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Jixuan Yang Qingsong Tan Wenhuan Zhu Chen Chen Xufang Liang Lei Pan 《Fish physiology and biochemistry》2014,40(1):93-104
The solute carrier family 7A, member 7 gene encodes the light chain- y+L amino acid transporter-1 (y+LAT1) of the heterodimeric carrier responsible for cationic amino acid (CAA) transport across the basolateral membranes of epithelial cells in intestine and kidney. Rising attention has been given to y+LAT1 involved in CAA metabolic pathways and growth control. The molecular characterization and function analysis of y+LAT1 in grass carp (Ctenopharyngodon idellus) is currently unknown. In the present study, full-length cDNA (2,688 bp), which encodes y+LAT1 and contains a 5′-untranslated region (319 bp), an open reading frame (1,506 bp) and a 3′-untranslated region (863 bp), has been cloned from grass carp. Amino acid sequence of grass carp y+LAT1 contains 11 transmembrane domains and shows 95 %, 80 % and 75 % sequence similarity to zebra fish, amphibian and mammalian y+LAT1, respectively. The tissue distribution and expression regulation by fasting of y+LAT1 mRNA were analyzed using real-time PCR. Our results showed that y+LAT1 mRNA was highly expressed in midgut, foregut and spleen while weakly expressed in hindgut, kidney, gill, brain, heart, liver and muscle. Nutritional status significantly influenced y+LAT1 mRNA expression in fish tissues, such as down-regulation of y+LAT1 mRNA expression after fasting (14 days). 相似文献
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ghrelin是一种在脊椎动物摄食调节过程中起重要作用的脑肠肽,具有明显的摄食促进作用。实验利用同源克隆技术获得了草鱼ghrelin基因的cDNA序列和DNA序列,其中cDNA序列全长506 bp,包括90 bp的5′端非编码区(5′-untranslated region,5′UTR),312 bp的开放阅读框(open reading frame,ORF),以及104 bp的3′端非编码区(3′-untranslated region,3′UTR)。开放阅读框编码的103个氨基酸的ghrelin前体肽,经剪切加工后形成含有19个氨基酸的成熟肽。氨基酸序列分析结果显示,草鱼ghrelin与硬骨鱼类ghrelin相似度最高,而与其他脊椎动物相似度较低,同时草鱼ghrelin成熟肽N端的"活性中心"(active core)为鲤科鱼类中常见的GTSF形式。与大多数硬骨鱼类的ghrelin基因结构相同,草鱼ghrelin基因也包括4个外显子和3个内含子。荧光定量PCR检测到ghrelin mRNA大量分布于草鱼的前肠和脾,脑、肾、肝、肌肉、皮和鳔等组织也有ghrelin mRNA分布。草鱼脑和肠中的ghrelin表达水平在摄食后下降,随着饥饿时间的延长表达水平逐步升高,最后维持在较高水平,表明ghrelin作为摄食启动信号对草鱼的摄食活动起到了促进作用。 相似文献
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固醇调节元件结合蛋白(SREBPs)是调控糖脂代谢相关基因表达的关键核转录因子。为获知草鱼SREBP-1基因的序列及其在肝脏中的表达规律,本实验采用同源克隆和RACE方法获得了草鱼SREBP-1基因的部分cDNA序列,并通过生物信息学方法对该基因及所编码蛋白的结构特征进行了分析;采用实时荧光定量PCR技术,对SREBP-1基因在8种不同组织的表达规律及低糖(糖含量24%)和高糖(糖含量42%)投喂条件下肝脏中的表达水平进行了研究。结果显示,所克隆到的草鱼SREBP-1基因cDNA长4 760 bp,其中包括开放读码框3 426bp,编码1 141个氨基酸;草鱼SREBP-1具有一个典型的碱性螺旋-环-螺旋亮氨酸拉链结构(bHLH-zip);氨基酸序列比对结果显示,草鱼SREBP-1与其他鱼类的同源性在76%~88%之间,与斑马鱼的进化关系最近;草鱼SREBP-1基因在脑中的表达量最高,肝脏和肠次之,在肾脏、脾脏、肌肉、脂肪和性腺中均有少量表达;与对照组相比,SREBP-1基因在高糖诱导下表达量显著提高(P0.05),低糖诱导下没有显著性差异。研究表明,在高糖负荷条件下,草鱼肝脏中SREBP-1可能会促进糖的利用和转化,从而参与糖代谢调节过程,为丰富鱼类糖代谢调控机理提供研究资料,并有望为提高鱼类对饲料糖的利用效率提供理论依据。 相似文献
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Hao Wang Libo He Yongyan Pei Pengfei Chu Rong Huang Yongming Li Lanjie Liao Zuoyan Zhu Yaping Wang 《Fish physiology and biochemistry》2016,42(5):1369-1382
Multidomain proapoptotic Bcl-2-associated X (Bax) protein is an essential effector responsible for mitochondrial outer membrane permeabilization, resulting in cell death via apoptosis. In this study, two Bax genes of grass carp (Ctenopharyngodon idellus), designated as CiBax1 and CiBax2, were isolated and analyzed. The obtained CiBax1 cDNA is 2058 bp long, with a 579 bp open reading frame (ORF) coding a protein of 192 amino acid residues. The full-length cDNA of CiBax2 is 1161 bp, with a 618 bp ORF coding 205 amino acids. Both CiBax1 and CiBax2 are typical members of Bcl-2 family containing conserved Bcl and C-terminal domains, and they share conserved synteny with zebrafish Bax genes despite the grass carp Bax mapping to different linkage groups. Phylogenetic analysis showed that CiBax1 was clustered with Bax from most teleost fish, and CiBax2 was close to Bax2 from teleost fish but far separated from that of Salmo salar. Quantitative real-time PCR analysis revealed broad expression of CiBax1 and CiBax2 in tissues from healthy grass carp, but the relative expression level differed. The mRNA expression of CiBax1 and CiBax2 was both upregulated significantly and peaked in all examined tissues at days 5 or 6 post-infection with grass carp reovirus. Subcellular localization indicated that CiBax1 protein was localized in both nucleus and cytosol, while CiBax2 protein only in cytosol. Moreover, CiBax2, but not CiBax1 was colocalized with mitochondrion under normal condition. Taken together, the findings would be helpful for further understanding of the function of Bax in teleost fish. 相似文献
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Ke Feng Gui-rong Zhang Kai-jian Wei Bang-xi Xiong Tao Liang Hai-chao Ping 《Fish physiology and biochemistry》2012,38(6):1825-1834
Cholecystokinin (CCK) is a multi-functional brain–gut peptide in fish and mammals. To investigate the role of CCK in appetite regulation in fish, a 770-bp full-length cDNA sequence of CCK gene was obtained by RT-PCR and rapid amplification of cDNA ends methods in grass carp Ctenopharyngodon idellus. Homology analysis showed that the CCK cDNA sequence of grass carp had the highest similarity (90 %) to that of goldfish Carassius auratus and a higher similarity (>70 %) to those of other teleosts than to mammals. The PCR amplification using genomic DNA identified that the CCK gene of grass carp was comprised of three exons and two introns. Real-time quantitative PCR was used to detect CCK mRNA expression in adult tissues. High levels of gene expression were found in the hypothalamus and pituitary; moderate levels in the intestine, muscle and white adipose tissue; and low levels in other tissues. During early development (i.e., fertilized eggs to 35-day post-hatching larvae) the levels of CCK mRNA expression were higher during embryonic developmental stages than during post-hatch larval stages. Fasting decreased CCK mRNA expression levels in the brain and intestine, whereas refeeding resulted in an increase of expression. The results suggest that CCK mRNA expression has obvious tissue specificity and may have a role in feed intake regulation in grass carp. 相似文献
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S. L. Hsieh H. T. Chang C. H. Wu C. M. Kuo 《Aquaculture (Amsterdam, Netherlands)》2004,230(1-4):527-546
The stearoyl-CoA desaturase cDNA in tilapia (Oreochromis mossambicus) was cloned by RT-PCR and RACE, and it was compared with those in grass carp, common carp and milkfish. Nucleotide sequence analysis revealed that the full length of cDNA (1172 bp) clone encompasses 1008 bp open reading frame (ORF) encoding 336 amino acid residues. The deduced amino acid sequence shares 78–82% identity with the teleosts and 64–66% with mammals compared, and like these fish, the cloned tilapia stearoyl-CoA desaturase amino acid sequence conserves three histidine cluster motifs (one HXXXXH and two HXXHH), which functioned as non-heme iron binding sites, essential for stearoyl-CoA desaturase activity. RT-PCR and Northern blot analysis reveal that tilapia stearoyl-CoA desaturase is expressed only in liver, but the stearoyl-CoA desaturase expression in multiple tissues was observed in milkfish, grass carp and carp. Further, the hormonal regulation of stearoyl-CoA desaturase gene expression was investigated by a single injection of 17β-estradiol and testosterone. The results showed that the administration of 17β-estradiol to tilapia led to a greater increase in desaturase activity than testosterone, and higher doses of steroids produced greater increases in enzyme activity. The comparative RT-PCR analysis showed that the stearoyl-CoA desaturase mRNA level increased significantly in 17β-estradiol treated animals, especially in the groups receiving a single injection of 50 mg 17β-estradiol. This was reflected in the decrease in the saturated fatty acids and the increase in the monounsaturated fatty acids. The proportion of the polyunsaturated fatty acids was not affected. 相似文献
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A full-length cDNA encoding the insulin-like growth factor binding protein-3 (IGFBP-3) was cloned from the liver of common carp (Cyprinus carpio) by RT-PCR. The IGFBP-3 cDNA sequence is 1,680 bp long and has an open reading frame of 882 bp encoding a predicted polypeptide of 293 amino acid residues. The deduced amino acid sequence contains a putative signal peptide of 25 amino acid residues resulting in a mature protein of 268 amino acids. A single band of approximate 1.9 kb was found in liver by Northern blot analysis. IGFBP-3 mRNA was observed in all regions of brain with high levels. In peripheral tissues, high levels of IGFBP-3 mRNA were found in retina, red muscle, liver, heart, posterior intestine, spleen, and testis. Relatively lower levels were found in white muscle, kidney, thymus gland, and ovary, while in head kidney, blood, skin, gill, middle intestine, and anterior intestine, the IGFBP-3 mRNA levels were much lower. IGFBP-3 mRNA was first detected in the blastula stage with significantly high level. The level sharply decreased in gastrula stage, and it became to increase in the following stages. During the reproductive cycle, the abundance of IGFBP-3 mRNA significantly decreased between the recrudescing stage and the matured stage in ovary, although in testis, IGFBP-3 mRNA expression level did not exhibit a significant change. The mRNA expression profiles in the present study imply that the IGFBP-3 may play important physiological functions in common carp development and reproduction. 相似文献
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实验获得了草鱼脂肪组织甘油三酯水解酶(ATGL) 部分cDNA序列(GenBank 登录号为HQ845211), 并进行了序列同源性分析; 采用实时定量反转录聚合酶链式反应(qRT-PCR)方法, 检测了ATGL基因在草鱼不同组织的表达状况; 研究了投喂n-3高不饱和脂肪酸(n-3 HUFAs)对草鱼肝胰脏ATGL基因时序表达的影响。结果显示, 所获得的草鱼ATGL基因部分cDNA序列长度为687 bp, 与人、牛、小鼠、长腭泥虎鱼、大黄鱼等物种的同源性为65%~75%; 该基因在草鱼心脏、肝胰脏、脾脏、鳃、肾脏、肌肉、腹腔脂肪组织、脑、小肠、精巢10个组织中均有表达, 其中在腹腔脂肪组织中表达丰度最高, 在肝胰脏和肌肉中表达丰度次之。处理组草鱼摄食n-3 HUFAs饲料后, ATGL基因的表达水平在第1周和第2周显著高于对照组, 第3周后, 该基因的表达水平在处理组与对照组间无显著差异。研究首次克隆得到草鱼ATGL基因部分cDNA序列, 并发现该基因在草鱼脂质蓄积及代谢较旺盛的组织中表达水平较高, 且其在肝胰脏中的表达受到n-3 HUFAs的影响, 其规律为先被诱导升高, 然后回复到正常水平。 相似文献
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Molecular Characterization and Dietary Regulation of the AlaSerCys Transporter 2 in Grass Carp,Ctenopharyngodon idella 
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下载免费PDF全文 Rong Hu Yonghong Wang Fufa Qu Jianzhou Tang Yi Zhou Shuangqing Lu Jianshe Zhang Zhen Liu Zhigang Zhou Xinhong Guo 《Journal of the World Aquaculture Society》2017,48(2):333-341
AlaSerCys Transporter 2 (ASCT2), a glutamine/amino acid transporter, plays an important role in the absorption of neutral amino acids. In this study, we cloned the full‐length cDNA of ASCT2 from grass carp, Ctenopharyngodon idella, which encodes a 541‐amino‐acid protein. Phylogenetic analysis revealed that the ASCT2 sequence of grass carp clustered with the ASCT2 from Danio rerio. ASCT2 mRNA was differentially expressed in different tissues, with a gradient expression from high to low in tissues of the liver, gill, muscle, midgut, hypophysis, hindgut, kidney, heart, foregut, brain, spleen, and gonad. Meanwhile, the quantitative real‐time polymerase chain reaction results showed that 22% crude protein diets could significantly stimulate ASCT2 gene expression. In addition, ASCT2 was expressed in higher quantities in the soybean meal group than in the fishmeal group. Furthermore, 7.5 g/kg glutamine and 2.5 g/kg glutamine dipeptide remarkably improved the expression of ASCT2. These results suggested that ASCT2 expression is regulated by the dietary protein levels and sources, as well as appropriate concentration of glutamine and glutamine dipeptide additives. 相似文献
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在草鱼中新发现的两种免疫球蛋白重链的cDNA和基因组序列,其中的一种IgZ命名为IgZ-2,以区别于已报道的IgZ,另一种只有两个恒定区,一个恒定区与IgM相似而另一个与IgZ相似,这一特征与已报道的鲤的IgM-IgZ相似,故同样称为IgM-IgZ。分泌型IgZ-2的cDNA序列包含1889bp,编码539个氨基酸,其3'编码区包含267bp,但缺乏5'非编码区及部分可变区序列。分泌型IgM-IgZ的cDNA全长为1316bp,编码361个氨基酸,其5'非编码区包含3bp,3'非编码区包含227bp。膜结合型IgM-IgZ由两个膜外显子与CH2中的一个剪切位点剪切而成。氨基酸序列比对结果显示IgZ-2和IgM-IgZ的恒定区存在保守的半胱氨酸。系统进化树分析显示,草鱼IgZ-2以较高的支持率与斑马鱼IgZ聚为一枝,再与草鱼IgZ、草鱼IgM-IgZ和鲤IgM-IgZ这一枝聚为一类。用半定量RT-PCR检测IgZ-2和IgM-IgZ在4条草鱼的器官/组织中的表达,发现分泌型IgZ-2、分泌型IgM-IgZ和膜结合型IgM-IgZ在4条鱼中的表达存在个体差异,但主要都在免疫器官中表达。 相似文献
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为探讨Adipo R1-B在鱼类糖类和脂质代谢中的作用,本实验采用RACE方法获得了草鱼Adipo R1-B的全长c DNA序列(登录号:KP733846),利用生物信息学技术对该基因及其所编码蛋白的结构特征进行了分析;采用实时定量PCR技术,检测了Adipo R1-B在19个不同组织中的表达特性以及高糖(45%)、高脂(8%)和高糖高脂饲喂对草鱼肝脏中该基因表达的影响。结果显示,草鱼Adipo R1-B c DNA全长2186 bp,其中开放读码框为1122 bp,编码373个氨基酸;跨膜结构分析表明草鱼Adipo R1-B为典型的7次跨膜蛋白;同源性分析结果显示,草鱼Adipo R1-B与其他物种的Adipo R1-B高度同源(氨基酸相似度78%以上),并与斑马鱼Adipo R1-B的进化关系最近;组织分布结果显示,Adipo R1-B在草鱼肝脏中表达量最高,中枢神经系统和红肌次之;此外,高脂和高糖高脂饲喂均能够显著提高草鱼肝脏中Adipo R1-B的表达水平。因此,草鱼肝脏中Adipo R1-B的表达水平受到日粮中脂类水平的调控,推测该受体可能在调节鱼类脂质代谢中发挥重要作用。 相似文献
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This study was conducted to assess the effect of eicosapentaenoic acid (EPA) on grass carp preadipocyte glycerol kinase (GyK) expression, as well as to explore the mechanism. Here, we cloned partial sequence of grass carp GyK gene and analyzed its tissue distribution. The result showed that GyK gene expressed most in the liver, followed by adipose tissue and the kidney. Besides, 400 μM oleic acid (18:1n-9, OA) was used to establish a hypertrophic preadipocyte model. GyK gene expression and enzyme activity were significantly enhanced after model cells were treated with 100 μM eicosapentaenoic acid (20:5n-3, EPA) for 6, 12, and 24 h. Meanwhile, peroxisome proliferative-activated receptor (PPAR)γ, adipose triglyceride lipase (ATGL), and the two isoforms of grass carp HSL gene were first identified by Sun et al (2016), and they defined the two isoforms as HSLa and HSLb. Therefore, maybe HSLa and HSLb are appropriate.. The content of triglyceride was dramatically increased by EPA treatment for 24 h. Further, a competitive ATGL antagonist, HY-15859, attenuated the increase in GyK induced by EPA at 12 h. Surprisingly, the enhanced lipolysis and PPARγ gene expression induced by serum deprivation were paralleled by an increase in GyK gene expression, whereas a stabilization in GyK enzyme activity. Other fatty acids, including docosahexaenoic acid, alpha-linolenic acid, linoleic acid, and OA also promoted GyK gene expression. Moreover, an irreversible PPARγ antagonist, GW9662, was used to investigate the role of PPARγ in GyK induction. Data showed that GW9662 abolished the induction of GyK by EPA at 12 h. Together, these data suggested that EPA elevated grass carp preadipocytes GyK expression. ATGL and PPARγ contributed to the induction of GyK. PPARγ may be a key regulator in response to GyK expression induced by EPA. 相似文献
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本研究根据半滑舌鳎(Cynoglossus semilaevis)基因组数据库中预测的多聚免疫球蛋白受体(Polymeric immunoglobulin receptor, pIgR)序列,通过PCR和RACE技术获得了半滑舌鳎pIgR基因cDNA,全长为1419 bp,开放阅读框(ORF)为1020 bp,编码339个氨基酸,5′UTR区域为109 bp,3′UTR区域为290bp。保守结构域分析显示,半滑舌鳎pIgR蛋白包含1个信号肽,2个免疫球蛋白功能域(Ig-like domain, ILD)和1个跨膜结构域。经蛋白序列同源比对和系统进化树分析,发现半滑舌鳎pIgR与大菱鲆(Scophthalmus maximus)和牙鲆(Paralichthy solivaceus)的pIgR亲缘关系最近。实时荧光定量PCR分析显示,pIgR基因在健康半滑舌鳎的不同组织中均有表达,在鳃中表达较高,在肌肉中表达最低。经哈维氏弧菌(Vibrio harveyi)病原感染刺激后,pIgR基因在半滑舌鳎的5个组织(肝脏、脾脏、肾脏、肠和鳃)中均呈先上升后下降的趋势,其中,在脾脏和鳃中48 h达到最高值,在肝脏、肾脏和肠中72 h达到峰值。与内脏组织不同的是,pIgR基因在皮肤中呈一直上升的趋势。上述结果表明,pIgR基因在半滑舌鳎抵御哈维氏弧菌的免疫应答中发挥重要作用。 相似文献