Dissolved oxygen and ammonia affect ammonia production via GDH/AMPK signaling pathway and alter flesh quality in Chinese perch (Siniperca chuatsi).

The dissolved oxygen (DO) and ammonia are crucial to the growth of Chinese perch (Siniperca chuatsi). Information on the effects of DO and total ammonia nitrogen (TAN) in regulating ammonia nitrogen excretion and flesh quality in Chinese perch is scanty. This study aimed to evaluate the effects of dissolved DO at oxygen levels of 3 mg/L and 9 mg/L, as well as the TAN concentrations of 0.3 mg/L and 0.9 mg/L on ammonia excretion and flesh quality. Results showed that the ammonia contents in plasma, muscle, and liver of the 9 mg/L DO group were significantly higher than those of the 3 mg/L DO group (P < 0.05). However, the expression of AMPK-related signaling pathway genes (gdh, lkb1, and ampd) and flesh quality indicators (gumminess, chewiness, hardness) in the 9 mg/L DO group were significantly lower than those in the 3 mg/L DO group. Under long-term exposure to 0.9 mg/L TAN, the ammonia contents in plasma and gill filaments, as well as muscle flesh quality (resilience, gumminess, chewiness, cohesiveness), were significantly lower than those in the 0.3 mg/L TAN group (P < 0.05). However, the activities of GDH and AMPD enzymes in the 0.9 mg/L TAN group were significantly higher than those in the 0.3 mg/L TAN group. In summary, when fish are exposed to 3 mg/L DO and 0.9 mg/L TAN in the environment for a long time, their amino acids are used for transamination and deamination, resulting in insufficient energy supply for Chinese perch, whereas 9 mg/L DO and 0.9 mg/L TAN caused deterioration of the flesh quality.

Knockout of lws1 in zebrafish (Danio rerio) reveals its role in regulating feeding and vision-guided behavior.

Long-wave sensitive (LWS) is a G protein-coupled receptor expressed in the retina, and zebrafish is a better model organism for studying vision, but the role of LWS1 in vision-guided behavior of larvae fish has rarely been reported. In this study, we found that zebrafish lws1 and lws2 are tandemly replicated genes, both with six exons, with lws1 being more evolutionarily conserved. The presence of Y277F in the amino acid sequence of lws2 may have contributed to the shift of λmax to green light. We established a lws1 knockout zebrafish model using CRISPR/Cas9 technology. Lws1-/- larvae showed significantly higher levels of feeding and appetite gene (agrp) expression than WT, and significantly lower levels of anorexia gene (pomc, cart) expression. In addition, green light gene compensation was observed in lws1-/- larvae with significantly increased expression levels of rh2-1. The light-dark movement test showed that lws1-/- larvae were more active under light-dark transitions or vibrational stimuli, and the expression of phototransduction-related genes was significantly up-regulated. This study reveals the important role of lws1 gene in the regulation of vision-guided behavior in larvae.

Addition of α-ketoglutaric acid (AKG) reduces deamination in Chinese perch (Siniperca chuatsi) fed with fermented soybean meal as a substitute for fishmeal.

To alleviate amino acid imbalances in fermented soybean meal as a replacement for fishmeal feeds, this study evaluated the effects of adding lysine (Lys), methionine (Met), and α-ketoglutaric acid (AKG) to fermented soybean meals for Chinese perch. Chinese perch (34 ± 3 g) were fed five diets for 66 days (fishmeal as the protein source of the basal diet [FM]; fermented soybean meal as a substitute for 30% fishmeal in the soybean meal diet [FSM]; addition of crystalline Lys and Met [AA]; addition of α-ketoglutaric acid [AKG]; and simultaneous addition of crystalline Lys, Met, and AKG [BA] to the soybean meal diet). At the end of the feeding trial, the FSM group had the highest feeding rate and the lowest weight gain rate among all the groups. The FM group had the highest protein retention and the lowest feed efficiency among the groups. The mRNA transcription level of genes related to the AMP-activating protein (AMPK) signaling pathway and amino acid response (AAR) signaling pathway (lkb1, atf4, and gcn2) were highest in the AA group (P < 0.05) but lower in the AKG and BA groups. In the AKG group, the mRNA transcription level of the gluconeogenesis pathway-related gene (pepck and g6pase) was significantly higher than that in the other four groups, but the mRNA transcription level of genes related to amino acid catabolism (gdh and ampd) was lower. Among all the groups, the FSM group had the lowest mRNA transcription level of genes associated with the mammalian target of rapamycin (mTOR) signaling pathway (mtor and s6k). These findings imply that the feeding rate of Chinese perch in the fermented soybean meal group was the highest, but the protein retention was the lowest, while the addition of Lys, Met, and AKG improved protein retention. In conclusion, the addition of AKG to fermented soybean meal as a fishmeal substitute reduced amino acid deamination, enhanced gluconeogenesis, and increased protein deposition, which contributed to the growth of Chinese perch, alleviated amino acid imbalances, and improved the feed utilization of Chinese perch.

Dietary supplementation of VA enhances growth, feed utilization, glucose and lipid metabolism, appetite, and antioxidant capacity of Chinese perch (Siniperca chuatsi).

The aim of this study was to investigate the effect of dietary vitamin A on juvenile Chinese perch (Siniperca chuatsi). Chinese perch were fed with five experimental diets containing 0, 20, 40, 60, and 80 mg VA·kg-1 for 8 weeks. Results showed that dietary vitamin A significantly influenced the fish's growth, feed utilization, glucose and lipid metabolism, appetite, and antioxidant capacity. Vitamin A-supplemented groups had higher weight gain rate (WGR) and specific growth rate (SGR) compared to the control group. Feed conversion ratio (FCR) was also lower in the vitamin A-supplemented groups. Dietary vitamin A had no significant effect on the survival rate (SR). Compared to the control group, fish fed with vitamin A had increased feed intake (FI), and the expression of appetite-promoting genes (npy and agrp) was significantly higher in the 40 mg VA·kg-1 group. Vitamin A also enhanced the utilization of dietary protein by Chinese perch. The serum glucose content of the fish fed with 40 mg VA·kg-1 diet was significantly higher than that of the control group and 20 mg VA·kg-1 diet, indicating that the promoting effect of VA on gluconeogenesis was greater than that on glycolysis. Additionally, dietary vitamin A increased the expression of lipid metabolism-related genes (hl and fas) and antioxidant genes (nrf2 and gpx) in the fish. These results suggest that the optimal vitamin A requirement of juvenile Chinese perch bream was estimated to be 37.32 mg VA·kg-1 based on broken-line regression analysis of WGR. In conclusion, this study provides valuable insights into the potential benefits of dietary vitamin A on the growth, metabolism, and antioxidant capacity of Chinese perch.

Knockout of SWS2 in zebrafish (Danio rerio) reveals its roles in feeding and phototactic behaviors.

Common ancestor of vertebrates had four cone opsin subfamilies to obtain color vision: ultraviolet-sensitive (SWS1), blue-sensitive (SWS2), middle wavelength sensitive (RH2) and long wavelength sensitive (LWS). Nevertheless, eutherian mammals had lost the SWS2 and RH2 opsins during their nocturnal lifestyle. Many studies had demonstrated the role of SWS1 and LWS cones in feeding, mate choice and skin pigment cell formation. However, the role of SWS2 and RH2 cones remain elusive. In the present study, we used an ideal model visual system, zebrafish, which still have the four cone opsins, to generate a SWS2 knockout zebrafish line. Through various behavioral test, we found that sws2-/- zebrafish larvae exhibited increased food intake compared with WT. Additionally, there were significantly increased the gene expression of phototransduction pathways in sws2-/- zebrafish larvae. Compared to WT, mutant zebrafish showed weaker phototaxis of red light and changed sensitivity of yellow, red and blue lights. But both mutant and WT zebrafish preferred the red light than other wavelengths of light. Taken together, we proposed that SWS2 cone is not necessary for feeding and phototaxis in zebrafish.

Neuropeptide Y receptor Y2 (npy2r) deficiency reduces anxiety and increases food intake in Japanese medaka (Oryzias latipes).

Neuropeptide Y receptor Y2 (npy2r) is an important receptor gene involved in anxiety and feeding regulation in mammals. Since NPY receptors have different receptor gene deletions in mammals and teleost fish, it is not clear whether npy2r has the similar function in fish as in mammals. In this study, we used the CRISPR/Cas9 system to establish npy2r-deficient medaka (Oryzias latipes). Unexpectedly, the deletion of npy2r resulted in the npy2r +/- medaka were all-male, therefore, npy2r homozygous mutant lines could not be established. The deletion of npy2r increased the food intake in medaka, and the expression levels of appetite stimulating genes (agrp, npy) increased significantly, while the expression levels of anorexia factors (cck, pomc) decreased significantly. Moreover, the absence of npy2r significantly increased the total length and body weight of medaka. The mirror test and open field test showed that npy2r +/- medaka improved sociability and reduced anxiety-like behavior, qRT-PCR analysis showed that the expression levels of anxiety related genes (th1, th2, gr1, gr2, and mr) in npy2r +/- medaka were significantly decreased. So far, this is the first npy2r gene knockout model established in fish and demonstrates that npy2r plays an important role in the regulation of reproduction, feeding and anxiety in fish.

Dietary soybean lecithin promoted growth performance and feeding in juvenile Chinese perch (Siniperca chuatsi) could be by optimizing glucolipid metabolism.

To explore the potential benefits of dietary phospholipids (PLs) in fish glucose metabolism and to promote feed culture of Chinese perch (Siniperca chuatsi), we set up six diets to feed Chinese perch (initial mean body weight 37.01 ± 0.20 g) for 86 days, including: Control diet (CT), 1% (SL1), 2% (SL2), 3% (SL3), 4% (SL4) soybean lecithin (SL) and 2% (KO2) krill oil (KO) supplemental diets (in triplicate, 20 fish each). Our study found that the SL2 significantly improved the weight gain rate and special growth rate, but the KO2 did not. In addition, the SL2 diet significantly improved feed intake, which is consistent with the mRNA levels of appetite-related genes (npy, agrp, leptin A). Additionally, in the CT and SL-added groups, leptin A expression levels were nearly synchronized with serum glucose levels. Besides, the SL2 significantly upregulated expression levels of glut2, gk, cs, fas and downregulated g6pase in the liver, suggesting that it may enhance glucose uptake, aerobic oxidation, and conversion to fatty acids. The SL2 also maintained the hepatic crude lipid content unchanged compared to the CT, possibly by significantly down-regulating the mRNA level of hepatic lipase gene (hl), and by elevating serum low-density lipoprotein (LDL) level and intraperitoneal fat ratio in significance. Moreover, the serum high-density lipoprotein levels were significantly increased by PL supplementation, and the SL2 further significantly increased serum total cholesterol and LDL levels, suggesting that dietary PLs promote lipid absorption and transport. Furthermore, dietary SL at 1% level could enhance non-specific immune capacity, with serum total protein level being markedly higher than that in the CT group. In conclusion, it is speculated that the promotion of glucose utilization and appetite by 2% dietary SL could be linked. We suggest a 1.91% supplementation of SL in the diet for the best growth performance in juvenile Chinese perch.

Regulatory effect of NK homeobox 1 (NKX2.1) on melanocortin 4 receptor (Mc4r) promoter in Mandarin fish.

The melanocortin 4 receptor (MC4R) is a G protein-coupled transporter that mediates the regulation of thyroid hormones and leptin on energy balance and food intake. However, the mechanisms of transcriptional regulation of Mc4r by thyroid hormone and leptin in fish have been rarely reported. The messenger RNA expression of Mc4r gene was significantly higher in brain than those in other tissues of mandarin fish. We analyzed the structure and function of a 2029 bp sequence of Mc4r promoter. Meanwhile, overexpression of NKX2.1 and incubation with leptin significantly increased Mc4r promoter activity, but triiodothyronine showed the opposite effect. In addition, mutations in the NKX2.1 binding site abolished not only the activation of Mc4r promoter activity by leptin but also the inhibitory effect of thyroid hormones on Mc4r promoter activity. In summary, these results suggested that thyroid hormones and leptin might regulate the transcriptional expression of Mc4r through NKX2.1.

Dietary zinc levels affect growth, appetite, and lipid metabolism of Chinese perch (Siniperca chuatsi).

An 84-day feeding experiment was conducted to investigate the effects of dietary Zn (zinc) on growth performance, food intake, and lipid metabolism of Chinese perch (Siniperca chuatsi). Five isonitrogenous and isolipidic diets with differential Zn contents (67, 100, 149, 230, and 410 mg/kg) were fed to 270 fish (35.47 ± 0.49 g). Results showed that fish growth and food intake increased markedly with the dietary 149 mg/kg Zn levels. Meanwhile, the food intake of 149 mg/kg group was significantly higher than that of other treatment groups after feeding for 8 weeks (P < 0.05). The qRT-PCR results showed that the expression of center appetite regulation factors in the hypothalamus was significantly regulated, and 149 mg/kg significantly increased mRNA expression of npy (neuropeptide Y) and decreased pomc (anorexigenic proopiomelanocortin) and cart (cocaine- and amphetamine-regulated transcript) gene expression. Meanwhile, the expressions of the main genes (such as leptin A and ghrelin) involved in peripheral appetite regulation factors were significantly up-regulated firstly and then reduced with the dietary Zn level increased, whereas the expression of cck (cholecystokinin) was significantly up-regulated. Serum AST (aspartate transaminase) and ALT (alanine transaminase) activities in fish fed the diets containing 230 and 410 mg/kg were significantly higher than that in other groups (P < 0.05). The lipid content of liver in 67 and 100 mg/kg groups was significantly higher than other groups (P < 0.05). Furthermore, dietary Zn significantly elevated the serum TG (triglyceride) and TCHO (total cholesterol) content levels (P < 0.05). Fish fed a high Zn diet (149, 230, and 410 mg/kg) dramatically down-regulated expression of srebp1 (sterol regulatory element binding proteins1c) and fas (fatty acid synthetase), but up-regulated expression of pparα (peroxisome proliferators-activated receptor-α) and cpt1 (carnitine palmitoyl transferase I) in the liver. The optimal dietary Zn inclusion level ranged from 146.69 to 152.86 mg/kg diet, based on two-slope broken-line regression analysis of WGR (weight gain rate) and FCR (feed conversion rate) for Chinese perch.

Altered Visual Function in Short-Wave-Sensitive 1 (sws1) Gene Knockout Japanese Medaka (Oryzias latipes) Larvae.

Visual perception plays a crucial role in foraging, avoiding predators, mate selection, and communication. The regulation of color vision is largely dependent on opsin, which is the first step in the formation of the visual transduction cascade in photoreceptor cells. Short-wave-sensitive 1 (sws1) is a visual pigment that mediates short-wavelength light transduction in vertebrates. The depletion of sws1 resulted in increased M-opsin in mice. However, there is still no report on the visual function of sws1 in teleost fish. Here, we constructed the sws1 knockout medaka using CRISPR/Cas9 technology. The 6 dph (days post-hatching) medaka sws1-/- larvae exhibited significantly decreased food intake and total length at the first feeding stage, and the mRNA levels of orexigenic genes (npy and agrp) were significantly upregulated after feeding. The swimming speed was significantly reduced during the period of dark-light transition stimulation in the sws1-mutant larvae. Histological analysis showed that the thickness of the lens was reduced, whereas the thickness of the ganglion cell layer (GCL) was significantly increased in sws1-/- medaka larvae. Additionally, the deletion of sws1 decreased the mRNA levels of genes involved in phototransduction (gnb3b, grk7a, grk7b, and pde6c). We also observed increased retinal cell apoptosis and oxidative stress in sws1 knockout medaka larvae. Collectively, these results suggest that sws1 deficiency in medaka larvae may impair visual function and cause retinal cell apoptosis, which is associated with the downregulation of photoconduction expression and oxidative stress.

Dietary supplementation of pyridoxine can enhance the growth performance and improve the protein, lipid utilization efficiency of mandarin fish (Siniperca chuatsi).

This study aimed to assess the effect of pyridoxine supplementation in the mandarin fish diet on growth performance, protein and lipid metabolism, and liver and intestinal histology. Mandarin fish were fed six diets with different levels of pyridoxine (2.67 mg/kg (control), 4.41 mg/kg, 6.57 mg/kg, 10.25 mg/kg, 17.93 mg/kg, 33.12 mg/kg diet) for 8 weeks, and samples were collected for analysis. The findings demonstrated that feeding mandarin fish a diet with 6.57 mg/kg pyridoxine led to a significant increase in weight gain rate (WGR), protein efficiency ratio (PER), whole-body crude protein, whole-body crude lipid, serum protein, cholesterol (CHO), triacylglycerol (TG), high-density lipoprotein-cholesterol (HDL-C), low-density lipoprotein-cholesterol (LDL-C), and alkaline phosphatase (ALP), as well as significantly lower serum glucose (GLU) and feed conversion ratio (FCR), compared to the control group (P < 0.05). Furthermore, we found a significant upregulation of the relative expression of genes associated with hepatic lipid oxidation and synthesis (hl, lpl, pparα, cpt1, cs, srebp1, and fas) and proteolysis (ast, alt, and gdh) in fish fed a diet containing 6.57 mg/kg pyridoxine (P < 0.05). Regarding the histological analysis, we observed a notable decrease in the quantity of intestinal mucus-secreting cells when the fish fed a diet containing 10.25 mg/kg pyridoxine (P < 0.05). These findings suggest that dietary pyridoxine supplementation promotes mandarin fish growth by improving the efficiency of protein and lipid utilization. Additionally, we used a broken-line regression analysis to estimate the optimal dietary pyridoxine requirement for mandarin fish in the range of 6.17-6.41 mg/kg based on WGR, FCR, and PER.

Role of short-wave-sensitive 1 (sws1) in cone development and first feeding in larval zebrafish.

Color vision is mediated by the expression of different major visual pigment proteins (opsins) on retinal photoreceptors. Vertebrates have four classes of cone opsins that are most sensitive to different wavelengths of light: short wavelength sensitive 1 (SWS1), short wavelength sensitive 2 (SWS2), medium wavelength sensitive (RH2), and long wavelength sensitive (LWS). UV wavelengths play important roles in foraging and communication. However, direct evidence provide links between sws1 and first feeding is lacking. Here, CRISPR/Cas9 technology was performed to generate mutant zebrafish lines with sws1 deletion. sws1 mutant zebrafish larvae exhibited decreased sws1, rh2-2, and lws1 expression, and increased rod gene (rho and gnat1) expression. Furthermore, the sws1-deficient larvae exhibited significantly reduced food intake, and the orexigenic genes npy and agrp signaling were upregulated at 6 days postfertilization (dpf). The transcription expression of sws1 and rh2-3 genes decreased in sws1-/- adults compared to wild type. Surprisingly, the results of feeding at the adult stage were not the same with larvae. sws1 deficiency did not affect food intake and appetite gene expression at adult stages. These results reveal a role for sws1 in normal cone development and first feeding in larval zebrafish.

Effects and regulatory pathway of proopinmelanocortin on feeding habit domestication in mandarin fish.

Proopiomelanocortin (POMC) is a hormone precursor, and has been reported to participate in domestication. However, its effects on feeding habit domestication in fish are poorly understood. Mandarin fish (Siniperca chuatsi) feeds solely on live prey fish since first-feeding. In the present study, the high expression of pomc in mandarin fish, both the pomc siRNA and MC4R inhibitor treatments increased the success rate of domestication from live prey fish to dead prey fish and food intake of dead prey fish, suggesting the role of pomc on the special feeding habit of live prey fish in mandarin fish. In addition, one c-fos binding site was identified in the region that from -1053 bp to -931 bp upstream of the transcription start site of pomc, and this region exhibited positive promoter activity. The mandarin fish brain cells treated with c-fos siRNA displayed suppressed pomc mRNA expression, indicating that c-fos positively regulated pomc expression. Furthermore, the mRNA expression of c-fos was higher in the mandarin fish which were more difficult to domesticate. The results of ChIP assay and inhibitor treatment confirmed that the activation of c-fos gene by histone H3K4me3 was catalyzed by Setd1b in mandarin fish. Three open peaks were found at the upstream regulatory region of setd1b by ATAC-seq, and the mRNA expression of setd1b was higher in the mandarin fish which were more difficult to domesticate. These results indicated that Setd1b could methylate histone H3K4 to activate the c-fos transcription, maintaining the high expression of pomc, which might contribute to the special feeding habit of mandarin fish.

Knockout of sws2a and sws2b in Medaka (Oryzias latipes) Reveals Their Roles in Regulating Vision-Guided Behavior and Eye Development.

The medaka (Oryzias latipes) is an excellent vertebrate model for studying the development of the retina. Its genome database is complete, and the number of opsin genes is relatively small compared to zebrafish. Short wavelength sensitive 2 (sws2), a G-protein-coupled receptor expressed in the retina, has been lost in mammals, but its role in eye development in fish is still poorly understood. In this study, we established a sws2a and sws2b knockout medaka model by CRISPR/Cas9 technology. We discovered that medaka sws2a and sws2b are mainly expressed in the eyes and may be regulated by growth differentiation factor 6a (gdf6a). Compared with the WT, sws2a-/- and sws2b-/- mutant larvae displayed an increase in swimming speed during the changes from light to dark. We also observed that sws2a-/- and sws2b-/- larvae both swam faster than WT in the first 10 s of the 2 min light period. The enhanced vision-guided behavior in sws2a-/- and sws2b-/- medaka larvae may be related to the upregulation of phototransduction-related genes. Additionally, we also found that sws2b affects the expression of eye development genes, while sws2a is unaffected. Together, these findings indicate that sws2a and sws2b knockouts increase vision-guided behavior and phototransduction, but on the other hand, sws2b plays an important role in regulating eye development genes. This study provides data for further understanding of the role of sws2a and sws2b in medaka retina development.

Role of phosphoenolpyruvate carboxykinase 1 (pck1) in mediating nutrient metabolism in zebrafish.

Carbohydrates are the most economical source of energy in fish feeds, but most fish have limited ability to utilize carbohydrates. It has been reported that phosphoenolpyruvate carboxykinase 1 (pck1) is involved in carbohydrate metabolism, lipid metabolism, and other metabolic processes. However, direct evidence is lacking to fully understand the relationship between pck1 and glucose and lipid metabolism. Here, we generated a pck1 knockout zebrafish by CRISPR/cas9 system, and a high-carbohydrate diet was provided to 60 days post-fertilization (dpf) for 8 weeks. We found that pck1-deficient zebrafish displayed decreased plasma glucose, elevated mRNA levels of glycolysis-related genes (gck, pfk, pk), and reduced the transcriptional levels of gluconeogenic genes (pck1, fbp1a) in liver. We also found decreased triglyceride, total cholesterol, and lipid accumulation and in pck1-/- zebrafish, along with downregulation of genes for lipolysis (acaca) and lipogenesis (cpt1). In addition, the observation of HE staining revealed that the total muscle area of pck1-/- was substantially less than that of WT zebrafish and real-time PCR suggested that GH/IGF-1 signaling (ulk2, stat1b) may be suppressed in pck1-deficient fish. Taken together, these findings suggested that pck1 may play an important role in the high-carbohydrate diet utilization of fish and significantly affected lipid metabolism and protein synthesis in zebrafish. pck1 knockout mutant line could facilitate a further mechanism study of pck1-associated metabolic regulation and provide new information for improving carbohydrate utilization traits.

DNA methylation of pck1 might contribute to the programming effects of early high-carbohydrate diets feeding to the glucose metabolism across two generations in zebrafish (Danio rerio).

The purpose of this study is to assess the effects of early high-carbohydrate stimulus on glucose metabolism in zebrafish (Danio rerio) over two generations and explore the mechanisms that explain those nutritional programming effects via epigenetic modifications. The larvae were delivered a high-carbohydrate diet (53.66%) that was used as an early nutritional stimulus from the first feeding to the end of the yolk sac (FF) and 5 days after yolk-sac exhaustion (YE). The larvae (F0) and their offspring (F1) were then both fed the control diet (22.69%) until adulthood (15 weeks), and they were challenged with a high-carbohydrate diet (35.36%) at the 16th week. The results indicated that early stimulus immediately raised the mRNA levels of genes involved in glycolysis and gluconeogenesis. At the end of F0 challenge, both treatment groups decreased the plasma glucose levels, increased the expression levels of glucokinase (gck), and inhibited the mRNA during gluconeogenesis. When challenged in F1, the glucose levels were lower in FF (F1), and the mRNA levels of phosphoenolpyruvate carboxykinase 1 (pck1) were decreased in FF (F1) and YE (F1). Besides, in both experimental groups (F0 and F1), the CpG island of pck1 maintained lower levels of hypermethylated expression from F0 adult, 24 h post-fertilization embryo, to F1 adult. In conclusion, these results indicated that an early high-carbohydrate stimulus could significantly reprogram glucose metabolism in adult zebrafish, that those modifications could be partially transmitted to the next generation, and that the DNA methylation of pck1 might work as a stable epigenetic marker to contribute to those processes.

Comparative Study of the Molecular Characterization, Evolution, and Structure Modeling of Digestive Lipase Genes Reveals the Different Evolutionary Selection Between Mammals and Fishes.

Vertebrates need suitable lipases to digest lipids for the requirement of energy and essential nutrients; however, the main digestive lipase genes of fishes have certain controversies. In this study, two types of digestive lipase genes (pancreatic lipase (pl) and bile salt-activated lipase (bsal)) were identified in mammals and fishes. The neighborhood genes and key active sites of the two lipase genes were conserved in mammals and fishes. Three copies of PL genes were found in mammals, but only one copy of the pl gene was found in most of the fish species, and the pl gene was even completely absent in some fish species (e.g., zebrafish, medaka, and common carp). Additionally, the hydrophobic amino acid residues (Ile and Leu) which are important to pancreatic lipase activity were also absent in most of the fish species. The PL was the main digestive lipase gene in mammals, but the pl gene seemed not to be the main digestive lipase gene in fish due to the absence of the pl gene sequence and the important amino acid residues. In contrast, the bsal gene existed in all fish species, even two to five copies of bsal genes were found in most of the fishes, but only one copy of the BSAL gene was found in mammals. The amino acid residues of bile salt-binding sites and the three-dimensional (3D) structure modeling of Bsal proteins were conserved in most of the fish species, so bsal might be the main digestive lipase gene in fish. The phylogenetic analysis also indicated that pl or bsal showed an independent evolution between mammals and fishes. Therefore, we inferred that the evolutionary selection of the main digestive lipase genes diverged into two types between mammals and fishes. These findings will provide valuable evidence for the study of lipid digestion in fish.

Lysine Deprivation Regulates Npy Expression via GCN2 Signaling Pathway in Mandarin Fish (Siniperca chuatsi).

Regulation of food intake is associated with nutrient-sensing systems and the expression of appetite neuropeptides. Nutrient-sensing systems generate the capacity to sense nutrient availability to maintain energy and metabolism homeostasis. Appetite neuropeptides are prominent factors that are essential for regulating the appetite to adapt energy status. However, the link between the expression of appetite neuropeptides and nutrient-sensing systems remains debatable in carnivorous fish. Here, with intracerebroventricular (ICV) administration of six essential amino acids (lysine, methionine, tryptophan, arginine, phenylalanine, or threonine) performed in mandarin fish (Siniperca chuatsi), we found that lysine and methionine are the feeding-stimulating amino acids other than the reported valine, and found a key appetite neuropeptide, neuropeptide Y (NPY), mainly contributes to the regulatory role of the essential amino acids on food intake. With the brain cells of mandarin fish cultured in essential amino acid deleted medium (lysine, methionine, histidine, valine, or leucine), we showed that only lysine deprivation activated the general control nonderepressible 2 (GCN2) signaling pathway, elevated α subunit of eukaryotic translation initiation factor 2 (eIF2α) phosphorylation, increased activating transcription factor 4 (ATF4) protein expression, and finally induced transcription of npy. Furthermore, pharmacological inhibition of GCN2 and eIF2α phosphorylation signaling by GCN2iB or ISRIB, effectively blocked the transcriptional induction of npy in lysine deprivation. Overall, these findings could provide a better understanding of the GCN2 signaling pathway involved in food intake control by amino acids.

Genome-wide identification and expression patterns of opsin genes during larval development in Chinese perch (Siniperca chuatsi).

Vision is important for fish to forage food and fishes express opsin genes to receive visual signals. Chinese perch (Siniperca chuatsi) larvae prey on other fish species larvae at firstfeeding but donoteat any zooplankton, the expression of opsin genes in S. chuatsilarvae is unknown. In this study, we conducted a whole-genome analysis and demonstrated that S. chuatsihave5cone opsin genes (sws1, sws2Aα, sws2Aß, rh2and lws)and 2 rod opsin genes (rh1and rh1-exorh). The syntenicanalysisshowedthe flanking genes ofall opsin genes were conserved during fish evolution, but the ancestorof S. chuatsimightlost some opsin gene copies duringtheevolution.The phylogeneticanalysisshowed sws1of S. chuatsiwas closest to those of Lates calcariferwhich had a truncated sws1gene; the sws2Aα, sws2Aß,lws,rh2,rh1 andrh1-exorh of S. chuatsihad a closer relationship with those of Percomorpha fishes.Importantly, results of in situhybridization showed the sws1 opsingene,which is related to forage zooplankton,had extremely low levelexpression in retinaat early stages.Surprisingly, the rh2 opsin gene had a high level expression at firstfeeding stage. The sws2Aα, sws2Aßand lwshad a little expression at early stages but the lwsshowed a increasing trend with larval development, rh1 opsin gene expression appeared at15 dph. In thisstudy, we found a specialpattern of visual opsin genes expression in S. chuatsi, it might influence the larval first feeding and feeding habit.

Dietary bile acids reduce liver lipid deposition via activating farnesoid X receptor, and improve gut health by regulating gut microbiota in Chinese perch (Siniperca chuatsi).

The aim of this study is to explore the effects of dietary bile acids (BAs) supplementation on lipid metabolism and gut health of Chinese perch (Siniperca chuatsi), and its possible mechanisms. Two isonitrogenous and isolipidic diets were formulated to supplement different levels of BAs (0 and 900 mg BAs kg-1 diet, respectively). All fish (Initial mean body weight: 171.29 ± 0.77g) were randomly divided into 2 groups (triplicate, 54 fish/group) and were fed with different experimental diets for 56 days, respectively. Dietary exogenous BAs supplementation at the concentration of 900 mg kg-1 significantly increased weight gain and survival rate, and decreased feed conversion ratio. BAs could inhibit lipid synthesis and promote lipid oxidation to reduce lipid deposition by activating farnesoid X receptor (FXR). Dietary BAs supplementation increased the abundance of Lactobacilli in Firmicutes, and the increase of Lactobacillus caused the increase of lactic acid level and the decrease of pH, which might be the reason for the gut villus length and gut wall high in this study. Dietary BAs supplementation increased the levels of catalase and superoxide dismutase and decreased the level of malondialdehyde in the gut and plasma, which might be contributed to the regulating the antioxidant stress phenotype of gut microbiota by the increased abundance of Firmicutes. Then it caused the increase of the globulin level in the plasma, meaning the enhancement of immune state. The increased immunity might also be thought to be responsible for increased survival rate. These results suggest dietary BAs reduce liver lipid deposition via activating FXR, and improve gut health by regulating gut microbiota in Chinese perch.