Expression of NADPH oxidase (NOX) 5 in rabbit corneal stromal cells.
Expression of NADPH oxidase (NOX) 5 in rabbit corneal stromal cells.
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DOI：
10.1371/journal.pone.0034440
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
O'Brien WJ
中科院分区：
文献类型：
--
作者：
Rizvi F;Heimann T;O'Brien WJ
目的：检测兔角膜基质细胞(RCSC)是否表达NOX5。NADPH氧化酶(NOXES)是一种优先利用NADPH作为底物并产生超氧化物的酶。NOXES的几种异构体以多蛋白复合体的形式发挥作用，而NOX5和DUOXs的活性不需要辅助蛋白，并具有钙结合的EF手。以人NOX5为模板，通过RT-PCR扩增兔NOX5基因。对扩增产物进行测序，以确定其身份。通过蛋白质印迹分析鉴定了NOX5编码的蛋白。NOX5 siRNA被用来降低转录、蛋白质和钙刺激活性。在电子计算机分析中，建立了兔NOX5的推测结构、功能和进化。以NADPH为底物，而不是NADH为底物，测定RCSC中NOx活性。用NOX5为引物的RT-PCR扩增出288个碱基的RCSC基因片段，测序证实其与人NOX5基因同源。利用该序列预测了兔(Oryctolagus Cunrulus)NOX5基因。NOX5 siRNA可减少RCSC中NOX5mRNA的表达，减少离子霉素可刺激超氧化物的产生。免疫印迹分析检测到NOX5编码约65~70 kDa的蛋白质。在电子分析中，预测了一种可能的兔子NOX5蛋白，含有801个氨基酸。Motif搜索预测在N末端至少存在3个推测的EF-Hand，在C末端存在一个NOx结构域。与啮齿动物不同，NOX5基因在兔形动物细胞中表达的数据证明，兔子是研究NOX5功能的有趣模型。兔NOX5的活性受钙刺激，这是NOX5的一个普遍特征。NOX5也可能被证明是一种有用的遗传标记，用于研究扇形动物的分类地位和Glires分类。
To determine whether NOX 5 is expressed in rabbit corneal stromal cells (RCSC). NADPH oxidases (NOXes) are enzymes that preferentially use NADPH as a substrate and generate superoxide. Several isoforms of NOXes function as multi-protein complexes while NOX5 and DUOXs do not require the accessory proteins for their activity and possess calcium binding EF hands. Human NOX5 primers were used to amplify the rabbit NOX5 by RT-PCR. Amplified product was sequenced to confirm its identity. The protein encoded by the NOX5 was identified by western blot analysis. NOX5 siRNA was used to reduce transcript, protein, and calcium stimulated activity. In silico analyses were performed to establish the putative structure, functions, and evolution of rabbit NOX5. NOX activity was measured in RCSC with NADPH rather than NADH as a substrate. RT-PCR with NOX5 primers amplified 288 bp product using RCSC cDNA, which, when sequenced, confirmed its identity to human NOX5 mRNA. This sequence was used to predict the rabbit (Oryctolagus cuniculus) NOX5 gene. NOX5 siRNA reduced amounts of NOX5 mRNA in RCSC and reduced ionomycin stimulated superoxide production. A protein of about 65 to 70 kDa encoded by the NOX5 was detected by western blot analysis. In silico analysis predicted a putative rabbit NOX5 protein containing 801 amino acids. Motif searches predicted the presence of at least 3 putative EF-hands in N-terminus and a NOX domain in C terminal region. The data document that the NOX5 gene was expressed in cells of lagomorphs unlike rodents, making the rabbit an interesting model to study NOX5 functions. The activity of the rabbit NOX5 was calcium stimulated, a trait of NOX5 in general. NOX5 may also prove to be a useful genetic marker for studying the taxonomic position of lagomorphs and the Glires classification.
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影响因子：
20.1
作者：
Lyle AN;Deshpande NN;Taniyama Y;Seidel-Rogol B;Pounkova L;Du P;Papaharalambus C;Lassègue B;Griendling KK
通讯作者：
Griendling KK
影响因子：
56.9
作者：
Asher, RJ;Meng, J;Novacek, MJ
通讯作者：
Novacek, MJ
影响因子：
2.5
作者：
Kawai, Yusuke K.;Ikenaka, Yoshinori;Ishizuka, Mayumi
通讯作者：
Ishizuka, Mayumi
影响因子：
3.3
作者：
Kawahara, Tsukasa;Lambeth, J. David
通讯作者：
Lambeth, J. David
影响因子：
7.4
作者：
BelAiba, Rachida S.;Djordjevic, Talija;Goerlach, Agnes
通讯作者：
Goerlach, Agnes