Ikaros通过靶向调控 c-KIT抑制B-ALL白血病细胞的增殖

Ikaros是一种重要的造血细胞分化与发育调控因子，其基因结构、蛋白质活性的改变与淋巴细胞白血病的发生密切相关。致癌基因c-KIT与白血病的发生有直接联系，但Ikaros与c-KIT之间的调控关系尚未见报道。本研究报道，在人急性B淋巴细胞白血病（B-acute lymphoblastic leukemia, B-ALL）细胞中，Ikaros可靶向调控c KIT基因的转录与蛋白质表达。通过在人B ALL细胞系Nalm6中分别高表达和shRNA干扰Ikaros后，qRT-PCR与 Western 印迹结果显示，Ikaros可直接抑制c-KIT基因的表达。双荧光素酶报告实验检测Ikaros及其突变体对c-KIT基因的直接靶向作用。结果显示，野生型Ikaros可明显抑制c-KIT的表达，而突变体则不能。进一步利用染色质免疫共沉淀技术（chromatin immunoprecipitation，ChIP），检测Ikaros对c-KIT上游启动子序列的结合活力。结果显示，Ikaros蛋白在c KIT的上游调控区约-500 bp处有明显的结合。Ikaros通过靶向c-KIT上游启动子，抑制c-KIT表达，抑制B-ALL细胞的增殖，为临床治疗白血病提供了新思路。     

The Oncogenic MicroRNA-21 Inhibits the Tumor Suppressive Activity of FBXO11 to Promote Tumorigenesis

The microRNA miR-21 is overexpressed in most human cancers and accumulating evidence indicates that it functions as an oncogene. Since miRNAs suppress the expression of their target genes, we hypothesized that some miR-21 targets may act as tumor suppressors, and thus their expression would be anticipated to be reduced by the high miR-21 levels observed in various human cancers. By microarray analysis and quantitative PCR we identified and validated FBXO11 (a member of the F-box subfamily lacking a distinct unifying domain) as a miR-21 target gene. FBXO11 is a component of the SKP1-CUL1-F-box ubiquitin ligase complex that targets proteins for ubiquitination and proteosomal degradation. By loss of function and gain of function studies, we show that FBXO11 acts as a tumor suppressor, promotes apoptosis and mediates the degradation of the oncogenic protein BCL6. The critical role that FBXO11 plays in miR-21-mediated tumorigenesis was demonstrated by a rescue experiment, in which silencing FBXO11 in miR-21KD cancer cells restored their high tumorigenicity. Expression of miR-21 and FBXO11 are inversely correlated in tumor tissue, and their expression correlates with patient survival and tumor grade. High FBXO11 expression correlates with better patient survival and lower tumor grade consistent with its tumor suppressor activity. In contrast high miR-21 expression, which correlates with poor patient survival and higher tumor grade, is consistent with its oncogenic activity. Our results identify FBXO11 as a novel miR-21 target gene, and demonstrate that the oncogenic miRNA miR-21 decreases the expression of FBXO11, which normally acts as a tumor suppressor, and thereby promotes tumorigenesis.     

Pseudomonas aeruginosa Inhibits the Growth of Cryptococcus Species

Pseudomonas aeruginosa is a ubiquitous and opportunistic bacterium that inhibits the growth of different microorganisms, including Gram-positive bacteria and fungi such as Candida spp. and Aspergillus fumigatus. In this study, we investigated the interaction between P. aeruginosa and Cryptococcus spp. We found that P. aeruginosa PA14 and, to a lesser extent, PAO1 significantly inhibited the growth of Cryptococcus spp. The inhibition of growth was observed on solid medium by the visualization of a zone of inhibition of yeast growth and in liquid culture by viable cell counting. Interestingly, such inhibition was only observed when P. aeruginosa and Cryptococcus were co-cultured. Minimal inhibition was observed when cell-cell contact was prevented using a separation membrane, suggesting that cell contact is required for inhibition. Using mutant strains of Pseudomonas quinoline signaling, we showed that P. aeruginosa inhibited the growth of Cryptococcus spp. by producing antifungal molecules pyocyanin, a redox-active phenazine, and 2-heptyl-3,4-dihydroxyquinoline (PQS), an extracellular quorum-sensing signal. Because both P. aeruginosa and Cryptococcus neoformans are commonly found in lung infections of immunocompromised patients, this study may have important implication for the interaction of these microbes in both an ecological and a clinical point of view.     

蛋白质SUMO化修饰研究进展

SUMO(small ubiquitin-related modifier)是类泛素蛋白家族的重要成员之一,可与多种蛋白结合发挥相应的功能,其分子结构及SUMO化反应途径都与泛素类似,但二者功能完全不同。SUMO化修饰可参与转录调节、核转运、维持基因组完整性及信号转导等多种细胞内活动,是一种重要的多功能的蛋白质翻译后修饰方式。SUMO化修饰功能的失调可能导致某些疾病的发生。     

Trophosome of the Deep-Sea Tubeworm Riftia pachyptila Inhibits Bacterial Growth

The giant tubeworm Riftia pachyptila lives in symbiosis with the chemoautotrophic gammaproteobacterium Cand. Endoriftia persephone. Symbionts are released back into the environment upon host death in high-pressure experiments, while microbial fouling is not involved in trophosome degradation. Therefore, we examined the antimicrobial effect of the tubeworm’s trophosome and skin. The growth of all four tested Gram-positive, but only of one of the tested Gram-negative bacterial strains was inhibited by freshly fixed and degrading trophosome (incubated up to ten days at either warm or cold temperature), while no effect on Saccharomyces cerevisiae was observed. The skin did not show antimicrobial effects. A liquid chromatography-mass spectrometric analysis of the ethanol supernatant of fixed trophosomes lead to the tentative identification of the phospholipids 1-palmitoleyl-2-lyso-phosphatidylethanolamine, 2-palmitoleyl-1-lyso-phosphatidylethanolamine and the free fatty acids palmitoleic, palmitic and oleic acid, which are known to have an antimicrobial effect. As a result of tissue autolysis, the abundance of the free fatty acids increased with longer incubation time of trophosome samples. This correlated with an increasing growth inhibition of Bacillus subtilis and Listeria welshimeri, but not of the other bacterial strains. Therefore, the free fatty acids produced upon host degradation could be the cause of inhibition of at least these two bacterial strains.     

Solar UV-B Radiation Inhibits the Growth of Antarctic Terrestrial Fungi

We tested the effects of solar radiation, and UV-B in particular, on the growth of Antarctic terrestrial fungi. The growth responses to solar radiation of five fungi, Geomyces pannorum, Phoma herbarum, Pythium sp., Verticillium sp., and Mortierella parvispora, each isolated from Antarctic terrestrial habitats, were examined on an agar medium in the natural Antarctic environment. A 3-h exposure to solar radiation of >287 nm reduced the hyphal extension rates of all species relative to controls kept in the dark. Pythium sp. cultures exposed to solar radiation for 1.5 h on five consecutive days were most sensitive to radiation of >287 nm, but radiation of >313 nm also inhibited growth to a lesser extent. Radiation of >400 nm had no effect on hyphal growth relative to controls kept in the dark. Short-wave solar UV-B radiation of between 287 and 305 nm inhibited the growth of Pythium sp. hyphae on and below the surface of the agar medium after 24 h, but radiation of ≥345 nm only reduced the growth of surface hyphae. Similar detrimental effects of UV-B on surface and, to a lesser extent, submerged hyphae of all five fungi were shown in the laboratory by using artificial UV-B from fluorescent lamps. A comparison of growth responses to solar radiation and temperature showed that the species that were most resistant to UV radiation grew fastest at higher temperatures. These data suggest that solar UV-B reduces the growth of fungi on the soil surface in the Antarctic terrestrial environment.     

Osmotic Shock Inhibits Auxin-stimulated Acidification and Growth

Cells of oat coleoptiles (Avena sativa L. cv. "Garry") have been osmotically shocked in order to observe the effect of alterations of the plasma membrane on some auxin responses. When coleoptile sections were treated sequentially with 0.5 m mannitol and 1 mm Na-phosphate (pH 6.4) at 4 C, polar auxin transport and acidification by 1 mM CaCl(2) were unaffected, but auxin-stimulated acidification and growth were eliminated. Shock treatment also had no effect on acid-stimulated growth or on freezing point depression by the cytoplasm. It is suggested that osmotic shock modifies a portion of the plasma membrane which interacts with auxin and eventually leads to growth.     

Gefitinib Inhibits the Growth of Toxoplasma gondii in HeLa Cells

Toxoplasma gondii is the causative agent of toxoplasmosis with symptoms of congenital neurological and ocular diseases and acquired lymphadenitis, retinochoroiditis, and meningoencephalitis. Small molecules which block the activity of protein kinases were tested in in vitro culture of T. gondii to find new therapeutic drugs of safer and more effective than the combined administration of pyrimethamine and sulfadoxine that sometimes provoke lethal Stevens-Johnson syndrome. Among them, Gefitinib and Crizotinib inhibited intracellular growth of T. gondii in HeLa cells by counting the number of T. gondii per parasitophorous vacuolar membrane whereas Sunitinib did not. Gefitinib inhibited the growth of T. gondii in a dose-dependent manner over 5 µM up to the tolerable concentration of HeLa cells and halted the division of the parasite immediately from the time point of treatment. Gefitinib inhibition suggests that tyrosine kinases of EGFR family or other homologous kinases of the parasite itself may be the target to cause the block of T. gondii growth.     

Overexpression of Autophagy-Related Genes Inhibits Yeast Filamentous Growth

Under conditions of nitrogen stress, the budding yeast S. cerevisiae initiates a cellular response involving the activation of autophagy, an intracellular catabolic process for the degradation and recycling of proteins and organelles. In certain strains of yeast, nitrogen stress also drives a striking developmental transition to a filamentous form of growth, in which cells remain physically connected after cytokinesis. We recently identified an interrelationship between these processes, with the inhibition of autophagy resulting in exaggerated filamentous growth. Our results suggest a model wherein autophagy mitigates nutrient stress, and filamentous growth is responsive to the degree of this stress. Here, we extended these studies to encompass a phenotypic analysis of filamentous growth upon overexpression of autophagy-related (ATG) genes. Specifically, overexpression of ATG1, ATG3, ATG7, ATG17, ATG19, ATG23, ATG24, and ATG29 inhibited filamentous growth. From our understanding of autophagy in yeast, overexpression of these genes does not markedly affect the activity of the pathway; thus, we do not expect that this filamentous growth phenotype is due strictly to diminished nitrogen stress in ATG overexpression mutants. Rather, these results highlight an additional undefined regulatory mechanism linking autophagy and filamentous growth, possibly independent of the upstream nitrogen-sensing machinery feeding into both processes.

Addendum to:

An Interrelationship Between Autophagy and Filamentous Growth in Budding Yeast

J. Ma, R. Jin, X. Jia, C.J. Dobry, L. Wang, F. Reggiori, J. Zhu and A. Kumar

Genetics 2007; In press     

Suppressive Activity of Streptomycin on the Growth of Mycobacterium lepraemurium in Macrophage Cultures

The effect of streptomycin on the growth of an obligate intracellular bacterium was studied in a new host-parasite cell system. The system consisted of Mycobacterium lepraemurium grown in cultures of mouse peritoneal macrophages. Since these organisms do not grow in bacteriological media, the influence of extracellular bacterial growth can be ruled out. The suppressive activity of streptomycin was observed in a total of five experiments. At the end of 4 weeks, the average number of organisms per macrophage for the controls was 65.7; for cultures with streptomycin at concentrations of 0.5, 1, 5, 10, 50, and 100 mug/ml of medium, it was 45.4, 38.3, 28.7, 22.7, 13.4, and 8.2, respectively. A good dose-response relationship was evident. M. lepraemurium which had been treated in macrophage cultures with various concentrations of the antibiotic for 6 to 8 weeks was used to infect fresh macrophages. These cultures were in turn treated with streptomycin. Resistance of the organisms to streptomycin did not occur.     

Erlotinib Inhibits Growth of a Patient-Derived Chordoma Xenograft

Chordomas are rare primary bone tumors that occur along the neuraxis. Primary treatment is surgery, often followed by radiotherapy. Treatment options for patients with recurrence are limited and, notably, there are no FDA approved therapeutic agents. Development of therapeutic options has been limited by the paucity of preclinical model systems. We have established and previously reported the initial characterization of the first patient-derived chordoma xenograft model. In this study, we further characterize this model and demonstrate that it continues to resemble the original patient tumor histologically and immunohistochemically, maintains nuclear expression of brachyury, and is highly concordant with the original patient tumor by whole genome genotyping. Pathway analysis of this xenograft demonstrates activation of epidermal growth factor receptor (EGFR). In vitro studies demonstrate that two small molecule inhibitors of EGFR, erlotinib and gefitinib, inhibit proliferation of the chordoma cell line U-CH 1. We further demonstrate that erlotinib significantly inhibits chordoma growth in vivo. Evaluation of tumors post-treatment reveals that erlotinib reduces phosphorylation of EGFR. This is the first demonstration of antitumor activity in a patient-derived chordoma xenograft model and these findings support further evaluation of EGFR inhibitors in this disease.     

Pseudomonas aeruginosa Inhibits the Growth of Scedosporium and Lomentospora In Vitro

Mycopathologia - In vitro bacterial–fungal interaction studies in cystic fibrosis (CF) have mainly focused on interactions between bacteria and Candida. Here we investigated the effect of...     

Supernatant from Cultured Intestinal Cells Inhibits the Growth of Gram-Negative Bacteria

Bacterial growth chambers of Transwell units bearing intestinal epithelial monolayers (C2BBe) was consistently observed to be stagnant during the course of transmigration studies with Salmonella typhi. This limitation could not be explained by varying the bacterial load in the inoculum. Conditioned media produced by cultured C2BBe cells would not support bacterial growth. Growth support of the media was restored by heating to 95 C for 10 min. C2BBe conditioned media had bacteriostatic activity for a large variety of gram-negative, enteropathogenic bacteria but had no effect on gram-positive bacteria.     

Blockade of the SNARE Protein Syntaxin 1 Inhibits Glioblastoma Tumor Growth

Glioblastoma (GBM) is the most prevalent adult brain tumor, with virtually no cure, and with a median overall survival of 15 months from diagnosis despite of the treatment. SNARE proteins mediate membrane fusion events in cells and are essential for many cellular processes including exocytosis and neurotransmission, intracellular trafficking and cell migration. Here we show that the blockade of the SNARE protein Syntaxin 1 (Stx1) function impairs GBM cell proliferation. We show that Stx1 loss-of-function in GBM cells, through ShRNA lentiviral transduction, a Stx1 dominant negative and botulinum toxins, dramatically reduces the growth of GBM after grafting U373 cells into the brain of immune compromised mice. Interestingly, Stx1 role on GBM progression may not be restricted just to cell proliferation since the blockade of Stx1 also reduces in vitro GBM cell invasiveness suggesting a role in several processes relevant for tumor progression. Altogether, our findings indicate that the blockade of SNARE proteins may represent a novel therapeutic tool against GBM.     

PHA-767491抑制多种肿瘤细胞生长

Cdc7激酶抑制剂PHA-767491是最新发现的一类抗肿瘤新药.本实验利用不同浓度的PHA-767491对肿瘤细胞进行抑制研究.实验结果显示,PHA-767491对肿瘤细胞有很强的生长抑制作用,且抑制效果随着药物浓度或时间的增加而增强;通过和化疗药物5-氟尿嘧啶对比发现,PHA-767491只需较低剂量就能发挥出抑制肿瘤的作用,且疗效远高于5-氟尿嘧啶.研究进一步还发现,PHA-767491可通过促使PARP和casepase3蛋白的剪切诱导肿瘤细胞凋亡,同时PHA-767491还可以引起肿瘤细胞自噬.综上研究表明,PHA-767491可以通过诱导细胞凋亡和引起细胞自噬作用对多种肿瘤细胞有较好的治疗效果,而对正常细胞毒性很低.因此该实验研究为今后抗肿瘤新药PHA-767491的进一步应用于癌症的临床治疗提供了重要的实验依据.     

Ivermectin Inhibits Growth of Chlamydia trachomatis in Epithelial Cells

Ivermectin is currently approved for treatment of both clinical and veterinary infections by nematodes, including Onchocerca cervicalis in horses and Onchocerca volvulus in humans. However, ivermectin has never been shown to be effective against bacterial pathogens. Here we show that ivermectin also inhibits infection of epithelial cells by the bacterial pathogen, Chlamydia trachomatis, at doses that could be envisioned clinically for sexually-transmitted or ocular infections by Chlamydia.