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• W613249969 abstract "catalaEls peixos carnivors presenten baixa capacitat per metabolitzar carbohidrats i per mantenir la glucemia. En aquests organismes, la baixa capacitat per metabolitzar carbohidrats condueix a estats d?hiperglicemia mes marcats i sostinguts que els descrits per a mamifers, despres de l?administracio de glucosa o la ingesta de dietes amb elevat contingut de carbohidrats. L?alanina aminotransferasa (ALT) catalitza la transaminacio reversible entre L-alanina i ?-cetoglutarat per formar L-glutamat i piruvat. Mitjancant la interconversio d?aquests quatre metabolits, l?ALT esdeve un nexe d?unio entre el metabolisme d?aminoacids i el de carbohidrats. En estudis previs del nostre grup es va descriure la presencia de tres isoformes ALT en S. aurata, els isoenzims citosolics, cALT1 i cALT2, i la isoforma mitocondrial mALT. L?expressio hepatica de cALT2 incrementa en situacions gluconeogeniques, mentre que la de cALT1 es predominant durant el periode postprandial per a l?utilitzacio dels nutrients de la dieta. L?objectiu d?aquest treball es incrementar el coneixement del metabolisme intermediari en peixos i aixi permetre realitzar futures intervencions biotecnologiques amb la intencio de millorar la utilitzacio metabolica dels nutrients de la dieta. Per comprendre millor la funcio d?mALT vam estudiar la distribucio tissular, la caracteritzacio cinetica i la regulacio nutricional i hormonal de l?expressio d?mALT en S. aurata. Addicionalment, vam analitzar l?expressio tissular i la regulacio hormonal dels enzims aspartat aminotransferasa mitocondrial (AST2), glutamat deshidrogenasa (GDH) i glutamina sintetasa (GlnS), relacionats tots ells amb el metabolisme d?aminoacids. En orades alimentades, mALT, GDH i GlnS s?expressen majoritariment a fetge, intesti i ronyo. Els nostres estudis indiquen que l?expressio de mALT, GDH, AST2 i GlnS es elevada en animals alimentats, mentre que disminueix en condicions associades amb la gluconegenesi, com ara el deju i el tractament amb estreptozotocina (STZ). Estudis cinetics de l?activitat enzimatica mALT indiquen que l?enzim catalitza de manera mes eficient la conversio d?L-alanina a piruvat, que no pas la reaccio inversa. Per coneixer el mecanisme molecular implicat en la regulacio transcripcional de l?expressio d?mALT, vam aillar el promotor mALT d?orada, i vam mostrar que HNF4? incrementa la transcipcio d?mALT per unio a una caixa HRE. El deju i l?administracio d?STZ disminuiren els nivells d?HNF4? en ronyo d?S. aurata, portant a un descens en la transcripcio d?mALT. Els nostres resultats suggereixen que HNF4? te un paper important en la regulacio transcripcional del gen mALT en ronyo d?S. aurata. En conclusio, els nostres resultats suggereixen que mALT i cALT1, que presenten una distribucio tissular i un patro d?expressio en deju i tractament amb STZ similars, podrien cooperar per direccionar els aminoacids de la dieta al mitocondri per destinar-los a l?obtencio d?energia. Per tant, ALT es podria utilitzar com a diana per realitzar una intervencio biotecnologica a fi de reduir la utilitzacio de proteines amb finalitats energetiques i optimitzar aixi l?us dels nutrients de la dieta en el cultiu de peixos. EnglishAbstract: Carnivorous fish have poor ability to use dietary carbohydrates and to control the blood glucose levels. Compared with mammals, these animals show prolonged hyperglycemia after a glucose load or when feeding on high carbohydrate diets. Alanine aminotransferase (ALT) links carbohydrate and amino acid metabolism through catalyzing the reversible transamination between L-alanine and 2-oxoglutarate to form pyruvate and L-glutamate. Our group, in previous studies showed the presence of three ALT isoforms in Sparus aurata: the cytosolic isoenzymes cALT1 and cALT2 and a mitochondrial isoform, mALT. In fish liver, increased expression of cALT2 is associated to enhanced gluconeogenesis while cALT1 is predominant during the postprandial utilization of dietary nutrients. The aim of the present study was to increase the current knowledge of fish intermediary metabolism to allow future biotechnological actions in order to improve metabolic utilization of dietary nutrients. To better understand the functional role of mALT we analysed the tissue distribution, kinetic characterization and nutritional and hormonal regulation of mALT expression in S. aurata. Furthermore, cloning and characterization of the mALT promoter was also addressed. Additionally, tissue expression and nutritional regulation of glutamate dehydrogenase (GDH), mitochondrial aspartate aminotransferase (AST2) and glutamine synthetase (GlnS), also involved in amino acid metabolism, was followed. In S. aurata under feeding conditions, mALT, GDH and GlnS are mainly expressed in liver, intestine and kidney. Our studies indicate that the expression of mALT, GDH, AST2 and GlnS is increased in fed animals, while decreased in conditions associated with gluconeogenesis, such as fasting or treatment with streptozotocin (STZ). Kinetic analysis of mALT enzyme activity indicated that this enzyme catalyses more efficiently the conversion of L-alanine to pyruvate than the reverse reaction. To understand the molecular mechanism underlying the transcriptional regulation of mALT expression, we isolated the S. aurata mALT promoter, and showed that HNF4? enhances mALT transcription through binding to an HRE box. Starvation and administration of STZ decreased HNF4? levels in the kidney of S. aurata, leading to downregulation of mALT transcription. Our results suggest that HNF4? may play an important role in the transcriptional regulation of mALT gene in kidney of S. aurata. In conclusion, our findings suggest that mALT and cALT1, which present a similar tissue distribution and pattern expression under starvation and STZ-treatment, can cooperate to redirect dietary amino acids to the mitochondria for energetic pourposes. This points to ALT as a target for a biotechnological action to spare protein and optimize the use of dietary nutrients for fish culture." @default.
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• W613249969 modified "2023-10-18" @default.
• W613249969 title "Control transcripcional i caracterització molecular de l’alanina aminotransferasa mitocondrial en l’orada (Sparus aurata)" @default.
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