衣原体调控宿主细胞凋亡的机制研究进展

衣原体是一类专性细胞内寄生的原核微生物,常引起性传播疾病、失明和肺炎等疾病。衣原体可能已经进化出调控宿主细胞的若干机制,通过改变宿主细胞蛋白位置,干扰宿主细胞凋亡信号通路等来抑制细胞凋亡,维持其自身的存活,核转录因子NF-κB也涉及到衣原体感染细胞凋亡抑制;衣原体可以经半胱氨酸蛋白酶依赖途径等机制诱导细胞凋亡,进而感染邻近细胞。     

蓝细菌病毒A-4L在鱼腥藻(Anabaena variabilis)藻苔中形成同心圆噬斑的成因

摘要:【目的】分析蓝细菌病毒A-4L在鱼腥藻(Anabaena sp.) PCC 7120藻苔中形成同心圆噬斑的原因,阐明A-4L的一个重要生物学特征,为分离、鉴定和筛选新的水华蓝藻病毒提供借鉴。【方法】用初始滴度为2.8×1010PFU/mL的A-4L悬液感染鱼腥藻,在不同时间点收集裂解液绘制一步生长曲线,获得A-4L对鱼腥藻的潜伏期和释放量。将适量A-4L悬液感染不同培养时间的藻苔,逐日观察和记录藻苔病变情况。培养藻苔并接种适量A-4L悬液,分别置于完全持续光照(Light:Dark=24 h:0 h,L:D=24 h:0 h)条件;完全周期光照(L:D=14 h:10h)条件;或前3 d周期光照转后3 d持续光照的条件下,比较不同光照条件对同心圆噬斑形成的影响。然后挑取单个噬斑进行扩大培养,纯化后,负染电镜观察A-4L的超微形态。【结果】A-4L的潜伏期为0.5－2h,释放量约为247 IU/cell (Infectious Units)。在周期光照条件下,藻苔接种A-4L 3－4 d后,出现同心圆噬斑,且同心圆数量与攻毒后的天数(n)有相关性,为“n－1”;同心圆间距约为3 mm。与周期光照条件相比,在持续光照条件下未形成同心圆噬斑。而在周期光照条件转持续光照条件下,由先周期光照时所形成的同心圆在转持续光照后逐渐消失,证实同心圆噬斑的形成依赖于周期光照。负染电镜观察显示A-4L具有一个近似球形的头部,直径约为50 nm以及长度约为10 nm的尾部,形态与短尾蓝细菌病毒相似。【结论】A-4L是一株能形成同心圆噬斑的蓝细菌病毒,并揭示其同心圆噬斑形成的关键条件是周期光照。     

The potato amylase inhibitor gene SbAI regulates cold‐induced sweetening in potato tubers by modulating amylase activity

Potato cold‐induced sweetening (CIS) is critical for the postharvest quality of potato tubers. Starch degradation is considered to be one of the key pathways in the CIS process. However, the functions of the genes that encode enzymes related to starch degradation in CIS and the activity regulation of these enzymes have received less attention. A potato amylase inhibitor gene known as SbAI was cloned from the wild potato species Solanum berthaultii. This genetic transformation confirmed that in contrast to the SbAI suppression in CIS‐resistant potatoes, overexpressing SbAI in CIS‐sensitive potatoes resulted in less amylase activity and a lower rate of starch degradation accompanied by a lower reducing sugar (RS) content in cold‐stored tubers. This finding suggested that the SbAI gene may play crucial roles in potato CIS by modulating the amylase activity. Further investigations indicated that pairwise protein–protein interactions occurred between SbAI and α‐amylase StAmy23, β‐amylases StBAM1 and StBAM9. SbAI could inhibit the activities of both α‐amylase and β‐amylase in potato tubers primarily by repressing StAmy23 and StBAM1, respectively. These findings provide the first evidence that SbAI is a key regulator of the amylases that confer starch degradation and RS accumulation in cold‐stored potato tubers.     

Melatonin inhibits triple-negative breast cancer progression through the Lnc049808-FUNDC1 pathway

Melatonin has been reported to have tumor-suppressive effects via comprehensive molecular mechanisms, and long non-coding RNAs (lncRNAs) may participate in this process. However, the mechanism by which melatonin affects the function of lncRNAs in triple-negative breast cancer (TNBC), the most aggressive subtype of breast cancer, is still unknown. Therefore, we aimed to investigate the differentially expressed mRNAs and lncRNAs in melatonin-treated TNBC cells and the interaction mechanisms. Microarray analyses were performed to identify differentially expressed mRNAs and lncRNAs in TNBC cell lines after melatonin treatment. To explore the functions and underlying mechanisms of the mRNAs and lncRNAs candidates, a series of in vitro experiments were conducted, including CCK-8, Transwell, colony formation, luciferase reporter gene, and RNA immunoprecipitation (RIP) assays, and mouse xenograft models were established. We found that after melatonin treatment, FUNDC1 and lnc049808 downregulated in TNBC cell lines. Knockdown of FUNDC1 and lnc049808 inhibited TNBC cell proliferation, invasion, and metastasis. Moreover, lnc049808 and FUNDC1 acted as competing endogenous RNAs (ceRNAs) for binding to miR-101. These findings indicated that melatonin inhibited TNBC progression through the lnc049808-FUNDC1 pathway and melatonin could be used as a potential therapeutic agent for TNBC.Subject terms: Breast cancer, Non-coding RNAs     

PIK-75 promotes homology-directed DNA repair

Homo!ogy-directed repair(HDR)is one of two major DNA repair pathways to mend the double-strand breaks(DSBs)formed in the genome(Liang et al.,1998;Pardo et al.,2009).Although less efficient compared with another DNA repair pathway,nonhomologous end joining(NHEJ),HDR is a type of precise repair to restore DNA damage and sustain genomic stability(Pardo et al.,2009;Ceccaldi et al.,2016).By contrast,NHEJ usually introduces mutations into the repaired site,thus probably harming the genomic integrity(Lieber et al.,2003).The error-free property enables HDR to be harnessed to correct a faulty mutation for therapeutic purpose in cells or in the body(Wu et al.,2013).In add让ion,HDR possesses great potential in the generation of genome-edited animals with precise genetic modifications,such as point mutation,DNA replacement,and DNA insertion in a specific genomic site(Wang et al.,2013).However,the low repair frequency mediated by HDR significantly limits让s application for efficient gene correction or establishment of various genetically modified animal models.Currently,multiple site-specific endonucleases have emerged as highly efficient tools to create targeted DSBs and markedly promote subsequent DNA repair either via HDR or NHEJ(Gaj et al.,2013).Nonetheless,the HDR-mediated modifications following the cleavage of engineering nucleases are still inefficient,usually with an efficiency less than 20%in cultured mammalian cells and embryos(Mali et al..2013;Wang et al.,2013;Yang et al.,2013).     

Heat shock protein 90 and tyrosine kinase regulate eNOS NO* generation but not NO* bioactivity

An increase in the association of heat shock protein 90 (HSP90) with endothelial nitric oxide (NO) synthase (eNOS) is well recognized for increasing NO (NO*) production. Despite the progress in this field, the mechanisms by which HSP90 modulates eNOS remain unclear due, in part, to the fact that geldanamycin (GA) redox cycles to generate superoxide anion (O(2)(-*) and the fact that inhibiting HSP90 with GA or radicicol (RAD) destabilizes tyrosine kinases that rely on the chaperone for maturation. In this report, we determine the extent to which these side effects alter vascular and endothelial cell function in physiologically relevant systems and in cultured endothelial cells. Vascular endothelial growth factor (VEGF)-stimulated vascular permeability, as measured by Evans blue leakage in the ears of male Swiss mice in vivo, and acetylcholine-induced vasodilation of isolated, pressurized mandibular arterioles from male C57BL6 mice ex vivo were attenuated by N(omega)-nitro-L-arginine methyl ester (L-NAME), GA, and RAD. Z-1[N-(2-aminoethyl)-N-(2-ammonoethyl)amino]diazen-1-ium-1,2-dioate (DETA-NONOate), a slow releasing NO. donor, increased vasodilation of arterioles pretreated with GA, RAD, and L-NAME equally well except at 10(-5) M, the highest concentration used, where vasodilation was greater in pressurized arterioles treated with L-NAME than in arterioles pretreated with GA or RAD alone. Both GA and RAD reduced NO* release from stimulated endothelial cell cultures and increased O(2)(-*) production in the endothelium of isolated aortas by an L-NAME-inhibitable mechanism. Pretreatment with RAD increased stimulated O(2)(-*) production from eNOS, whereas pretreatment with genistein (GE), a broad-spectrum tyrosine kinase inhibitor, did not; however, pretreatment with GE + RAD resulted in a super-induced state of uncoupled eNOS activity upon stimulation. These data suggest that the tyrosine kinases, either directly or indirectly, and HSP90-dependent signaling pathways act in concert to suppress uncoupled eNOS activity.     

Assessment of the effects of transgenic Bt cotton Bollgard II on the abundance of nontarget arthropods in Burkina Faso

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Quantitative profiling of drug-associated proteomic alterations by combined 2-nitrobenzenesulfenyl chloride (NBS) isotope labeling and 2DE/MS identification

The identification of drug-responsive biomarkers in complex protein mixtures is an important goal of quantitative proteomics. Here, we describe a novel approach for identifying such drug-induced protein alterations, which combines 2-nitrobenzenesulfenyl chloride (NBS) tryptophan labeling with two-dimensional gel electrophoresis (2DE)/mass spectrometry (MS). Lysates from drug-treated and control samples are labeled with light or heavy NBS moiety and separated on a common 2DE gel, and protein alterations are identified by MS through the differential intensity of paired NBS peptide peaks. Using NBS/2DE/MS, we profiled the proteomic alterations induced by tamoxifen (TAM) in the estrogen receptor (ER) positive MCF-7 breast cancer cell line. Of 88 protein spots that significantly changed upon TAM treatment, 44 spots representing 23 distinct protein species were successfully identified with NBS-paired peptides. Of these 23 TAM-altered proteins, 16 (70%) have not been previously associated with TAM or ER activity. We found the NBS labeling procedure to be both technically and biologically reproducible, and the NBS/2DE/MS alterations exhibited good concordance with conventional 2DE differential protein quantitation, with discrepancies largely due to the comigration of distinct proteins in the regular 2DE gels. To validate the NBS/2DE/MS results, we used immunoblotting to confirm GRP78, CK19, and PA2G4 as bona fide TAM-regulated proteins. Furthermore, we demonstrate that PA2G4 expression can serve as a novel prognostic factor for disease-free survival in two independent breast cancer patient cohorts. To our knowledge, this is the first report describing the proteomic changes in breast cancer cells induced by TAM, the most commonly used selective estrogen receptor modulator (SERM). Our results indicate that NBS/2DE/MS may represent a more reliable approach for cellular protein quantitation than conventional 2DE approaches.