Peixos transgènics

La generació d'organismes modificats genèticament (OMG) ha representat un avanç tecnològic molt important en totes les àrees de recerca biològica. Així, la producció d'animals transgènics ha estat una eina molt valuosa per a l'establiment d'organismes model en recerca biomèdica, la producció massiva de macromolècules biològicament actives i el desenvolupament de cultius d'aliments, entre d'altres. Malgrat que tradicionalment els mamífers han estat els vertebrats més emprats per a la generació d'animals transgènics, els peixos constitueixen una alternativa excel·lent. Comparats amb els mamífers, els peixos presenten tot un conjunt de característiques, tant evolutives com fisiològiques, i també una sèrie d'interessos comercials i tecnològics, que estan fent d'aquests organismes els animals amb més potencial en els diferents camps d'aplicació de la tecnologia de manipulació genètica. En aquesta revisió, a més de presentar els darrers avenços tecnològics disponibles per a la generació de peixos transgènics, es repassen les diferents aplicacions de la transferència gènica en peixos, i també els beneficis i els possibles riscos derivats d'aquest ús. Paraules clau: peix, transgènesi, vector, tècniques de transferència gènica, aqüicultura,The generation of genetically modified organisms (GMOs) has provided an important technological advance to many areas of biological research. The production of these GMOs has been a valuable tool for the establishment of model organisms for biomedical research and the commercial production of biologically active macromolecules and food, amongst others. Traditionally, mammals have been used as the model organisms however in recent years the production of transgenic fish has provided an excellent alternative. Compared with mammals, fish presents some ideal physiological characteristics and evolutionary features, which together with recent technological advances and commercial interests, make fish an ideal candidate for genetic manipulation. This paper provides a brief summary of the most recent relevant data in this field

Evolution shapes the responsiveness of the D-box enhancer element to light and reactive oxygen species in vertebrates

The circadian clock is a highly conserved cell-autonomous mechanism that directs daily rhythms in most aspects of biology. Daily entrainment by environmental signals, notably light, is essential for its function. However, our understanding of the mechanisms and the evolution of photic entrainment remains incomplete. Fish represent attractive models for exploring how light regulates the circadian clock due to the direct light sensitivity of their peripheral clocks. Central to this property is the light induced expression of clock genes that is mediated by D-box enhancer elements. Here, using zebrafish cells, we reveal that the light responsive D-box enhancer serves as a nuclear target for reactive oxygen species (ROS). We demonstrate that exposure to short wavelengths of visible light triggers increases in ROS levels via NADPH oxidase activity. Elevated ROS activates the JNK and p38 MAP kinases and in turn, induces clock gene expression via the D-box. In blind cavefish and mammals, where peripheral clocks are no longer entrained by direct illumination, ROS levels are still increased upon light exposure. However, in these species ROS no longer induces D-box driven clock gene transcription. Thus, during evolution, alterations in ROS-responsive signal transduction pathways underlie fundamental changes in peripheral clock photoentrainment.Universidad de Ferrara | Ref. FAR2014–201

Mutations in blind cavefish target the light-regulated circadian clock gene, period 2

Light represents the principal signal driving circadian clock entrainment. However, how light influences the evolution of the clock remains poorly understood. The cavefish Phreatichthys andruzzii represents a fascinating model to explore how evolution under extreme aphotic conditions shapes the circadian clock, since in this species the clock is unresponsive to light. We have previously demonstrated that loss-of-function mutations targeting non-visual opsins contribute in part to this blind clock phenotype. Here, we have compared orthologs of two core clock genes that play a key role in photic entrainment, cry1a and per2, in both zebrafish and P. andruzzii. We encountered aberrantly spliced variants for the P. andruzzii per2 transcript. The most abundant transcript encodes a truncated protein lacking the C-terminal Cry binding domain and incorporating an intronic, transposon-derived coding sequence. We demonstrate that the transposon insertion leads to a predominantly cytoplasmic localization of the cavefish Per2 protein in contrast to the zebrafish ortholog which is distributed in both the nucleus and cytoplasm. Thus, it seems that during evolution in complete darkness, the photic entrainment pathway of the circadian clock has been subject to mutation at multiple levels, extending from opsin photoreceptors to nuclear effectors.Universidad de Ferrara | Ref. FAR2014-201

Unraveling the periprandial changes in brain serotonergic activity and its correlation with food intake-related neuropeptides in rainbow trout Oncorhynchus mykiss

This study explored changes in brain serotonin content and activity together with hypothalamic neuropeptide mRNA abundance around feeding time in rainbow trout, as well as the effect of one-day fasting. Groups of trout fed at two (ZT2) and six (ZT6) hours after lights on were sampled from 90 minutes before to 240 minutes after feeding, while additional groups of non-fed trout were also included in the study. Changes in brain amine and metabolite contents were measured in hindbrain, diencephalon and telencephalon, while in the diencephalon the mRNA abundance of tryptophan hydroxylase ( tph1 , tph2 ), serotonin receptors (5htr1a , 5htr1b and 5htr2c ) and several neuropeptides ( npy , agrp1 , cartpt , pomca1 , crfb ) involved in the control of food intake were also assessed. The results showed changes in the hypothalamic neuropeptides that were consistent with the expected role for each in the regulation of food intake in rainbow trout. Serotonergic activity increased rapidly at the time of food intake in the diencephalon and hindbrain and remained high for much of the postprandial period. This increase in serotonin abundance was concomitant with elevated levels of pomca1 mRNA in the diencephalon, suggesting that serotonin might act on brain neuropeptides to promote a satiety profile. Furthermore, serotonin synthesis and neuronal activity appear to increase already before the time of feeding, suggesting additional functions for this amine before and during food intake. Exploration of serotonin receptors in the diencephalon revealed only small changes for gene expression of 5htr1b and 5htr2c receptors during the postprandial phase. Therefore, the results suggest that serotonin may play a relevant role in the regulation of feeding behavior in rainbow trout during periprandial time, but a better understanding of its interaction with brain centers involved in receiving and processing food-related signals is still needed.Agencia Estatal de Investigación | Ref. PID2022-136288OB-C31Xunta de Galicia | Ref. ED431B 2019/37Agencia Estatal de Investigación | Ref. BES-2017-079708Xunta de Galicia | Ref. ED481B-2022-08

Peixos transgènics

La generació d'organismes modificats genèticament (OMG) ha representat un avanç tecnològic molt important en totes les àrees de recerca biològica. Així, la producció d'animals transgènics ha estat una eina molt valuosa per a l'establiment d'organismes model en recerca biomèdica, la producció massiva de macromolècules biològicament actives i el desenvolupament de cultius d'aliments, entre d'altres. Malgrat que tradicionalment els mamífers han estat els vertebrats més emprats per a la generació d'animals transgènics, els peixos constitueixen una alternativa excel·lent. Comparats amb els mamífers, els peixos presenten tot un conjunt de característiques, tant evolutives com fisiològiques, i també una sèrie d'interessos comercials i tecnològics, que estan fent d'aquests organismes els animals amb més potencial en els diferents camps d'aplicació de la tecnologia de manipulació genètica. En aquesta revisió, a més de presentar els darrers avenços tecnològics disponibles per a la generació de peixos transgènics, es repassen les diferents aplicacions de la transferència gènica en peixos, i també els beneficis i els possibles riscos derivats d'aquest ús. Paraules clau: peix, transgènesi, vector, tècniques de transferència gènica, aqüicultura,The generation of genetically modified organisms (GMOs) has provided an important technological advance to many areas of biological research. The production of these GMOs has been a valuable tool for the establishment of model organisms for biomedical research and the commercial production of biologically active macromolecules and food, amongst others. Traditionally, mammals have been used as the model organisms however in recent years the production of transgenic fish has provided an excellent alternative. Compared with mammals, fish presents some ideal physiological characteristics and evolutionary features, which together with recent technological advances and commercial interests, make fish an ideal candidate for genetic manipulation. This paper provides a brief summary of the most recent relevant data in this field

Differential circadian and light-driven rhythmicity of clock gene expression and behaviour in the turbot, Scophthalmus maximus

In fish, the circadian clock represents a key regulator of many aspects of biology and is controlled by combinations of abiotic and biotic factors. These environmental factors are frequently manipulated in fish farms as part of strategies designed to maximize productivity. The flatfish turbot, Scophthalmus maximus, represents one of the most important species within the aquaculture sector in Asia and Europe. Despite the strategic importance of this species, the function and regulation of the turbot circadian system remains poorly understood. Here, we have characterized the core circadian clock genes, clock1, per1, per2 and cry1 in turbot and have studied their daily expression in various tissues under a range of lighting conditions and feeding regimes. We have also explored the influence of light and feeding time on locomotor activity. Rhythmic expression of the four core clock genes was observed in all tissues studied under light dark (LD) cycle conditions. Rhythmicity of clock gene expression persisted upon transfer to artificial free running, constant conditions confirming their endogenous circadian clock control. Furthermore, turbot showed daily cycles of locomotor activity and food anticipatory activity (FAA) under LD and scheduled-feeding, with the activity phase as well as FAA coinciding with and being dependent upon exposure to light. Thus, while FAA was absent under constant dark (DD) conditions, it was still detected in constant light (LL). In contrast, general locomotor activity was arrhythmic in both constant darkness and constant light, pointing to a major contribution of light, in concert with the circadian clock, in timing locomotor activity in this species. Our data represents an important contribution to our understanding of the circadian timing system in the turbot and thereby the optimization of rearing protocols and the improvement of the well-being of turbot within fish farming environments