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The occurrerce of phosphoenolpyruvate carboxykinase (pepck) in the pericarp of different grapevine genotypes and in grape leaves and developing seeds

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JOURNAL OF HORTICULTURAL SCIENCE & BIOTECHNOLOGY 2018年第5期93卷 456-465页

作者： Famiani, Franco Paoletti, Andrea Proietti, Primo Battistelli, Alberto Moscatello, Stefano Cruz-Castillo, Juan G. Walker, Robert P. Univ Perugia Dipartimento Sci Agr Alimentari & Ambientali Perugia Italy CNR Ist Biol Agroambientale & Forestale Porano TR Italy Univ Autonoma Chapingo Ctr Reg Univ Oriente Chapingo Veracruz Mexico

The occurrence of phosphoenolpyruvate carboxykinase (pepck) and pyruvate orthophosphate dikinase (PPDK) was determined in the ripe pericarps of several grape genotypes. Moreover, the abundance pattern of pepck during the development of grape berries was studied in a range of cultivars. In addition, the presence of pepck in grape leaves and the location/abundance of pepck in developing grape seeds were investigated. pepck was present in the pericarp of all the cultivars of Vitis vinifera studied and in that of the hybrid 420A (Vitis berlandieri x Vitis riparia). By contrast, PPDK was either not present or present at very low abundance. The results strongly indicated that the pepck and not the PPDK pathway is the predominant one utilised in gluconeogenesis in the berry pericarp of grape genotypes. Further, the results showed that the abundance of pepck increased greatly at the start of ripening in all the cultivars, showing a similar pattern in all of them. pepck was not present (or at very low abundance) in grape leaves throughout their development. In developing grape seeds, the results indicated a similar abundance of pepck in both the seed coat and the tissues enclosed by it.

关键词： Grape gluconeogenesis malate pepck PPDK

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Deacetylated nimbin analog N2 fortifies alloxan-induced pancreatic β-cell damage in insulin-resistant zebrafish larvae by upregulating phosphoenolpyruvate carboxykinase (pepck) and insulin levels

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TOXICOLOGY AND APPLIED PHARMACOLOGY 2022年 454卷 116229页

作者： Sudhakaran, Gokul Rajesh, Ravi Guru, Ajay Haridevamuthu, B. Murugan, Raghul Bhuvanesh, Nattamai Wadaan, Mohammad Ahmad Mahboob, Shalid Juliet, Annie Gopinath, Pushparathinam Arockiaraj, Jesu SRM Inst Sci & Technol Coll Sci & Humanities Dept Biotechnol Chennai 603203 Tamil Nadu India SRM Inst Sci & Technol Dept Chem Coll Engn & Technol Chennai 603203 Tamil Nadu India Texas A&M Univ Dept Chem College Stn TX 77842 USA King Saud Univ Dept Zool Coll Sci POB 2455 Riyadh 11451 Saudi Arabia Univ Texas Austin Inst Cellular & Mol Biol Univ Stn A4800 Austin TX 78712 USA

This study aims to evaluate the protective behaviour of N2, a semi-natural analog of nimbin, for its anti-diabetic efficacy against alloxan-induced oxidative damage and beta-cell dysfunction in in-vivo zebrafish larvae. A 500 mu M of alloxan was exposed to zebrafish larvae for 24 h to induce oxidative stress in the pancreatic beta-cells and co-exposed with N2 to study the protection of N2 by inhibiting ROS by DCFH-DA, DHE and NDA staining along with Cellular damage, apoptosis and lipid peroxidation. The zebrafish was further exposed to 500 mu M alloxan for 72 h to induce beta-cell destruction along with depleted glucose uptake and co-exposed to N2 to study the protective mechanism. Glucose levels were estimated, and PCR was used to verify the mRNA expression of phosphoenolpyruvate carboxykinase (pepck) and insulin. Alloxan induced (24 h) oxidative stress in the pancreatic beta-cells in which N2's co-exposure inhibited ROS by eliminating O-2(-) radicals and restoring the glutathione levels, thus preventing cellular damage and lipid peroxidation. The zebrafish exposed to 500 mu M alloxan for 72 h was observed with beta-cell destruction along with depleted glucose uptake when stained with 2NBDG, wherein N2 was able to protect the pancreatic beta-cells from oxidative damage, promoted high glucose uptake and reduced glucose levels. N2 stimulated insulin production and downregulated pepck by inhibiting gluconeogenesis, attenuating post-prandial hyperglycemia. N2 may contribute to anti-oxidant protection against alloxan-induced beta-cell damage and anti-hyperglycemic activity, restoring insulin function and suppressing pepck expression.

关键词： beta-cell destruction Diabetic Nimbin Insulin pepck

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紫菜叶状体遗传分析与pepck、PEPC在不同世代间的表达分析

紫菜叶状体遗传分析与PEPCK、PEPC在不同世代间的表达分析

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作者：何林文中国科学院研究生院（海洋研究所）

学位级别：博士

紫菜是重要的经济海藻，其生活史包括单倍的叶状体世代和双倍的丝状体世代。与高等植物不同的是，其孢子体世代为微观阶段，配子体世代为宏观阶段。紫菜的减数分裂发生在壳孢子萌发时期，分裂产生的四个细胞不分开，共同发育为一株紫菜... 详细信息

紫菜是重要的经济海藻，其生活史包括单倍的叶状体世代和双倍的丝状体世代。与高等植物不同的是，其孢子体世代为微观阶段，配子体世代为宏观阶段。紫菜的减数分裂发生在壳孢子萌发时期，分裂产生的四个细胞不分开，共同发育为一株紫菜，也就是说紫菜的叶状体是嵌合体。在使用嵌合的叶状体进行遗传分析时可能得到不准确的结果。另外，紫菜的叶状体固着生长于潮间带的石块上，随着潮水的涨落经历脱水和复水过程；而紫菜的丝状体钻入贝壳等碳酸钙基质中生长，不能脱离海水生存。这两个世代生活环境的差异可能导致其光合特性的不同。为此本文进行了如下研究： 1)以条斑紫菜为材料，利用AFLP标记对单个丝状藻丝接种贝壳释放出的売孢子发育来的叶状体和由随机选择的売孢子苗释放出的单孢子苗分别进行遗传分析。15对引物组合在40个壳孢子苗中扩增得到538个位点，其中多态性位点为434个，多态性比率为80.7%。对40个个体进行遗传相似性分析发现其遗传相似性系数最大为0.93，最小为0.47，平均为0.71。19对引物组合在88个单孢子苗中扩增得到708个位点，其中多态性位点为698个，多态性比率为98.6%。对88个个体进行遗传相似性分析发现其遗传相似性系数最大为0.78，最小为0.43，平均为0.61。聚类分析发现売孢子苗和单孢子苗个体均可分为明显的两大类。结果表明，单孢子苗中检测到的多态性比率高于壳孢子苗，单孢子苗个体间的相似性低于壳孢子苗个体间的相似性，因此使用单孢子苗进行遗传分析获得的结果比使用壳孢子苗更准确。 2)以坛紫菜丝状体（丝状藻丝）和叶状体为材料，以18S和GAPDH为内参基因，使用荧光定量PCR技术对pepck、PEPC在不同世代的差异进行分析；利用酶偶联法分别测定了叶状体和丝状体中PEPC、pepck羧化和脱羧的活性；利用pepck、PEPC的保守区序列，使用PMAL表达系统对其进行原核表达，并对蛋白表达温度及所需IPTG浓度进行优化，纯化并浓缩得到的蛋白，制备相应蛋白的抗体，利用得到的抗体分析叶状体和丝状体中pepck、PEPC含量的差异。结果表明，丝状体中的pepck、PEPC的mRNA含量、酶活以及蛋白含量圴高于叶状体。因此推测在坛紫菜丝状体和叶状体中可能存在不同的碳固定方式，坛紫菜丝状体中可能存在类C4途径。

关键词：紫菜 pepck PEPC RTQ-PCR C4

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Diurnal modulation of pepck decarboxylation activity impacts photosystem II light-energy use in a drought-induced CAM species

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ENVIRONMENTAL AND EXPERIMENTAL BOTANY 2020年 173卷

作者： Pikart, Filipe C. Matiz, Alejandra Alves, Frederico R. R. Mercier, Helenice Univ Sao Paulo Inst Biociencias Dept Bot Rua Matao 277 BR-05508090 Sao Paulo Brazil Univ Fed Goias Inst Ciencias Biol Dept Bot Ave Esperanca S-NCampus Samambaia BR-74690900 Goiania Go Brazil

Crassulacean acid metabolism (CAM) is a photosynthetic pathway characterized by nocturnal CO2 fixation into organic acids followed by their diurnal decarboxylation. Decarboxylation activity is time regulated, usually presenting higher activity in the middle of the day. This process can be performed by phosphoenolpyruvate carboxykinase (pepck) and/or by malic enzyme, resulting in an increase in internal CO2 concentration which may act to suppress photorespiration. It is suggested that the CO2 provided by the decarboxylation can improve the photochemical use of energy in CAM plants. This photosynthetic pathway also contributes to improve water use efficiency, and, in an interesting way, some species can engage this photosynthesis when subjected to water deficit. Guzmania monostachia is a bromeliad that occupies habitats with intermittent water availability and CAM can be induced by water deficit. To test the light-time modulation of decarboxylation activity and light-energy use in G. monostachia, we analyzed the activity of pepck, the main decarboxylating enzyme for this bromeliad, as well as the photochemical and non-photochemical quenching and organic acids contents at three times of the day. Results showed a diurnal modulation where the highest pepck decarboxylation activity coincided with an increase in the photochemical quenching and a decrease in the non-photochemical quenching. In addition, a reduction in acid content was observed, strongly indicating the interface between CO2 generation through acid decarboxylation and its effect on PSII photochemical activity and photorespiration in CAM-induced plants. The same was not observed for well-watered plants. Also, pepck activity modulation may be related to transcriptional and post-translational regulation, since an increase in transcript abundance and a modulation in the dephosphorylated enzyme activity was noted. Moreover, we suggest that the time modulation of pepck plays an important role in regulating PSII ene

关键词： pepck Guzmania monostachia Decarboxylation Photosynthesis Drought Malic acid

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Data on the expression of pepck in HepG2 hepatocytes transfected with miR-195

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DATA IN BRIEF 2017年 15卷 747-751页

作者： Yang, Won-Mo Min, Kyung-Ho Park, Se-Whan Lee, Wan Dongguk Univ Coll Med Dept Biochem Gyeongju Si 38067 Gyeongsangbuk D South Korea Dongguk Univ Coll Med Channelopathy Res Ctr Endocrine Channelopathy Goyang Si 10326 Gyeonggi Do South Korea

Dietary fats rich in saturated fatty acid (SFA) increase the risk of metabolic diseases, and certain microRNAs (miRNAs) dysregulated by SFA are associated with the pathogenesis of insulin resistance and type 2 diabetes. A previous study found that miR-195 is increased by SFA and impairs hepatic insulin signaling through the suppression of INSR (Yang et al., 2014) [1]. This article reports accompanying data to determine the effect of miR-195 on the expression of pepck, a key player in hepatic gluconeogenesis. The transfection of miR-195 in HepG2 hepatocytes was found to increase the mRNA and protein expression of pepck. Moreover, the insulin-stimulated reduction of pepck expression was attenuated drastically by miR-195. More detailed analysis and understanding of the role of miR-195 in diet-induced hepatic insulin resistance can be found in "Saturated fatty acid-induced miR-195 impairs insulin signaling and glycogen metabolism in HepG2 cells" (Yang et al., 2014)111. (C) 2017 The Authors. Published by Elsevier Inc.

关键词： MicroRNAs miR-195 pepck Obesity Hepatocyte

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Influence of IGF-1 on pyruvate carboxylase(PC) and phosphoenolpyruvate carboxykinase(pepck)mRNA expression and enzyme activity in calf hepatocytes cultured in vitro

Influence of IGF-1 on pyruvate carboxylase(PC) and phosphoen...

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第三届中国兽医临床大会

作者： WANG Jian-guo,Li Xiao-bing,SHI Xiao-xia,WANG Zhe~*,LIU Guo-wen~* (College of Animal Science and Veterinary Medicine,Jilin University,Changchun 130062,China)

The major role of insulin-like growth factor-1(IGF-1)in the liver is to mediate glucose uptake in hepatocytes to synthesize glycogen and maintain blood glucose homeostasis.(Objective and Method)In the present study,to evaluate the role of IGF-1 on gluconeogenesis and nutrient metabolism in dairy cattle,pyruvate carboxylase(PC)and phosphoenolpyruvate carboxykinase(pepck)expression and enzyme activity were evaluated in primary cultures of bovine hepatocytes treated with different concentrations of IGF-1 by quantitative polymerase chain reaction and spectrophotometry,respectively.(Result)The results showed that expression of PC and pepck were significantly lower in bovine hepatocytes by IGF-1 treatment in test group compare to the control group(P<0.01).As IGF-1 concentration increased,PC and pepck enzyme activity in bovine hepatocytes decreased.(Conclusion)Evaluating PC and pepck mRNA levels and enzyme activity may thus be useful to monitor subclinical ketosis in dairy cows.

关键词： IGF-1 PC pepck hepatocyte

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Breed-dependent transcriptional regulation of pepck-c expression in the liver of broiler chickens

Breed-dependent transcriptional regulation of PEPCK-c expres...

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第11届世界畜产大会(WCAP2013)

作者： Feng Guo Yanhong Zhang Lanli Su Abdelkareem A.Ahmed Yingdong Ni Ruqian Zhao

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RHIGF-Ⅰ对体外单层培养的肝细胞pepck mRNA丰度的影响

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中国兽医学报 2008年第4期28卷 434-437,445页

作者：邓俊良王娅崔恒敏徐闯刘国文王哲四川农业大学动物科技学院四川雅安625014 吉林大学畜牧兽医学院吉林长春130062

根据牛肝细胞中持家基因表达的恒定性,选用β肌动蛋白(β-actin)作为内参照,检测外源性添加不同质量浓度的重组人胰岛素样生长因子(RHIGF-Ⅰ)对体外培养牛肝细胞磷酸烯醇式丙酮酸羧激酶(pepck)mRNA表达水平的变化情况。选用体外培养的... 详细信息

根据牛肝细胞中持家基因表达的恒定性,选用β肌动蛋白(β-actin)作为内参照,检测外源性添加不同质量浓度的重组人胰岛素样生长因子(RHIGF-Ⅰ)对体外培养牛肝细胞磷酸烯醇式丙酮酸羧激酶(pepck)mRNA表达水平的变化情况。选用体外培养的犊牛单层肝细胞为实验模型,当细胞贴壁后在无血清培养液中添加RHIGF-Ⅰ,其终质量浓度分别为0、5、10、15、20、30、40、50、60、100、150μg/L,待添加8h后,提取总RNA,反转录,用相同的模板扩增pepck基因及β内参,再比较各梯度目的基因与内参的灰度。结果表明,随着添加RHIGF-Ⅰ质量浓度的升高,pepck mRNA表达呈下降趋势,且均低于对照组。

关键词：内源性竞争PCR RHIGF-Ⅰ pepck mRNA丰度肝细胞

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神经肽Y对犊牛肝细胞pepck mRNA丰度及其活性的影响

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中国兽医学报 2008年第10期28卷 1213-1215页

作者：陈承祯李小兵谢光洪徐闯刘国文张永宏王哲吉林大学畜牧兽医学院吉林长春130062

取单层培养36h生长良好的犊牛肝细胞,采用单因素重复试验,分别添加0、50、100、200、500、1000ng/L的羊体外合成神经肽Y(Neuropeptide Y,NPY),每个处理3个重复(每个重复2孔),再培养12h后分别提取RNA和制备细胞上清液。应用荧光定量PCR... 详细信息

取单层培养36h生长良好的犊牛肝细胞,采用单因素重复试验,分别添加0、50、100、200、500、1000ng/L的羊体外合成神经肽Y(Neuropeptide Y,NPY),每个处理3个重复(每个重复2孔),再培养12h后分别提取RNA和制备细胞上清液。应用荧光定量PCR方法检测外源NPY对肝细胞糖异生关键酶磷酸烯醇式丙酮酸羧激酶(Phosphoenolpyruvate carboxykinase,pepck)基因表达的影响,同时用比色法检测其对肝细胞pepck活性的影响。结果表明,一定浓度的NPY显著促进肝细胞pepck mRNA表达,增强了pepck活性。

关键词：神经肽Y 肝细胞磷酸烯醇式丙酮酸羧激酶 NPY pepck

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沙棘pepck基因的克隆与生物信息学分析

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分子植物育种 2023年第8期21卷 2591-2597页

作者：李安娜李贺陈思羽王立奇侯远琳李帅大连民族大学环境与资源学院大连116600

磷酸烯醇式丙酮酸羧激酶(pepck)是调节植物糖异生途径的限速酶。本研究从沙棘果实的转录组测序结果中筛选出pepck基因序列,利用RT-PCR技术对其进行克隆,利用生物信息学相关方法分析其编码蛋白的理化性质和结构特征。结果表明,该基因开... 详细信息

磷酸烯醇式丙酮酸羧激酶(pepck)是调节植物糖异生途径的限速酶。本研究从沙棘果实的转录组测序结果中筛选出pepck基因序列,利用RT-PCR技术对其进行克隆,利用生物信息学相关方法分析其编码蛋白的理化性质和结构特征。结果表明,该基因开放阅读框(open reading frame, ORF)长度为1 989 bp,编码一个由662个氨基酸组成的亲水性蛋白,主要由α-螺旋(30.21%)、β-折叠(17.22%),无规则卷曲(47.13%)和β-转角(5.44%)组成,其ORF序列与克莱门柚(Citrus clementina)同源性最高,氨基酸序列与川桑(Moras notabilis)和洋蓟(Cynara cardunculus var. scolymus)的有较高的同源性。该结果为进一步研究沙棘pepck蛋白功能以及揭示植物糖异生途径的调控机制提供理论基础。

关键词：沙棘 pepck 克隆生物信息学分析

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