RESUMEN
Long-chain (≥C20 ) polyunsaturated fatty acids (LC-PUFAs) are physiologically important fatty acids for most animals, including humans. Although most LC-PUFA production occurs in aquatic primary producers such as microalgae, recent research indicates the ability of certain groups of (mainly marine) invertebrates for endogenous LC-PUFA biosynthesis and/or bioconversion from dietary precursors. The genetic pathways for and mechanisms behind LC-PUFA biosynthesis remain unknown in many invertebrates to date, especially in non-model species. However, the numerous genomic and transcriptomic resources currently available can contribute to our knowledge of the LC-PUFA biosynthetic capabilities of metazoans. Within our previously generated transcriptome of the benthic harpacticoid copepod Platychelipus littoralis, we detected expression of one methyl-end desaturase, one front-end desaturase, and seven elongases, key enzymes responsible for LC-PUFA biosynthesis. To demonstrate their functionality, we characterized eight of them using heterologous expression in yeast. The P. littoralis methyl-end desaturase has Δ15/17/19 desaturation activity, enabling biosynthesis of α-linolenic acid, eicosapentaenoic acid and docosahexaenoic acid (DHA) from 18:2 n-6, 20:4 n-6 and 22:5 n-6, respectively. Its front-end desaturase has Δ4 desaturation activity from 22:5 n-3 to DHA, implying that P. littoralis has multiple pathways to produce this physiologically important fatty acid. All studied P. littoralis elongases possess varying degrees of elongation activity for saturated and unsaturated fatty acids, producing aliphatic hydrocarbon chains with lengths of up to 30 carbons. Our investigation revealed a functionally diverse range of fatty acid biosynthesis genes in copepods, which highlights the need to scrutinize the role that primary consumers could perform in providing essential nutrients to upper trophic levels.
Asunto(s)
Ácido Eicosapentaenoico , Ácidos Grasos Insaturados , Humanos , Animales , Elongasas de Ácidos Grasos/genética , Elongasas de Ácidos Grasos/metabolismo , Ácidos Grasos Insaturados/genética , Ácidos Grasos Insaturados/metabolismo , Genoma , Saccharomyces cerevisiae/genética , Ácido Graso Desaturasas/genética , Ácido Graso Desaturasas/metabolismoRESUMEN
The splendid alfonsino Beryx splendens is a commercially important deep-sea fish in East Asian countries. Because the wild stock of this species has been declining, there is an urgent need to develop aquaculture systems. In the present study, we investigated the long-chain polyunsaturated fatty acid (LC-PUFA) requirements of B. splendens, which are known as essential dietary components in many carnivorous marine fish species. The fatty acid profiles of the muscles, liver, and stomach contents of B. splendens suggested that it acquires substantial levels of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) from its natural diet. The functional characterization of a fatty acid desaturase (Fads2) and three elongases (Elovl5, Elovl4a, and Elovl4b) from B. splendens confirmed their enzymatic capabilities in LC-PUFA biosynthesis. Fads2 showed Δ6 and Δ8 bifunctional desaturase activities. Elovl5 showed preferential elongase activities toward C18 and C20 PUFA substrates, whereas Elovl4a and Elovl4b showed activities toward various C18-22 substrates. Given that Fads2 showed no Δ5 desaturase activity and no other fads-like sequence was found in the B. splendens genome, EPA and arachidonic acid cannot be synthesized from C18 precursors; hence, they can be categorized as dietary essential fatty acids in B. splendens. EPA can be converted into DHA in B. splendens via the so-called Sprecher pathway. However, given that fads2 is only expressed in the brain, it is unlikely that the capacity of B. splendens to biosynthesize DHA from EPA can fulfill its physiological requirements. These results will be useful to researchers developing B. splendens aquaculture methods.
Asunto(s)
Proteínas de Peces , Peces , Animales , Elongasas de Ácidos Grasos/genética , Elongasas de Ácidos Grasos/metabolismo , Proteínas de Peces/metabolismo , Peces/metabolismo , Ácido Graso Desaturasas/genética , Ácidos Grasos Esenciales , Ácido Eicosapentaenoico , Ácidos Docosahexaenoicos , Dieta/veterinaria , Ácidos GrasosRESUMEN
The nutritional diversity of resources can affect the adaptive evolution of consumer metabolism and consumer diversification. The omega-3 long-chain polyunsaturated fatty acids eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3) have a high potential to affect consumer fitness, through their widespread effects on reproduction, growth and survival. However, few studies consider the evolution of fatty acid metabolism within an ecological context. In this review, we first document the extensive diversity in both primary producer and consumer fatty acid distributions amongst major ecosystems, between habitats and amongst species within habitats. We highlight some of the key nutritional contrasts that can shape behavioural and/or metabolic adaptation in consumers, discussing how consumers can evolve in response to the spatial, seasonal and community-level variation of resource quality. We propose a hierarchical trait-based approach for studying the evolution of consumers' metabolic networks and review the evolutionary genetic mechanisms underpinning consumer adaptation to EPA and DHA distributions. In doing so, we consider how the metabolic traits of consumers are hierarchically structured, from cell membrane function to maternal investment, and have strongly environment-dependent expression. Finally, we conclude with an outlook on how studying the metabolic adaptation of consumers within the context of nutritional landscapes can open up new opportunities for understanding evolutionary diversification.
Asunto(s)
Ácidos Grasos Omega-3 , Ácidos Grasos , Ácidos Docosahexaenoicos , Ecosistema , FenotipoRESUMEN
Long-chain (C20-24) polyunsaturated fatty acids (LC-PUFAs) are essential nutrients that are mostly produced in marine ecosystems. Previous studies suggested that gammarids have some capacity to endogenously produce LC-PUFAs. This study aimed to investigate the repertoire and functions of elongation of very long-chain fatty acid (Elovl) proteins in gammarids. Our results show that gammarids have, at least, three distinct elovl genes with putative roles in LC-PUFA biosynthesis. Phylogenetics allowed us to classify two elongases as Elovl4 and Elovl6, as they were bona fide orthologues of vertebrate Elovl4 and Elovl6. Moreover, a third elongase was named as "Elovl1/7-like" since it grouped closely to the Elovl1 and Elovl7 found in vertebrates. Molecular analysis of the deduced protein sequences indicated that the gammarid Elovl4 and Elovl1/7-like were indeed polyunsaturated fatty acid (PUFA) elongases, whereas Elovl6 had molecular features typically found in non-PUFA elongases. This was partly confirmed in the functional assays performed on the marine gammarid Echinogammarus marinus Elovl, which showed that both Elovl4 and Elovl1/7-like elongated PUFA substrates ranging from C18 to C22. E. marinus Elovl6 was only able to elongate C18 PUFA substrates, suggesting that this enzyme does not play major roles in the LC-PUFA biosynthesis of gammarids.
Asunto(s)
Anfípodos/enzimología , Clonación Molecular , Elongasas de Ácidos Grasos/metabolismo , Ácidos Grasos Insaturados/biosíntesis , Anfípodos/genética , Animales , Evolución Molecular , Elongasas de Ácidos Grasos/genética , Regulación Enzimológica de la Expresión Génica , Filogenia , Especificidad por SustratoRESUMEN
Insect pheromones are often derived from fatty acid metabolism. Fatty acid desaturases, enzymes introducing double bonds into fatty acids, are crucial for the biosynthesis of these chemical signals. Δ12-desaturases catalyse the biosynthesis of linoleic acid by introducing a second double bond into oleic acid, but have been identified in only a few animal species. Here, we report the functional characterisation of two Δ12-desaturases, Nvit_D12a and Nvit_D12b, from the parasitic wasp Nasonia vitripennis. We demonstrate that Nvit_D12a is expressed in the rectal vesicle of males where they produce a linoleic acid-derived sex pheromone to attract virgin females. 13C-labelling experiments with Urolepis rufipes, a closely related species belonging to the 'Nasonia group', revealed that females, but not males, are able to synthesise linoleic acid. U. rufipes males produce an isoprenoid sex pheromone in the same gland and do not depend on linoleic acid for pheromone production. This suggests that Δ12-desaturases are common in the 'Nasonia group', but acquired a specialised function in chemical communication of those species that use linoleic acid as a pheromone precursor. Phylogenetic analysis suggests that insect Δ12-desaturases have evolved repeatedly from Δ9-desaturases in different insect taxa. Hence, insects have developed a way to produce linoleic acid independent of the omega desaturase subfamily which harbours all of the eukaryotic Δ12-desaturases known so far.
Asunto(s)
Ácido Graso Desaturasas/genética , Proteínas de Insectos/genética , Ácido Linoleico/metabolismo , Atractivos Sexuales/biosíntesis , Avispas/metabolismo , Animales , Ácido Graso Desaturasas/metabolismo , Femenino , Proteínas de Insectos/metabolismo , MasculinoRESUMEN
BACKGROUND: Provision of long-chain polyunsaturated fatty acids (LC-PUFA) in vertebrates occurs through the diet or via endogenous production from C18 precursors through consecutive elongations and desaturations. It has been postulated that the abundance of LC-PUFA in the marine environment has remarkably modulated the gene complement and function of Fads in marine teleosts. In vertebrates two fatty acyl desaturases, namely Fads1 and Fads2, encode ∆5 and ∆6 desaturases, respectively. To fully clarify the evolutionary history of LC-PUFA biosynthesis in vertebrates, we investigated the gene repertoire and function of Fads from species placed at key evolutionary nodes. RESULTS: We demonstrate that functional Fads1Δ5 and Fads2∆6 arose from a tandem gene duplication in the ancestor of vertebrates, since they are present in the Arctic lamprey. Additionally, we show that a similar condition was retained in ray-finned fish such as the Senegal bichir and spotted gar, with the identification of fads1 genes in these lineages. Functional characterisation of the isolated desaturases reveals the first case of a Fads1 enzyme with ∆5 desaturase activity in the Teleostei lineage, the Elopomorpha. In contrast, in Osteoglossomorpha genomes, while no fads1 was identified, two separate fads2 duplicates with ∆6 and ∆5 desaturase activities respectively were uncovered. CONCLUSIONS: We conclude that, while the essential genetic components involved LC-PUFA biosynthesis evolved in the vertebrate ancestor, the full completion of the LC-PUFA biosynthesis pathway arose uniquely in gnathostomes.
Asunto(s)
Evolución Molecular , Ácido Graso Desaturasas/genética , Ácidos Grasos Insaturados/biosíntesis , Peces/genética , Peces/metabolismo , Secuencia de Aminoácidos , Animales , Ácido Graso Desaturasas/química , Filogenia , Homología de Secuencia de Ácido NucleicoRESUMEN
The biosynthetic capability of the long-chain polyunsaturated fatty acids (LC-PUFA) in teleosts are highly diversified due to evolutionary events such as gene loss and subsequent neo- and/or sub-functionalisation of enzymes encoded by existing genes. In the present study, we have comprehensively characterised genes potentially involved in LC-PUFA biosynthesis, namely one front-end desaturase (fads2) and eight fatty acid elongases (elovl1a, elovl1b, elovl4a, elovl4b, elovl5, elovl7, elovl8a and elovl8b) from an amphidromous teleost, Ayu sweetfish, Plecoglossus altivelis. Functional analysis confirmed Fads2 with Δ6, Δ5 and Δ8 desaturase activities towards multiple PUFA substrates and several Elovl enzymes exhibited elongation capacities towards C18-20 or C18-22 PUFA substrates. Consequently, P. altivelis possesses a complete enzymatic capability to synthesise physiologically important LC-PUFA including arachidonic acid (ARA, 20:4n-6), eicosapentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA, 22:6n-3) from their C18 precursors. Interestingly, the loss of elovl2 gene in P. altivelis was corroborated by genomic and phylogenetic analyses. However, this constraint would possibly be overcome by the function of alternative Elovl enzymes, such as Elovl1b, which has not hitherto been functionally characterised in teleosts. The present study contributes novel insights into LC-PUFA biosynthesis in the relatively understudied teleost group, Osmeriformes (Stomiati), thereby enhancing our understanding of the complement of LC-PUFA biosynthetic genes within teleosts.
Asunto(s)
Ácido Graso Desaturasas , Elongasas de Ácidos Grasos , Ácidos Grasos Insaturados , Osmeriformes , Animales , Ácidos Grasos Insaturados/metabolismo , Ácidos Grasos Insaturados/biosíntesis , Ácidos Grasos Insaturados/genética , Osmeriformes/metabolismo , Osmeriformes/genética , Ácido Graso Desaturasas/metabolismo , Ácido Graso Desaturasas/genética , Elongasas de Ácidos Grasos/metabolismo , Elongasas de Ácidos Grasos/genética , Filogenia , Proteínas de Peces/metabolismo , Proteínas de Peces/genética , Vías Biosintéticas/genética , Acetiltransferasas/metabolismo , Acetiltransferasas/genéticaRESUMEN
Studies have suggested that dietary purine nucleotides (NT) affect the muscle and liver fatty acid composition of rainbow trout. To examine the direct regulation of liver fatty acid metabolism by purine NT in rainbow trout, the liver cells were cultured in the presence of 500 µmol/L inosine, adenosine, or guanosine monophosphate (IMP, AMP, or GMP). The expression of pparα was significantly decreased in the liver cells cultured with purine NT for 24 h, whereas the expression of fads2 (Δ5) was increased. Docosahexaenoic acid (DHA) content in the liver cells was significantly higher after culturing with GMP. To determine the dose-dependent effects of NT, 50, 100, and 500 µmol/L GMP was added to the liver cells cultured in L-15 medium. At 48 h, 20:4n - 6, 22:5n - 3, 22:6n - 3, Æ© PUFA, and Æ© n - 3 PUFA content in the 50 µM GMP-containing medium was significantly higher compared with the other medium. The expression of Δ5 fads2, elovl2, and elovl5 in the liver cells was significantly higher in the 500 µmol/L GMP-containing medium at 48 h along with increased srebp-1 expression. These results suggest that purine NT directly affect fatty acid composition through modification of fatty acid metabolism-related genes in the liver of rainbow trout.
Asunto(s)
Oncorhynchus mykiss , Animales , Oncorhynchus mykiss/genética , Oncorhynchus mykiss/metabolismo , Metabolismo de los Lípidos , Hígado/metabolismo , Ácidos Grasos/metabolismo , Purinas/farmacología , Purinas/metabolismoRESUMEN
Previous data revealed the unexpected presence of genes encoding for long-chain polyunsaturated fatty acid (LC-PUFA) biosynthetic enzymes in transcriptomes from freshwater gammarids but not in marine species, even though closely related species were compared. This study aimed to clarify the origin and occurrence of selected LC-PUFA biosynthesis gene markers across all published gammarid transcriptomes. Through systematic searches, we confirmed the widespread occurrence of sequences from seven elongases and desaturases involved in LC-PUFA biosynthesis, in transcriptomes from freshwater gammarids but not marine species, and clarified that such occurrence is independent from the gammarid species and geographical origin. The phylogenetic analysis established that the retrieved elongase and desaturase sequences were closely related to bdelloid rotifers, confirming that multiple transcriptomes from freshwater gammarids contain contaminating rotifers' genetic material. Using the Adineta steineri genome, we investigated the genomic location and exon-intron organization of the elongase and desaturase genes, establishing they are all genome-anchored and, importantly, identifying instances of horizontal gene transfer. Finally, we provide compelling evidence demonstrating Bdelloidea desaturases and elongases enable these organisms to perform all the reactions for de novo biosynthesis of PUFA and, from them, LC-PUFA, an advantageous trait when considering the low abundance of these essential nutrients in freshwater environments.
Asunto(s)
Ácido Graso Desaturasas , Transcriptoma , Elongasas de Ácidos Grasos/genética , Elongasas de Ácidos Grasos/metabolismo , Filogenia , Ácido Graso Desaturasas/genética , Ácido Graso Desaturasas/metabolismo , Ácidos Grasos Insaturados , Agua DulceRESUMEN
The long-standing paradigm establishing that global production of Omega-3 (n-3) long-chain polyunsaturated fatty acids (LC-PUFA) derived almost exclusively from marine single-cell organisms, was recently challenged by the discovery that multiple invertebrates possess methyl-end (or ωx) desaturases, critical enzymes enabling the biosynthesis of n-3 LC-PUFA. However, the question of whether animals with ωx desaturases have complete n-3 LC-PUFA biosynthetic pathways and hence can contribute to the production of these compounds in marine ecosystems remained unanswered. In the present study, we investigated the complete enzymatic complement involved in the n-3 LC-PUFA biosynthesis in Tigriopus californicus, an intertidal harpacticoid copepod. A total of two ωx desaturases, five front-end desaturases and six fatty acyl elongases were successfully isolated and functionally characterized. The T. californicus ωx desaturases enable the de novo biosynthesis of C18 PUFA such as linoleic and α-linolenic acids, as well as several n-3 LC-PUFA from n-6 substrates. Functions demonstrated in front-end desaturases and fatty acyl elongases unveiled various routes through which T. californicus can biosynthesize the physiologically important arachidonic and eicosapentaenoic acids. Moreover, T. californicus possess a Δ4 desaturase, enabling the biosynthesis of docosahexaenoic acid via the 'Δ4 pathway'. In conclusion, harpacticoid copepods such as T. californicus have complete n-3 LC-PUFA biosynthetic pathways and such capacity illustrates major roles of these invertebrates in the provision of essential fatty acids to upper trophic levels.
Asunto(s)
Copépodos/fisiología , Ácidos Docosahexaenoicos/biosíntesis , Regulación Enzimológica de la Expresión Génica , Metabolismo de los Lípidos , Animales , Cromatografía de Gases , Copépodos/clasificación , Ácido Graso Desaturasas/genética , Ácido Graso Desaturasas/metabolismo , Ácidos Grasos/análisis , Ácidos Grasos/metabolismo , Redes y Vías Metabólicas , FilogeniaRESUMEN
Long-chain (≥C20) polyunsaturated fatty acids (LC-PUFA), including eicosapentaenoic acid (EPA, 20:5n-3), arachidonic acid (ARA, 20:4n-6) and docosahexaenoic acid (DHA, 22:6n-3), are essential in multiple physiological processes, especially during early development of vertebrates. LC-PUFA biosynthesis is achieved by two key families of enzymes, fatty acyl desaturases (Fads) and elongation of very long-chain fatty acid (Elovl). The present study determined the expression patterns of genes encoding desaturases (fads1 and fads2) and elongases (elovl2 and elovl5) involved in the LC-PUFA biosynthesis during early life-stages of the tropical gar Atractosteus tropicus. We further analyzed the fatty acid profiles during early development of A. tropicus to evaluate the impact of Fads and Elovl enzymatic activities. Specific oligonucleotides were designed from A. tropicus transcriptome to perform qPCR (quantitative polymerase chain reaction) on embryonic and larval stages, along with several organs (intestine, white muscle, brain, liver, heart, mesenteric adipose, kidney, gill, swim bladder, stomach, and spleen) collected from juvenile specimens. Fatty acid content of feeds and embryonic and larval stages were analyzed. Results show that fads1, fads2, elovl2 and elovl5 expression was detected from embryonic stages with expression peaks from day 15 post hatching, which could be related to transcriptional and dietary factors. Moreover, fads1, fads2 and elovl2 showed a higher expression in intestine, while elovl5 showed a higher expression in liver, suggesting that the tropical gar activates its LC-PUFA biosynthetic machinery to produce ARA, EPA and DHA to satisfy physiological demands at crucial developmental milestones during early development.
Asunto(s)
Ácido Graso Desaturasas/genética , Elongasas de Ácidos Grasos/genética , Ácidos Grasos Insaturados/biosíntesis , Proteínas de Peces/metabolismo , Peces/metabolismo , Regulación del Desarrollo de la Expresión Génica , Lipogénesis , Animales , Proteínas de Peces/genética , Peces/genética , Peces/crecimiento & desarrollo , TranscriptomaRESUMEN
The production of xenogenic gametes from large-bodied, commercially important marine fish species in closely related smaller host fish species with short generation times may enable rapid and simple seed production of the target species. As a first step toward this goal, we assessed the suitability of chub mackerel, Scomber japonicus, as a small-bodied recipient species for xenogenic spermatogonial transplantation. Histological observation of the early gonadal development of chub mackerel larvae and transplantation of fluorescent-labeled spermatogonia from Nibe croaker, Nibea mitsukurii, revealed that 5.3-mm chub mackerel larvae were suitable recipients for successful transplantation. Intraperitoneally transplanted xenogenic spermatogonia efficiently colonized the gonads of these recipient larvae, and donor-derived Nibe croaker germ cells proliferated rapidly soon after colonization. Moreover, gonadal soma-derived growth factor (gsdf) mRNA, a gonadal somatic cell marker, was expressed in recipient-derived cells surrounding the incorporated donor-derived germ cells, suggesting that donor-derived germ cells had settled at an appropriate location in the recipient gonad. Our data show that xenogenic spermatogonial transplantation was successful in chub mackerel and that the somatic microenvironment of the chub mackerel gonad can support the colonization, survival, and proliferation of intraperitoneally transplanted xenogenic germ cells derived from a donor species of a different taxonomic family.
Asunto(s)
Especies en Peligro de Extinción , Supervivencia de Injerto , Técnicas Reproductivas Asistidas/veterinaria , Espermatogonias/trasplante , Trasplante Heterólogo/veterinaria , Animales , Proliferación Celular , Explotaciones Pesqueras , Masculino , Perciformes , Especificidad de la Especie , Espermatogénesis/fisiología , Espermatogonias/crecimiento & desarrollo , Trasplante Heterólogo/métodosRESUMEN
Omega-3 (ω3 or n-3) long-chain polyunsaturated fatty acids (PUFA), including eicosapentaenoic acid and docosahexaenoic acid (DHA), play physiologically important roles in vertebrates. These compounds have long been believed to have originated almost exclusively from aquatic (mostly marine) single-cell organisms. Yet, a recent study has discovered that many invertebrates possess a type of enzymes called methyl-end desaturases (ωx) that enables them to endogenously produce n-3 long-chain PUFA and could make a significant contribution to production of these compounds in the marine environment. Polychaetes are major components of benthic fauna and thus important to maintain a robust food web as a recycler of organic matter and a prey item for higher trophic level species like fish. In the present study, we investigated the ωx enzymes from the common ragworm, Hediste diversicolor, a common inhabitant in sedimentary littoral ecosystems of the North Atlantic. Functional assays of the H. diversicolorωx demonstrated unique desaturation capacities. An ω3 desaturase mediated the conversion of n-6 fatty acid substrates into their corresponding n-3 products including DHA. A further enzyme possessed unique regioselectivities combining both ω6 and ω3 desaturase activities. These results illustrate that the long-chain PUFA biosynthetic enzymatic machinery of aquatic invertebrates such as polychaetes is highly diverse and clarify that invertebrates can be major contributors to fatty acid trophic upgrading in aquatic food webs. This article is part of the theme issue 'The next horizons for lipids as 'trophic biomarkers': evidence and significance of consumer modification of dietary fatty acids'.
Asunto(s)
Ácido Graso Desaturasas/metabolismo , Ácidos Grasos Omega-3/metabolismo , Ácidos Grasos Omega-6/metabolismo , Cadena Alimentaria , Poliquetos/metabolismo , AnimalesRESUMEN
The colonisation of freshwater environments by marine fishes has historically been considered a result of adaptation to low osmolality. However, most marine fishes cannot synthesise the physiologically indispensable fatty acid, docosahexaenoic acid (DHA), due to incomplete DHA biosynthetic pathways, which must be adapted to survive in freshwater environments where DHA is poor relative to marine environments. By analysing DHA biosynthetic pathways of one marine and three freshwater-dependent species from the flatfish family Achiridae, we revealed that functions of fatty acid metabolising enzymes have uniquely and independently evolved by multi-functionalisation or neofunctionalisation in each freshwater species, such that every functional combination of the enzymes has converged to generate complete and functional DHA biosynthetic pathways. Our results demonstrate the elaborate patchwork of fatty acid metabolism and the importance of acquiring DHA biosynthetic function in order for fish to cross the nutritional barrier at the mouth of rivers and colonise freshwater environments.
Asunto(s)
Vías Biosintéticas/genética , Monitoreo del Ambiente , Peces Planos/genética , Animales , Peces Planos/fisiología , Agua Dulce , Humanos , FilogeniaRESUMEN
The long-chain (≥C20) polyunsaturated fatty acid biosynthesis capacity of fish varies among species, with trophic level hypothesised as a major factor. The biosynthesis capacity is largely dependent upon the presence of functionally diversified fatty acyl desaturase 2 (Fads2) enzymes, since many teleosts have lost the gene encoding a Δ5 desaturase (Fads1). The present study aimed to characterise Fads2 from four teleosts occupying different trophic levels, namely Sarpa salpa, Chelon labrosus, Pegusa lascaris and Atherina presbyter, which were selected based on available data on functions of Fads2 from closely related species. Therefore, we had insight into the variability of Fads2 within the same phylogenetic group. Our results showed that Fads2 from S. salpa and C. labrosus were both Δ6 desaturases with further Δ8 activity while P. lascaris and A. presbyter Fads2 showed Δ4 activity. Fads2 activities of herbivorous S. salpa are consistent with those reported for carnivorous Sparidae species. The results suggested that trophic level might not directly drive diversification of teleost Fads2 as initially hypothesised, and other factors such as the species' phylogeny appeared to be more influential. In agreement, Fads2 activities from P. lascaris and A. presbyter were similar to their corresponding phylogenetic counterparts Solea senegalensis and Chirostoma estor.
Asunto(s)
Ácido Graso Desaturasas/metabolismo , Ácidos Grasos Insaturados/biosíntesis , Proteínas de Peces/metabolismo , Peces/fisiología , Secuencia de Aminoácidos/genética , Animales , Carnivoría/fisiología , Pruebas de Enzimas , Ácido Graso Desaturasas/química , Ácido Graso Desaturasas/genética , Ácidos Grasos Insaturados/química , Proteínas de Peces/química , Proteínas de Peces/genética , Herbivoria/fisiología , Filogenia , Alineación de Secuencia , Especificidad de la Especie , Especificidad por Sustrato/fisiologíaRESUMEN
Since the late 19th century, the Amazon species Colossoma macropomum (tambaqui) has been exploited commercially and the climate change has contributed to decline in tambaqui numbers. Although germ cell cryopreservation and transplantation can help preserve the species' genetic resources semipermanently, its germ cell behavior has not been analyzed to date. In this study, we isolated the tambaqui's dead end gene (dnd) homolog (tdnd) and used it as a molecular marker for germ cells to obtain basic information essential for transplantation. The amino acid sequence showed 98% similarity and 53% identity with the zebrafish dnd. Phylogenetic analysis and the presence of consensus motifs known for dnd revealed that tdnd encodes the dnd ortholog and its transcript is detectable only in the testes and ovaries, showing a strong positive signal in oocytes and spermatogonia. The tambaqui possesses, at least, three different transcripts of tdnd which show dissimilar expression profile in undifferentiated and sexually mature animals, suggesting that they play distinct roles in germline development and they may influence the choice of donors for the cell transplantation study.
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Empalme Alternativo/genética , Células Germinativas/crecimiento & desarrollo , Proteínas de Unión al ARN/genética , Proteínas de Pez Cebra/genética , Pez Cebra/genética , Animales , Biomarcadores/metabolismo , Embrión no Mamífero , Femenino , Regulación del Desarrollo de la Expresión Génica/genética , Células Germinativas/metabolismo , Masculino , Filogenia , Pez Cebra/crecimiento & desarrolloRESUMEN
The native Amazonian fish tambaqui (Colossoma macropomum) is the second-largest scaled fish in South America and the most common native species in Brazil. To preserve genetic resources with sufficient genetic diversity through germ cell cryopreservation and transplantation techniques, a molecular marker for identifying the cells is required to trace them during the manipulation processes. The vasa gene is a promising candidate, as its specific expression in germ cell lineage has been well-conserved throughout animal evolution. In this study, the full sequence of the vasa cDNA homolog from tambaqui was isolated and characterized, showing an open reading frame of 2010â¯bp encoding 669 amino acids. The putative protein was shown to contain eight conserved motifs of the DEAD-box protein family and high similarity to vasa homologs of other species. Tambaqui vasa (tvasa) mRNA expression was specific to the gonads, and in situ hybridization showed signals only in oocytes and spermatogonia. The results suggested that tvasa could be a useful germ cell marker in this species.
Asunto(s)
Characidae/genética , Clonación Molecular/métodos , ARN Helicasas DEAD-box/genética , Células Germinativas/metabolismo , Animales , Characidae/metabolismo , Conservación de los Recursos Naturales , Femenino , Proteínas de Peces/genética , Masculino , Sistemas de Lectura Abierta , Especificidad de Órganos , FilogeniaRESUMEN
In vertebrates, the essential fatty acids (FA) that satisfy the dietary requirements for a given species depend upon its desaturation and elongation capabilities to convert the C18 polyunsaturated fatty acids (PUFA), namely linoleic acid (LA, 18:2n-6) and α-linolenic acid (ALA, 18:3n-3), into the biologically active long-chain (C20-24) polyunsaturated fatty acids (LC-PUFA), including arachidonic acid (ARA, 20:4n-6), eicosapentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA, 22:6n-3). Recent studies have established that tambaqui (Colossoma macropomum), an important aquaculture-produced species in Brazil, is a herbivorous fish that can fulfil its essential FA requirements with dietary provision C18 PUFA LA and ALA, although the molecular mechanisms underpinning such ability remained unclear. The present study aimed at cloning and functionally characterizing genes encoding key desaturase and elongase enzymes, namely fads2, elovl5 and elovl2, involved in the LC-PUFA biosynthetic pathways in tambaqui. First, a fads2-like desaturase was isolated from tambaqui. When expressed in yeast, the tambaqui Fads2 showed Δ6, Δ5 and Δ8 desaturase capacities within the same enzyme, enabling all desaturation reactions required for ARA, EPA and DHA biosynthesis. Moreover, tambaqui possesses two elongases that are bona fide orthologs of elovl5 and elovl2. Their functional characterization confirmed that they can operate towards a variety of PUFA substrates with chain lengths ranging from 18 to 22 carbons. Overall our results provide compelling evidence that demonstrates that all the desaturase and elongase activities required to convert LA and ALA into ARA, EPA and DHA are present in tambaqui within the three genes studied herein, i.e. fads2, elovl5 and elovl2.
Asunto(s)
Ácido Araquidónico/metabolismo , Characidae/fisiología , Ácidos Docosahexaenoicos/metabolismo , Ácido Eicosapentaenoico/metabolismo , Hígado/enzimología , Acetiltransferasas/química , Acetiltransferasas/genética , Acetiltransferasas/metabolismo , Secuencia de Aminoácidos , Animales , Acuicultura , Brasil , Characidae/crecimiento & desarrollo , Ácido Graso Desaturasas/química , Ácido Graso Desaturasas/genética , Ácido Graso Desaturasas/metabolismo , Elongasas de Ácidos Grasos , Proteínas de Peces/química , Proteínas de Peces/genética , Proteínas de Peces/metabolismo , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Hígado/crecimiento & desarrollo , Hígado/metabolismo , Necesidades Nutricionales , Organismos Modificados Genéticamente , Filogenia , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Ríos , Saccharomyces cerevisiae/enzimología , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Especificidad por SustratoRESUMEN
The interest in understanding the capacity of aquatic invertebrates to biosynthesise omega-3 (ω3) long-chain (≥C20) polyunsaturated fatty acids (LC-PUFA) has increased in recent years. Using the common octopus Octopus vulgaris as a model species, we previously characterised a ∆5 desaturase and two elongases (i.e. Elovl2/5 and Elovl4) involved in the biosynthesis of LC-PUFA in molluscs. The aim of this study was to characterise both molecularly and functionally, two methyl-end (or ωx) desaturases that have been long regarded to be absent in most animals. O. vulgaris possess two ωx desaturase genes encoding enzymes with ∆12 and ω3 regioselectivities enabling the de novo biosynthesis of the C18 PUFA 18:2ω6 (LA, linoleic acid) and 18:3ω3 (ALA, α-linolenic acid), generally regarded as dietary essential for animals. The O. vulgaris ∆12 desaturase ("ωx2") mediates the conversion of 18:1ω9 (oleic acid) into LA, and subsequently, the ω3 desaturase ("ωx1") catalyses the ∆15 desaturation from LA to ALA. Additionally, the O. vulgaris ω3 desaturase has ∆17 capacity towards a variety of C20 ω6 PUFA that are converted to their ω3 PUFA products. Particularly relevant was the affinity of the ω3 desaturase towards 20:4ω6 (ARA, arachidonic acid) to produce 20:5ω3 (EPA, eicosapentaenoic acid), as supported by yeast heterologous expression, and enzymatic activity exhibited in vivo when paralarvae were incubated in the presence of [1-14C]20:4ω6. These results confirmed that several routes enabling EPA biosynthesis are operative in O. vulgaris whereas ARA and docosahexaenoic acid (DHA, 22:6ω3) should be considered essential fatty acids since endogenous production appears to be limited.
Asunto(s)
Ácido Graso Desaturasas/metabolismo , Ácidos Grasos Insaturados/biosíntesis , Octopodiformes/metabolismo , Animales , Ácido Araquidónico/biosíntesis , Ácido Araquidónico/metabolismo , Ácidos Docosahexaenoicos/biosíntesis , Ácidos Docosahexaenoicos/metabolismo , Ácido Eicosapentaenoico/biosíntesis , Ácido Graso Desaturasas/genética , Ácido Linoleico/biosíntesis , Octopodiformes/enzimología , Ácido alfa-Linolénico/biosíntesisRESUMEN
Colonization of new ecological niches has triggered large adaptive radiations. Although some lineages have made use of such opportunities, not all do so. The factors causing this variation among lineages are largely unknown. Here, we show that deficiency in docosahexaenoic acid (DHA), an essential ω-3 fatty acid, can constrain freshwater colonization by marine fishes. Our genomic analyses revealed multiple independent duplications of the fatty acid desaturase gene Fads2 in stickleback lineages that subsequently colonized and radiated in freshwater habitats, but not in close relatives that failed to colonize. Transgenic manipulation of Fads2 in marine stickleback increased their ability to synthesize DHA and survive on DHA-deficient diets. Multiple freshwater ray-finned fishes also show a convergent increase in Fads2 copies, indicating its key role in freshwater colonization.