关键词： 非小细胞肺癌   IQGAP1   ERK   信号通路   细胞增殖  

摘要： 该研究探讨了IQGAP1(IQ domain GTPase-activating protein 1)对非小细胞肺癌(nonsmall cell lung cancer,NSCLC)细胞增殖的影响及其对ERK信号通路的调节作用。将内源性IQGAP1低表达的A549细胞中分为空白组、空载组和IQGAP1过表达组;将内源性IQGAP1高表达的H1299细胞中分为空白组、阴性对照si RNA组和IQGAP1 si RNA组;采用ERK1/2磷酸化抑制剂U0126处理上述两株细胞。MTT法检测细胞增殖能力,Western blot法检测ERK1/2和p-ERK1/2蛋白质水平。结果显示,在A549细胞中,过表达IQGAP1能促进细胞增殖并促进ERK1/2磷酸化;在H1299中,敲低IQGAP1表达能够抑制细胞增殖并下调ERK1/2磷酸化水平。用U0126处理后能抑制IQGAP1对细胞增殖的促进作用。研究结果表明,IQGAP1可通过ERK信号通路促进体外非小细胞肺癌细胞增殖。

关键词： Allele specific expression   eQTL   Multiple sclerosis   CD69   IKZF3   IQGAP1  

摘要： Background: Multiple sclerosis is a chronic inflammatory, demyelinating disease of the central nervous system. Recent genome-wide studies have revealed more than 110 single nucleotide polymorphisms as associated with susceptibility to multiple sclerosis, but their functional contribution to disease development is mostly unknown. Results: Consistent allelic imbalance was observed for rs907091 in IKZF3 and rs11609 in IQGAP1, which are in strong linkage disequilibrium with the multiple sclerosis associated single nucleotide polymorphisms rs12946510 and rs8042861, respectively. Using multiple sclerosis patients and healthy controls heterozygous for rs907091 and rs11609, we showed that the multiple sclerosis risk alleles at IKZF3 and IQGAP1 are expressed at higher levels as compared to the protective allele. Furthermore, individuals homozygous for the multiple sclerosis risk allele at IQGAP1 had a significantly higher total expression of IQGAP1 compared to individuals homozygous for the protective allele. Conclusions: Our data indicate a possible regulatory role for the multiple sclerosis-associated IKZF3 and IQGAP1 variants. We suggest that such cis-acting mechanisms may contribute to the multiple sclerosis association of single nucleotide polymorphisms at IKZF3 and IQGAP1.

关键词： Cell signaling   IQGAP   Phosphatidylinositol phosphate kinase   Phosphoinositide  

摘要： Phosphatidylinositol 4,5-bisphosphate (PI4,5P₂) is a lipid messenger that regulates a wide variety of cellular functions. The majority of cellular PI4,5P₂ is generated by isoforms of the type I phosphatidylinositol phosphate kinases (PIPKI) that are generated from three genes, and each PIPKI isoform has a unique distribution and function in cells. It has been shown that the signaling specificity of PI4,5P₂ can be determined by a physical association of PIPKs with PI4,5P₂ effectors. IQGAP1 is newly identified as an interactor of multiple isoforms of PIPKs. Considering the versatile roles of IQGAP1 in cellular signaling pathways, IQGAP1 may confer isoform-specific roles of PIPKs in distinct cellular locations. In this mini review, the emerging roles of PIPKs that are regulated by an association with IQGAP1 will be summarized. Focuses will be on cell migration, vesicle trafficking, cell signaling, and nuclear events. © 2015 Elsevier Ltd.

关键词： HBx   CDC42   IQGAP1   细胞恶性转化   定量蛋白质组学  

摘要： 肝细胞癌是世界第五大流行癌症,致死率非常高。慢性乙型肝炎病毒(Hepatitis B virus, HBV)感染被认为是诱发肝癌的主要因素。研究发现HBV基因组x基因编码的X蛋白(hepatitsis B virus X protein, HBx)在HBV复制和诱发肝癌过程中具有重要作用。课题组前期对HBx转基因小鼠SILAC (Stable Isotope Labeling with Amino acids in Cell culture)定量蛋白质组学研究发现,在HBx诱发小鼠肝脏由重度炎症转变为癌症过程中,CDC42信号通路相关分子及CDC42蛋白本身蛋白表达量上调,提示在HBx诱发小鼠肝癌过程中,CDC42信号通路被激活。激活的CDC42可参与多条信号通路,引起细胞骨架、细胞周期、细胞侵袭转移能力等改变。为了解CDC42蛋白是否参与以及如何参与HBx介导的Huh7细胞恶性转化,本研究首先通过CRISPR/Cas9技术对Huh7-HBx细胞中的CDC42基因进行修饰,而后通过添加CDC42特异性抑制剂CASIN抑制Huh7-mock细胞和Huh7-HBx细胞中CDC42活性,探究CDC42功能破坏时HBx介导的Huh7恶性转化程度变化,进而揭示CDC42在HBx诱发人肝细胞恶性转化中的作用。研究结果将为深入理解HBV致癌机制、寻找HBV相关肝癌的治疗靶标提供理论依据。具体研究结果如下：***介导Huh7细胞恶性转化为了解HBx基因的异常表达是否可引起人肝癌细胞系Huh7细胞系的恶性转化,首先应用实时定量PCR检测Huh7-mock细胞和Huh7-HBx细胞中HBx基因表达情况,采用CCK-8试剂盒检测2种细胞的增殖能力、Annex V和FITC试剂盒检测细胞凋亡水平,并通过细胞划痕实验检测HBx基因转染前后Huh7细胞迁移能力的变化情况。结果显示：(1)Huh7-HBx细胞中HBx基因的表达显著高于Huh7-mock细胞(p<0.01)；(2)Huh7-HBx细胞的增殖能力显著高于Huh7-mock细胞(p<0.01)；(3)Huh7-HBx细胞的抗凋亡能力显著高于Huh7-mock细胞(流式细胞仪分析)。(4)Huh7-HBx细胞的迁移能力高于Huh7-mock细胞；(5)与Huh7-mock细胞相目比,Huh7-HBx细胞中CDC42蛋白表达量升高(Western blot分析)。***介导Huh7细胞恶性转化依赖CDC42为了验证CDC42蛋白是否参与HBx介导的Huh7细胞恶性转化,首先应用CRISPR/Cas9系统对Huh7-HBx细胞中CDC42基因进行编辑,检测CDC42基因敲除前后细胞的增殖能力、凋亡水平及迁移能力,而后采用CDC42蛋白特异性抑制剂CASIN处理Huh7-mock细胞和Huh7-HBx细胞,了解其对细胞增殖及凋亡水平的影响。结果发现：(1)获得在CDC42基因第二外显子处具有4个碱基缺失的突变体细胞(Huh7-HBx CDC42 KO);(2)Huh7-HBx CDC42 KO细胞增殖能力显著低于Huh7-HBx细胞(p<0.01)；(3)Huh7-HBx CDC42 KO细胞群体中晚期凋亡细胞比例升高,即细胞抗凋亡能力显著下降；(4)Huh7-HBx CDC42 KO细胞的迁移能力低于Huh7-HBx细胞；(5)不同浓度CASIN处理一定时间后,Huh7-HBx细胞的增殖能力显著降低(p<0.01),而Huh7细胞的增殖能力没有显著变化；Huh7-HBx细胞的抗凋亡能力显著下降(p

关键词： microRNA-506   乳腺癌   含IQ模序的GTP1酶活化蛋白(IQGAP1)   ERK信号通路  

摘要： 乳腺癌是人类最常见的一类恶性肿瘤,发病率高居妇女癌症第二位。Micro RNA-506（mi R506）与多种肿瘤的发生发展密切相关,且在肿瘤组织中表达异常。然而,mi RNA-506调节乳腺癌发生发展的作用机制目前尚不清楚。在多种肿瘤组织和肿瘤细胞中含IQ模序的GTP1酶活化蛋白（IQGAP1）表达上调,发挥致癌作用。因此,我们推测mi RNA-506在乳腺癌中发挥抑癌作用,且其下游靶基因为IQGAP1。本研究以mi RNA-506为切入点,为进一步探讨mi RNA-506调节乳腺癌发生发展的作用机制,为开发更有效的乳腺癌治疗靶点和方法奠定理论基础。在本研究中,我们首先采用RT-q PCR法检测mi RNA-506在恶性乳腺组织以及乳腺癌细胞系中的表达,结果显示在人乳腺组织及乳腺癌细胞系中,mi R-506表达量显著下降,且随着肿瘤分期增加,呈现逐渐减少的趋势。功能性实验结果显示,mi R-506低表达的MDA-MB-231细胞的增殖、侵袭以及粘附能力显著增加,而过表达mi R-506的HCC1937细胞的增殖、侵袭以及粘附能力显著下降,提示mi R-506在乳腺癌中发挥抑癌作用。接下来,我们检测到在人乳腺组织及乳腺癌细胞系中,IQGAP1蛋白表达显著上调,且随着肿瘤分期增加,表达逐渐增高,提示IQGAP1在乳腺癌中发挥促癌作用。荧光素酶实验证实mi R-506通过3’UTR端靶向作用于IQGAP1并下调IQGAP1表达,提示IQGAP1为mi R-506的下游靶基因。进一步的研究指出IQGAP1可以中和mi R-506所诱导的肿瘤细胞的增殖侵袭以及粘附能力。为进一步探讨mi R-506在乳腺癌中的作用机制,我们对相关的信号通路分子进行了检测,研究结果表明,mi R-506能够下调Erk1/2磷酸化,且IQGAP1可缓解mi R-506的这种作用。本研究首次证实mi R-506在乳腺癌细胞中能够作为一种肿瘤抑制因子,可直接下调IQGAP1的表达,进而抑制ERK信号通路发挥抑癌作用。本研究为将mi R-506/IQGAP1/ERK通路作为成为新的乳腺癌治疗靶点提供理论依据。

关键词： 人肾脏足细胞   细胞凋亡   IQGAP1   ERK1/2蛋白   p-ERK1/2蛋白  

摘要： 研究背景糖尿病是临床上一种常见内分泌性疾病,它是由于多种病因引起的以慢性高血糖为特征的代谢性疾病,是由于胰岛素分泌和（或）作用缺陷所致。持续高血糖与长期代谢紊乱等可导致全身组织器官的多种并发症,尤其是眼、肾、心血管及神经系统的慢性进行性病变、功能减退或衰竭。糖尿病的全身血管并发症分为大血管并发症和微血管并发症,糖尿病肾病（diabetic nephropathy,DN）是糖尿病最常见的微血管并发症之一,其临床表现主要为高血压、水肿、严重蛋白尿、低白蛋白血症和进行性肾功能减退;肾脏病理主要表现为结节状或弥漫性系膜细胞增殖、基底膜(（GBM）增厚,系膜细胞外基质积聚、足细胞足突融合,肾小球硬化及肾小管间质纤维化。糖尿病肾病是目前世界范围内导致终末期肾病（end stage renal disease,ESRD）的首位位病因,增加各国卫生保健事业支出的同时也给予社会、家庭、个人带来沉重负担。在我国糖尿病肾病发病率以及在终末期肾衰竭透析患者中所占的比例也成逐年增加趋势,严重影响了患者的生存质量。因此,探讨糖尿病肾病的发病机制、及早发现糖尿病肾病并给予适当干预治疗措施对广大糖尿病肾病患者具有重要的社会和现实意义。DN的发病机制十分复杂,涉及到糖脂代谢紊乱、炎症介质释放、血流动力学异常、细胞因子、氧化应激以及细胞凋亡等多因素的作用。近年来的研究显示,肾小球滤过屏障结构和功能的改变,尤其是作为肾小球滤过屏障重要组成成分的足细胞损伤,在糖尿病肾病的进展中发挥了举足轻重的作用,被认为是引起DN蛋白尿和肾小球硬化的关键因素。因此,研究足细胞的损伤为DN的预防和治疗带来了新的契机。足细胞又称为脏层肾小球上皮细胞,附着于肾小球基底膜外侧,与肾小球基底膜（glomerular basement membrane,GBM）、内皮细胞共同构成肾小球滤过屏障。足细胞的主要作用是对蛋白质的滤过及GBM成分的更新起着举足轻重的作用。由于足细胞是高度分化细胞,增殖能力较差,当足细胞受到损伤时,尿蛋白漏出增加,从而加重肾衰竭的发生,而蛋白尿又进一步加重足细胞的损伤。在DN的发生发展中,足细胞的改变主要包括足细胞数目减少,细胞凋亡增加,流动性增强,直至足细胞从GBM上脱落随尿液排出,导致GBM裸露,形成局灶粘连和肾小球硬化。DN足细胞损伤机制复杂,其不是单由某个因素所诱发,而是多个因素的联合作用。主要包括肾蛋白（nephrin）、高血糖、整合素、机械应力、炎性反应、肾素-血管紧张素-醛固酮系统（RAAS）的激活、AGEs的蓄积、GH-胰岛素样生长因子（insulin-like growth factor,IGF）-I轴、血管内皮生长因子（VEGF）、类肝素硫酸蛋白聚糖（HSPG）、肿瘤抑制基因（wT1）、Podocalyxin蛋白、过氧化物酶体增殖物激活受体（perox-isome proliferatoractivated receptor,PPAR）、脂联素、微小 RNA（micro RNA,miRNA）、TGF-β1、氧化应激及细胞凋亡等。有研究显示,IQ结构域GTP酶活化蛋白1（IQdomainGTPase-activating protein 1,IQGAP1）是一种含有多个蛋白结合域的支架蛋白,近期研究发现IQGAP1在细胞黏附迁移、维持细胞形态以及调节细胞凋亡等众多生命过程中发挥了举足轻重的作用[1,2]。IQGAP1是否参与高糖诱导的足细胞凋亡尚未见报道,基于上述理论基础,我们设计并进行了本次研究。本课题分为两部分,第一步部分,采用体外培养人肾脏足细胞（HPC）,观察高糖刺激对IQGAP1表达和分布以及对足细胞凋亡的影响。第二部分,进一步观察IQGAP1与丝裂原活化蛋白激酶（MAPK）信号通路以及肾素□血管紧张素□醛固酮系统（RAAS）在足细胞凋亡中的作用,探讨高糖诱导肾脏足细胞凋亡的分子机制,从而为更好的治疗糖尿病肾病提供理论支持。第一部分高糖刺激对足细胞中IQGAP1表达和分布以及凋亡的影响研究目的:1、观察人肾脏足细胞在体外高糖刺激下IQGAP1表达情况;2、观察人肾脏足细胞在体外高糖刺激下IQGAP1分布情况;3、观察人肾脏足细胞在体外高糖刺激下细胞凋亡情况。研究方法:HPC在33℃、5%CO2培养箱中,以含有10%胎牛血清及γ-干扰素（10 U/ml）的RPMI 1640培养液培养细胞使其增殖,传代后更换为不含Y-干扰素的RPMI 1640培养液,置入37° C、5%C02培养箱中培养10～14天,促进细胞分化成熟,用于后续实验。HPC被随机分为3组:A组:正常对照组（Normal Control,NC,）、B组:甘露醇对照组（Manitol+Glucose Control,MG）和C组:高糖组（High Glucose,HG）。NC组细胞给

关键词： Legionella pneumophila   Cell migration   Dictyostelium discoideum   Macrophages   Host cell migration   Guanosine triphosphatase   Amoebas   Small interfering RNA  

摘要： Small molecule signaling promotes the communication between bacteria as well as between bacteria and eukaryotes. The opportunistic pathogenic bacterium Legionella pneumophila employs LAI-1 (3-hydroxypentadecane-4-one) for bacterial cell-cell communication. LAI-1 is produced and detected by the Lqs (Legionella quorum sensing) system, which regulates a variety of processes including natural competence for DNA uptake and pathogen-host cell interactions. In this study, we analyze the role of LAI-1 in inter-kingdom signaling. L. pneumophila lacking the autoinducer synthase LqsA no longer impeded the migration of infected cells, and the defect was complemented by plasmid-borne lqsA. Synthetic LAI-1 dose-dependently inhibited cell migration, without affecting bacterial uptake or cytotoxicity. The forward migration index but not the velocity of LAI-1-treated cells was reduced, and the cell cytoskeleton appeared destabilized. LAI-1-dependent inhibition of cell migration involved the scaffold protein IQGAP1, the small GTPase Cdc42 as well as the Cdc42-specific guanine nucleotide exchange factor ARHGEF9, but not other modulators of Cdc42, or RhoA, Rac1 or Ran GTPase. Upon treatment with LAI-1, Cdc42 was inactivated and IQGAP1 redistributed to the cell cortex regardless of whether Cdc42 was present or not. Furthermore, LAI-1 reversed the inhibition of cell migration by L. pneumophila, suggesting that the compound and the bacteria antagonistically target host signaling pathway(s). Collectively, the results indicate that the L. pneumophila quorum sensing compound LAI-1 modulates migration of eukaryotic cells through a signaling pathway involving IQGAP1, Cdc42 and ARHGEF9.

关键词： IQGAP1   口腔鳞状细胞癌(OSCC)   增殖   侵袭  

摘要： 目的:研究IQ结构域三磷酸鸟苷合酶-激活蛋白1(IQGAP1)在口腔鳞状细胞癌(OSCC)组织中的表达,探讨其对肿瘤细胞增殖和侵袭的影响及其作用机制。方法:免疫印迹和荧光定量PCR检测IQGAP1在癌组织和癌旁组织中的表达水平;利用脂质体转染法将重组质粒pc DNA3.1-IQGAP1转入人口腔鳞状细胞癌细胞系SCC-4,并用Akt抑制剂预处理,免疫印迹检测Akt磷酸化水平、MTT法和Transwell侵袭实验分别测定细胞增殖和侵袭能力的变化。结果:IQGAP1蛋白和mRNA表达量显著高于癌旁组织(P<0.05),重组质粒转染后,SCC-4细胞中IQGAP1表达量明显增加,细胞的增殖能力和侵袭能力增强(P<0.05)。IQGAP1过表达后细胞中p Akt水平明显升高;当用Akt抑制剂处理后,IQGAP1诱导的p Akt的水平明显降低,细胞增殖和侵袭能力降低(P<0.05)。结论:IQGAP1在OSCC组织中高表达,可能通过调节Akt通路的活化来介导癌细胞的增殖和侵袭能力。

关键词： 肝细胞肝癌   IQ模体的Ras   GTP酶活化蛋白1   肝切除术   预后  

摘要： 目的探讨含有IQ模体的Ras GTP酶活化蛋白1（IQGAP1）在肝细胞肝癌组织中的表达及其临床意义。方法收集2007~2009年期间在笔者所在医院行肝癌切除治疗的79例肝细胞肝癌患者的临床资料,采用免疫组化法检测肝癌组织中IQGAP1的表达,并分析其与肝癌患者临床病理特征的关系,比较IQGAP1表达阳性和阴性患者肝癌切除术后的累积无瘤生存率及总体生存率。结果本组79例肝癌患者中肝癌组织中IQGAP1表达阳性者43（54.4%）,IQGAP1表达阳性的患者多为低分化及存在微血管侵犯者。IQGAP1表达阳性与阴性患者术后1、3及5年的累积无瘤生存率分别为67.4%、39.5%及23.3%和100%、94.4%及83.3%,前者明显低于后者（P〈0.001）;IQGAP1表达阳性患者术后1、3及5年的累积总体生存率为97.7%、71.5%和53.3%,明显低于IQGAP1表达阴性者的100%、97.2%和88.9%（P〈0.001）。结论肝细胞肝癌患者中IQGAP1阳性表达者预后较差。IQGAP1可能是肝细胞肝癌预后的指标。

关键词： colorectal cancer   cell invasion   DNAJB6   IQGAP1  

摘要： DNAJB6 is a member of the heat shock protein 40 (Hsp40) family. We here investigated the clinical correlation and biological role of DNAJB6 overexpression in colorectal cancer (CRC). The expression of DNAJB6 protein was examined in 200 cases of colorectal adenocarcinomas by immunohistochemistry (IHC) technology. Gene transfection and RNA interference were performed to determine the effect of DNAJB6 expression on the invasion of CRC cells and to explore the underlying molecular mechanisms in vitro and in vivo. Overexpression of DNAJB6 was found in 39% (78/200) of the CRC tissues, especially in tumors at pT4 as compared with at pT1-3 (P = 0.02). A Kaplan-Meier survival analysis revealed a correlation between DNAJB6 expression and overall survival (OS) times (P = 0.003). Multivariate analysis confirmed that DNAJB6 overexpression was an independent prognostic factor for CRC (P = 0.002). RNA interference-mediated silencing of the DNAJB6 gene inhibited the invasion of CRC cells in vitro were accompanied by a significant reduction in the protein levels of IQ-domain GTPase-activating protein 1 (IQGAP1) and phosphorylated ERK (pERK). An in vivo assay showed that inhibition of DNAJB6 expression decreased the lung metastases of CRC cells. IHC analysis of serial sections showed that there was a positive correlation between DNAJB6 and IQGAP1 expression in primary CRC tissues (P = 0.013). The data suggest that DNAJB6 plays an important oncogenic role in CRC cell invasion by up-regulating IQGAP1 and activating the ERK signaling pathway and that DNAJB6 may be used as a prognostic marker for CRC. (C) 2014 Wiley Periodicals, Inc.