in the developing of DLBD

Proteins in human brain cortex are modified by oxidation, glycoxidation, and lipoxidation. Effects of alzheimer disease and identification of lipoxidation targets

Diverse oxidative pathways, such as direct oxidation of amino acids, glycoxidation, and lipoxidation could contribute to Alzheimer disease pathogenesis. A global survey for the amount of structurally characterized probes for these reactions is lacking and could overcome the lack of specificity derived from measurement of 2,4-dinitrophenylhydrazine reactive carbonyls. Consequently we analyzed (i) the presence and concentrations of glutamic and aminoadipic semialdehydes, N(epsilon)-(carboxymethyl)-lysine, N(epsilon)-(carboxyethyl)-lysine, and N(epsilon)-(malondialdehyde)-lysine by means of gas chromatography/mass spectrometry, (ii) the biological response through expression of the receptor for advanced glycation end products, (iii) the fatty acid composition in brain samples from Alzheimer disease patients and age-matched controls, and (iv) the targets of N(epsilon)-(malondialdehyde)-lysine formation in brain cortex by proteomic techniques. Alzheimer disease was associated with significant, although heterogeneous, increases in the concentrations of all evaluated markers. Alzheimer disease samples presented increases in expression of the receptor for advanced glycation end products with high molecular heterogeneity. Samples from Alzheimer disease patients also showed content of docosahexaenoic acid, which increased lipid peroxidizability. In accordance, N(epsilon)-(malondialdehyde)-lysine formation targeted important proteins for both glial and neuronal homeostasis such as neurofilament L, alpha-tubulin, glial fibrillary acidic protein, ubiquinol-cytochrome c reductase complex protein I, and the beta chain of ATP synthase. These data support an important role for lipid peroxidation-derived protein modifications in Alzheimer disease pathogenesis

PDR-1/hParkin negatively regulates the phagocyosis of apoptotic cell corpses in Caenorhabditis elegans

Apoptotic cell death is an integral part of cell turnover in many tissues, and proper corpse clearance is vital to maintaining tissue homeostasis in all multicellular organisms. Even in tissues with high cellular turnover, apoptotic cells are rarely seen because of efficient clearance mechanisms in healthy individuals. In Caenorhabditis elegans, two parallel and partly redundant conserved pathways act in cell corpse engulfment. The pathway for cytoskeletal rearrangement requires the small GTPase CED-10 Rac1 acting for an efficient surround of the dead cell. The CED-10 Rac pathway is also required for the proper migration of the distal tip cells (DTCs) during the development of the C. elegans gonad. Parkin, the mammalian homolog of the C. elegans PDR-1, interacts with Rac1 in aged human brain and it is also implicated with actin dynamics and cytoskeletal rearrangements in Parkinsons's disease, suggesting that it might act on engulfment. Our genetic and biochemical studies indicate that PDR-1 inhibits apoptotic cell engulfment and DTC migration by ubiquitylating CED-10 for degradation

PDR-1/hParkin negatively regulates the phagocyosis of apoptotic cell corpses in Caenorhabditis elegans

Apoptotic cell death is an integral part of cell turnover in many tissues, and proper corpse clearance is vital to maintaining tissue homeostasis in all multicellular organisms. Even in tissues with high cellular turnover, apoptotic cells are rarely seen because of efficient clearance mechanisms in healthy individuals. In Caenorhabditis elegans, two parallel and partly redundant conserved pathways act in cell corpse engulfment. The pathway for cytoskeletal rearrangement requires the small GTPase CED-10 Rac1 acting for an efficient surround of the dead cell. The CED-10 Rac pathway is also required for the proper migration of the distal tip cells (DTCs) during the development of the C. elegans gonad. Parkin, the mammalian homolog of the C. elegans PDR-1, interacts with Rac1 in aged human brain and it is also implicated with actin dynamics and cytoskeletal rearrangements in Parkinsons's disease, suggesting that it might act on engulfment. Our genetic and biochemical studies indicate that PDR-1 inhibits apoptotic cell engulfment and DTC migration by ubiquitylating CED-10 for degradation