Kermit Interacts with Gαo, Vang, and Motor Proteins in Drosophila Planar Cell Polarity
Kermit Interacts with Gαo, Vang, and Motor Proteins in Drosophila Planar Cell Polarity
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DOI：
10.1371/journal.pone.0076885
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发表时间：
2013-10-03
期刊：
影响因子：
3.7
通讯作者：
Katanaev, Vladimir L.
中科院分区：
文献类型：
--
作者：
Lin, Chen;Katanaev, Vladimir L.
氧化固醇是胆固醇的氧化产物。胆甾烷-3 β，5 α，6 β-三醇（简称三醇）是最丰富和最活跃的氧固醇之一。在这里，我们报告三醇对人前列腺癌细胞具有抗癌活性。用三醇处理细胞可剂量依赖性地抑制LNCaP CDXR-3、DU-145和PC-3人前列腺癌细胞的增殖，并减少软琼脂中的集落形成。三醇以20 mg/kg每日口服给药三周，显著延缓了裸鼠中PC-3异种移植物的生长。流式细胞术分析表明，三醇处理在10-40 μ M引起G1期细胞周期停滞，而TUNEL检测表明，三醇处理在20-40 μ M诱导所有三个细胞系的凋亡。Micro-Western阵列和传统的Western印迹方法表明，三醇处理导致Akt 1，磷酸化Akt Ser 473，磷酸化Akt Thr 308，PDK 1，c-Myc和Skp 2蛋白水平的表达降低以及细胞周期抑制剂p27（Kip）的积累。三醇处理也导致PC-3异种移植物中Akt 1蛋白表达降低。Skp 2在PC-3细胞中的过表达部分地挽救了由三醇引起的生长抑制。三醇处理抑制DU-145、PC-3和CDXR-3细胞的迁移和侵袭。与上皮间质转化相关的蛋白质以及粘着斑激酶的表达水平受到三醇处理这些细胞的影响。三醇处理引起E-钙粘蛋白蛋白水平的表达增加，但N-钙粘蛋白、波形蛋白、Slug、FAK、磷酸化FAK Ser 722和磷酸化FAK Tyr 861蛋白水平的表达降低。激光共聚焦显微镜显示β-肌动蛋白和α-微管蛋白在CDXR-3和DU-145细胞的外周重新分布。我们的观察结果表明，三醇可能是一个有前途的治疗剂晚期转移性前列腺癌。大多数情况下，成人发病的结核病（TB）的结果，重新激活预先存在的结核分枝杆菌感染。结核分枝杆菌通常侵入呼吸道，大多数患者发展为肺内结核;然而，一些患者同时发展为肺结核和肺外结核。本研究的目的是确定与肺结核患者并发肺外疾病风险增加相关的人口统计学和临床因素。我们比较了孤立性肺结核患者与并发肺结核和肺外结核的患者。我们最初分析了从基于人口的台湾全民健康保险数据库中随机选择的100万名受试者。基于对该数据库中5414例肺结核患者的分析，女性比男性更容易并发肺外结核（OR：1.30，p = 0.013）。对高雄医学大学医院数据库的一项单独分析，依赖于痰培养证实的肺结核，表明女性比男性更容易并发肺外结核（OR：1.62，p = 0.039）。在两个数据库中，年龄小于45岁的患者的肺外结核没有显著的性别差异。然而，对于45岁及以上的患者，女性比男性更可能并发肺外结核（保险数据库：9.0% vs. 6.8%，p = 0.016，OR：1.36;医院数据库：27.3% vs. 16.0%，p = 0.008，OR = 1.98）。我们的研究结果表明，在肺结核患者中，老年女性并发肺外结核的风险增加。除了普遍存在的顶部-基底极性，上皮细胞往往是极化的平面内的组织-被称为平面细胞极性（PCP）的现象。在果蝇中，PCP的表现在眼睛、翅膀和角质层中可见。PCP信号传导的几个组分在苍蝇和脊椎动物中已被表征，包括异源三聚体Go蛋白。然而，PCP中的Go信号传导伙伴在很大程度上仍然未知。使用遗传筛选，我们发现Kermit，以前涉及G蛋白和PCP信号，作为一种新的结合伙伴的围棋。通过下拉和遗传相互作用的研究，我们发现，Kermit与Go和另一个PCP组件旺，已知在PCP建立过程中进行细胞内重新定位相互作用。我们进一步证明，在PCP的Kermit的活动差异依赖于马达蛋白：微管为基础的动力蛋白和驱动蛋白马达和肌动蛋白为基础的肌球蛋白VI。我们的研究结果将Kermit作为Go的潜在换能器，将旺与马达蛋白联系起来，以便在PCP建立期间将其递送到专用细胞隔室。
Oxysterols are oxidation products of cholesterol. Cholestane-3 beta, 5 alpha, 6 beta-triol (abbreviated as triol) is one of the most abundant and active oxysterols. Here, we report that triol exhibits anti-cancer activity against human prostate cancer cells. Treatment of cells with triol dose-dependently suppressed proliferation of LNCaP CDXR-3, DU-145, and PC-3 human prostate cancer cells and reduced colony formation in soft agar. Oral administration of triol at 20 mg/kg daily for three weeks significantly retarded the growth of PC-3 xenografts in nude mice. Flow cytometric analysis revealed that triol treatment at 10-40 mu M caused G1 cell cycle arrest while the TUNEL assay indicated that triol treatment at 20-40 mu M induced apoptosis in all three cell lines. Micro-Western Arrays and traditional Western blotting methods indicated that triol treatment resulted in reduced expression of Akt1, phospho-Akt Ser473, phospho-Akt Thr308, PDK1, c-Myc, and Skp2 protein levels as well as accumulation of the cell cycle inhibitor p27(Kip). Triol treatment also resulted in reduced Akt1 protein expression in PC-3 xenografts. Overexpression of Skp2 in PC-3 cells partially rescued the growth inhibition caused by triol. Triol treatment suppressed migration and invasion of DU-145, PC-3, and CDXR-3 cells. The expression levels of proteins associated with epithelial-mesenchymal transition as well as focal adhesion kinase were affected by triol treatment in these cells. Triol treatment caused increased expression of E-cadherin protein levels but decreased expression of N-cadherin, vimentin, Slug, FAK, phospho-FAK Ser722, and phospho-FAK Tyr861 protein levels. Confocal laser microscopy revealed redistribution of beta-actin and alpha-tubulin at the periphery of the CDXR-3 and DU-145 cells. Our observations suggest that triol may represent a promising therapeutic agent for advanced metastatic prostate cancer.Most cases of adult-onset tuberculosis (TB) result from reactivation of a pre-existing Mycobacterium tuberculosis infection. Mycobacterium tuberculosis usually invades the respiratory tract and most patients develop intrapulmonary TB; however, some patients develop concurrent pulmonary and extra-pulmonary TB. The purpose of the present study was to identify the demographic and clinical factors associated with an increased risk of concurrent extra-pulmonary diseases in patients with pulmonary TB. We compared patients who had isolated pulmonary TB with patients who had concurrent pulmonary and extra-pulmonary TB. We initially analyzed one-million randomly selected subjects from the population-based Taiwan National Health Insurance database. Based on analysis of 5414 pulmonary TB patients in this database, women were more likely than men to have concurrent extra-pulmonary TB (OR: 1.30, p = 0.013). A separate analysis of the Kaohsiung Medical University Hospital database, which relied on sputum culture-proven pulmonary TB, indicated that women were more likely than men to have concurrent extra-pulmonary TB (OR: 1.62, p = 0.039). There was no significant gender difference in extra-pulmonary TB for patients younger than 45 years in either database. However, for patients 45 years and older, women were more likely than men to have concurrent extra-pulmonary TB (insurance database: 9.0% vs. 6.8%, p = 0.016, OR: 1.36; hospital database: 27.3% vs. 16.0%, p = 0.008, OR = 1.98). Our results indicate that among patients who have pulmonary TB, older females have an increased risk for concurrent extra-pulmonary TB.In addition to the ubiquitous apical-basal polarity, epithelial cells are often polarized within the plane of the tissue - the phenomenon known as planar cell polarity (PCP). In Drosophila, manifestations of PCP are visible in the eye, wing, and cuticle. Several components of the PCP signaling have been characterized in flies and vertebrates, including the heterotrimeric Go protein. However, Go signaling partners in PCP remain largely unknown. Using a genetic screen we uncover Kermit, previously implicated in G protein and PCP signaling, as a novel binding partner of Go. Through pull-down and genetic interaction studies, we find that Kermit interacts with Go and another PCP component Vang, known to undergo intracellular relocalization during PCP establishment. We further demonstrate that the activity of Kermit in PCP differentially relies on the motor proteins: the microtubule-based dynein and kinesin motors and the actin-based myosin VI. Our results place Kermit as a potential transducer of Go, linking Vang with motor proteins for its delivery to dedicated cellular compartments during PCP establishment.