Nucleolytic degradation of 3′-ending overhangs is essential for DNA-end resection in RecA-loading deficient recB mutants of Escherichia coli

Degradation of a 5′-ending strand is the hallmark of the universal process of DNA double strand break (DSB) resection, which results in creation of the central recombination intermediate, a 3′-ending overhang. Here we show that in Escherichia coli recB1080/recB1067 mutants, which are devoid of RecBCD's nuclease and RecA loading activities, degradation of the unwound 3′ tail is as essential as is degradation of its 5′-ending complement. Namely, a synergistic action of ExoI, ExoVII, SbcCD and ExoX single-strand specific exonucleases (ssExos) of 3′-5′ polarity was essential for preserving cell viability, DNA repair and homologous recombination in the recB1080/recB1067 mutants, to the same extent as the redundant action of 5′-tail trimming ssExos RecJ and ExoVII. recB1080 derivatives lacking 3′-5′ ssExos also showed a strong induction of the SOS response and greatly increased SOS-dependent mutagenesis. Furthermore, we show that ExoI and ExoVII ssExos act synergistically in suppressing illegitimate recombination in the recB1080 mutant but not in a wt strain, while working in concert with the RecQ helicase. Remarkably, 3′-5′ ssExos show synergism with RecQ helicase in the recB1080 mutant in all the assays tested. The effect of inactivation of 3′-5′ ssExos in the recB1080/recB1067 mutants was much stronger than in wt, recD, and recB strains. These results demonstrate that the presence of a long, reactive 3′ overhang can be as toxic for a cell as its complete absence, i.e. it may prevent DSB repair. Our results indicate that coupling of helicase and RecA-loading activity during dsDNA-end resection is crucial in avoiding the deleterious effects of a long and stabile 3′ tail in E. coli.     

DNA End Resection: Nucleases Team Up with the Right Partners to Initiate Homologous Recombination

The repair of DNA double-strand breaks by homologous recombination commences by nucleolytic degradation of the 5′-terminated strand of the DNA break. This leads to the formation of 3′-tailed DNA, which serves as a substrate for the strand exchange protein Rad51. The nucleoprotein filament then invades homologous DNA to drive template-directed repair. In this review, I discuss mainly the mechanisms of DNA end resection in Saccharomyces cerevisiae, which includes short-range resection by Mre11-Rad50-Xrs2 and Sae2, as well as processive long-range resection by Sgs1-Dna2 or Exo1 pathways. Resection mechanisms are highly conserved between yeast and humans, and analogous machineries are found in prokaryotes as well.     

The dystonia gene THAP1 controls DNA double-strand break repair choice

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Control of DNA end resection by yeast Hmo1p affects efficiency of DNA end-joining

The primary pathways for DNA double strand break (DSB) repair are homologous recombination (HR) and non-homologous end–joining (NHEJ). The choice between HR and NHEJ is influenced by the extent of DNA end resection, as extensive resection is required for HR but repressive to NHEJ. Conversely, association of the DNA end-binding protein Ku, which is integral to classical NHEJ, inhibits resection. In absence of key NHEJ components, a third repair pathway is exposed; this alternative-end joining (A-EJ) is a highly error-prone process that uses micro-homologies at the breakpoints and is initiated by DNA end resection. In Saccharomyces cerevisiae, the high mobility group protein Hmo1p has been implicated in controlling DNA end resection, suggesting its potential role in repair pathway choice. Using a plasmid end-joining assay, we show here that absence of Hmo1p results in reduced repair efficiency and accuracy, indicating that Hmo1p promotes end-joining; this effect is only observed on DNA with protruding ends. Notably, inhibition of DNA end resection in an hmo1Δ strain restores repair efficiency to the levels observed in wild-type cells. In absence of Ku, HMO1 deletion also reduces repair efficiency further, while inhibition of resection restores repair efficiency to the levels observed in kuΔ. We suggest that Hmo1p functions to control DNA end resection, thereby preventing error-prone A-EJ repair and directing repairs towards classical NHEJ. The very low efficiency of DSB repair in kuΔhmo1Δ cells further suggests that excessive DNA resection is inhibitory for A-EJ.     

eIF-4E 和MMP9 在子宫内膜癌中的表达及临床意义

目的:探讨真核翻译起始因子4E(e IF-4E)和基质金属蛋白酶9(MMP9)在子宫内膜癌组织中的表达及其临床意义。方法:应用免疫组化法检测e IF-4E和MMP9在16例正常子宫内膜组织、10例增生子宫内膜组织及42例子宫内膜癌组织中的表达,分析其与子宫内膜癌临床病理之间的关系,并研究二者的相关性。结果:e IF-4E与MMP9在三组组织中的表达阳性率逐渐增加,三组间的差异有统计学意义(P0.05),两两比较,正常内膜和内膜癌之间的差异有统计学意义(P0.0167)。e IF-4E和MMP9的表达与子宫内膜癌的FIGO分期、组织学分级及淋巴结转移具有显著相关性(P0.05);子宫内膜癌组织中e IF-4E与MMP9的阳性表达呈正相关(r=0.717,P0.05)。结论:e IF-4E和MMP9在子宫内膜癌组织中表达显著上调,随FIGO分期的增加、组织学分级的增高及有淋巴结转移而增高,可能与子宫内膜癌的发生发展相关,二者的联合检测对于评估子宫内膜癌患者的病情及预后有重要的参考意义。     

内镜黏膜切除术治疗低位直肠侧向发育型肿瘤随机对照研究

目的:探讨内镜黏膜切除术治疗低位直肠侧向发育型肿瘤的临床效果。方法:收集我院收治的低位直肠侧向发育型肿瘤患者40例,随机分为对照组和实验组,每组各20例,对照组患者给予常规内镜下粘膜切除术,实验组患者给予内镜反转黏膜切除术,治疗结束后,对所有患者的病变残留例数、手术时间以及住院时间、并发症发生情况、手术切除效果进行检测并比较。结果:与对照组患者相比,实验组患者复发率较低(P0.05),手术时间以及住院时间、并发症发生率较低(P0.05);两组患者切除效果相比较,实验组分次切除、肿瘤残留例数患者较少,完全切除患者例数较多(P0.05),两组患者的整块切除例数无差异(P0.05)。结论:内镜反转黏膜切除术对于低位直肠侧向发育型肿瘤患者的临床疗效优于常规内镜下粘膜切除术,对临床有指导意义。     

Modulating effects of vanillic acid on circadian pattern of indices of redox homeostasis in N-Methly-N′-Nitro-N-Nitrosoguanidine induced endometrial carcinoma in rats

The circadian timing system controls drug metabolism and cellular proliferation over the 24-h period through molecular clocks in every cell. Accumulating epidemiological and genetic evidences indicate that the disruption of circadian rhythms might be directly linked to cancer. This study evaluates the effect of vanillic acid on the circadian rhythms of circulatory lipid peroxidation and antioxidant status during N-Methyl-N′-Nitro-N-Nitrosoguanidine (MNNG)-induced endometrial carcinoma in rats. The characteristics of circadian rhythms (acrophase, amplitude and mesor) of lipid peroxidation products – thiobarbituric acid reactive substances (TBARS), lipid hydroperoxides (LOOH) and enzymatic antioxidants like superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and non-enzymatic antioxidants such as reduced glutathione (GSH), vitamins C and E were markedly declined in MNNG-treated rats when compared to other groups. Pre and co-treatment of vanillic acid to MNNG-treated animals significantly increased the mesor and altered amplitudes of antioxidants and significantly decreased the mesor values of TBARS and LOOH. Further, delays in acrophase in MNNG-induced rats were reversed by vanllic acid administration. Thus, oral treatment of vanillic acid results in normalization of the altered rhythms of these indices of redox homeostasis (compared to controls) by its anticarcinogenic, cytoprotective and antioxidant effects.     

子宫内膜异位症合并不孕的原因及治疗方案探讨

目的：探讨子宫内膜异位症并不孕的原因及治疗方案的临床意义。方法：随机选取2007年6月至2012年8月就诊于我院的不孕症或疑似子宫内膜异位症（EMS）经检查确诊为EMS的患者216例,按照用药区别平均分为六组,分别为三苯氧胺组、米非司酮组、丹那唑组、诺雷德组、无效成分安慰剂组及两阶段复合治疗组,记录上述患者经治疗后随访1年内受孕率情况。结果：无效成分安慰剂组无疗效,各方面与其他组比较有明显差异（P〈0.05）,其余各组疗效相比无明显差异（P〉0.05）;三苯氧胺组与丹那唑组不良反应发生率高于其他组,比较有明显差异（P〈0.05）;两阶段复合治疗组的复发率最低,受孕率最高,除安慰剂组外其他组之间比较无明显差异（P〉0.05）。结论：此次研究的药物治疗效果近似,但是副作用、复发率及受孕率具有差异,临床应根据患者具体情况施治。     

Predictive factors for biochemical pregnancy in intracytoplasmic sperm injection cycles

The aim of this study was to investigate which factors contribute to the incidence of biochemical pregnancy (BP) in intracytoplasmic sperm injection (ICSI) cycles. This cohort study included cycles performed from June 2010 to September 2016 in a private, university-affiliated IVF centre. Cycles were split into four groups, depending on the pregnancy outcomes: Clinical Pregnancy (CP, n?=?903), Biochemical Pregnancy (BP, n?=?55), Miscarriage (MI, n?=?142) and Negative Pregnancy (NP, n?=?2034). The effects of ovarian stimulation, laboratory data and seminal parameters on pregnancy outcomes were evaluated using adjusted general linear models. Discriminant analyses were conducted to construct a model for pregnancy prediction and to establish cut-offs for BP. The total sperm count (p?=?0.035), total and progressive sperm motility (p?=?0.001 and p?=?0.023, respectively), total motile sperm count (TMSC, p?=?0.029) and the endometrial thickness (p?<?0.001) were lower among BP group cycles. Lower rates of high-quality cleavage-stage embryos were observed in the BP group compared to CP and MI groups (p?<?0.001). In discriminant analyses, cut-offs for BP prediction were established for the following factors: endometrial thickness < 11?mm, sperm motility < 55.5% and total dose of follicle-stimulating hormone (FSH)> 2400 IU. The incidence of biochemical pregnancy was four times higher when the aforementioned factors did not meet the defined cut-offs. The combination of suboptimal endometrial development and poor seminal and embryo quality contribute to an increased incidence of biochemical pregnancy in ICSI cycles.     

The role of progesterone elevation in IVF

Elevation of progesterone during the late follicular phase of stimulated in-vitro fertilization cycles is a frequent event, which negatively impacts the outcome. Over the years evidence has demonstrated a direct relationship between late-follicular elevated progesterone and endometrial receptivity. In this regard, elective cryopreservation of all good quality embryos and transfer in a subsequent frozen/thawed cycle is the most common strategy adopted by clinicians in case of elevated progesterone. Nonetheless, recent evidence suggests that elective cryopreservation might not entirely resolve the reduced pregnancy outcomes associated with the elevation of progesterone, considering that the increase may affect not only implantation, but also embryo quality.