Structure at restriction endonuclease MboI cleavage sites protected by actinomycin D or distamycin A

Restriction endonuclease MboI cleavage of DNA was inhibited by actinomycin D and distamycin A. The two inhibitors protected different subsets of the 8 cleavage sites in polyoma DNA. The cleavage reactions were analyzed both in the presence of minimal inhibitory concentrations of the compounds and at higher concentrations, allowing cleavage at only 1 site/DNA molecule. The experiments showed that cleavage sites most efficiently protected by actinomycin D had putative inhibitor binding sites at a distance of 1-2 base pairs from the MboI recognition sequence. Distamycin A, in contrast, apparently has to bind immediately adjacent to the MboI recognition sequence to protect from cleavage.     

Specific effect of manganese on the allantoinase of Vigna Radiata

Vigna radiata seedlings germinated in the presence of Mn²⁺ show an unusual increase in allantoinase activity which is proportional to Mn²⁺ concentration up to 5 mM. Though Mn²⁺ is not an activator for V. radiata allantoinase, it specifically protects allantoinase against thermal as well as papain-catalysed inactivation. Evidence is presented to show that the primary effect of Mn²⁺ is a protective one, both in vitro and in vivo, and that this is reflected in the observed enhancement of allantoinase activity in Mn²⁺ grown seedlings. That this unusual effect of Mn²⁺ is a specific one is indicated by the lack of a similar effect with Mg²⁺. Cu²⁺ is shown to destabilize V. radiata allantoinase in vitro as well as in vivo.     

Specific protection from phenocopy induction by heat shock

Mild heat treatments applied to whole animals or cell cultures of Drosophila prior to lethal heat shocks result in increased survival and protection against phenocopy induction. The optimal condition for the preliminary mild heat treatment is that which induces the synthesis of heat-shock proteins but does not turn off the protein synthesis that is in progress. Recovery of protein synthesis but not RNA synthesis following a drastic heat shock is much enhanced by the pretreatments. The results suggest that the protection for survival and against phenocopy induction is due to storage of messenger RNA.     

Diagnostic accuracy of fine-needle aspiration cytology in histological grade 1 breast carcinomas: are we good enough?

Background: Fine-needle aspiration cytology (FNAC) of both palpable and non-palpable breast carcinomas has a high accuracy and sensitivity in dedicated centres. It is generally thought that low-grade carcinomas have a distinctly lower sensitivity due to discrete cellular atypia that may be difficult to appreciate. Grade 1 carcinomas make up about 45% of screening-detected breast carcinomas and about 20% of symptomatic breast cancers. The aim of this study was to evaluate the diagnostic sensitivity of grade 1 carcinomas and identify the critical features in the cytological diagnostic work-up of these tumours. Methods: There were FNAC smears from 494 histologically confirmed grade 1 carcinomas diagnosed during 1996–2004. The cytological diagnoses were compared with the histology. Results: A definitive malignant diagnosis (absolute sensitivity) was given in 382 cases (77.3%). Equivocal or suspicious diagnoses were given in 75 (15.2%), benign or probably benign (false negative) in 24 (4.8%). Thirteen cases (2.6%) were unsatisfactory. Complete sensitivity was 92.7%. Invasive ductal carcinomas comprised 81.3% of all cases; absolute sensitivity for these was 80.9%. Invasive lobular and tubular carcinomas comprised 7.3% and 5.9% of cases, respectively; absolute sensitivity for these diagnosis was 50.0% and 57.1%, respectively, significantly lower than for other subtypes (P ≤ 0.0001) whereas the difference for complete sensitivity was less but still significant (P = 0.017). Absolute and complete sensitivities were lower for tumours less than 1 cm size compared with more than 1 cm (P ≤ 0.00001). Conclusion: Preoperative FNAC diagnosis of grade 1 breast carcinoma has a high sensitivity, especially in ductal carcinomas. Invasive lobular and tubular carcinomas were less likely to receive a definite preoperative diagnosis. The main reason for not reaching a definitive malignant diagnosis was sampling error due to small tumours less than 1 cm in diameter, irrespective of tumour subtype.     

Proteasome-dependent degradation of replisome components regulates faithful DNA replication

The replication machinery, or the replisome, collides with a variety of obstacles during the normal process of DNA replication. In addition to damaged template DNA, numerous chromosome regions are considered to be difficult to replicate owing to the presence of DNA secondary structures and DNA-binding proteins. Under these conditions, the replication fork stalls, generating replication stress. Stalled forks are prone to collapse, posing serious threats to genomic integrity. It is generally thought that the replication checkpoint functions to stabilize the replisome and replication fork structure upon replication stress. This is important in order to allow DNA replication to resume once the problem is solved. However, our recent studies demonstrated that some replisome components undergo proteasome-dependent degradation during DNA replication in the fission yeast Schizosaccharomyces pombe. Our investigation has revealed the involvement of the SCF^Pof3 (Skp1-Cullin/Cdc53-F-box) ubiquitin ligase in replisome regulation. We also demonstrated that forced accumulation of the replisome components leads to abnormal DNA replication upon replication stress. Here we review these findings and present additional data indicating the importance of replisome degradation for DNA replication. Our studies suggest that cells activate an alternative pathway to degrade replisome components in order to preserve genomic integrity.     

Evolutionary trade-offs at the Arabidopsis WRR4A resistance locus underpin alternate Albugo candida race recognition specificities

The oomycete Albugo candida causes white rust of Brassicaceae, including vegetable and oilseed crops, and wild relatives such as Arabidopsis thaliana. Novel White Rust Resistance (WRR) genes from Arabidopsis enable new insights into plant/parasite co-evolution. WRR4A from Arabidopsis accession Columbia (Col-0) provides resistance to many but not all white rust races, and encodes a nucleotide-binding, leucine-rich repeat immune receptor. Col-0 WRR4A resistance is broken by AcEx1, an isolate of A. candida. We identified an allele of WRR4A in Arabidopsis accession Øystese-0 (Oy-0) and other accessions that confers full resistance to AcEx1. WRR4A^Oy-0 carries a C-terminal extension required for recognition of AcEx1, but reduces recognition of several effectors recognized by the WRR4A^Col-0 allele. WRR4A^Oy-0 confers full resistance to AcEx1 when expressed in the oilseed crop Camelina sativa.     

MT2A对脂多糖介导的人肺毛细血管内皮细胞损伤的保护作用

目的:探索不同浓度的金属硫蛋2A(Metallothionein 2A, MT2A)对脂多糖(Lipopolysaccharid, LPS)介导的人肺微血管内皮细胞损伤的保护作用。方法:培养人肺毛细血管内皮细胞株(Human lung microvascular endothelial cells, HPMVECs),经过一定浓度LPS溶液进行刺激后,利用不同浓度的MT2A与对照组共同培养,一段时间后观察炎性介质IL-6、TNF-α释放的量及荧光显微镜观察组HPMVECs骨架形态变化。结果:各组TNF-α浓度均在0 h最低,随之逐渐升高,到6 h达到高峰;从各时间点来看,除0 h各组TNF-α浓度无显著差异外(F=0.717, P=0.549),其余各时间点B1、B2、B3均显著高于A组(均P0.05)。各组中IL-6浓度均在0 h最低,A组在2 h达到高峰,随后逐渐下降;B1组在4 h达到高峰,随之下降;B2、B3组从0 h开始逐渐升高,到6 h达到峰值;从各时间点来看,除0 h各处理因素无显著差异外(F=2.341, P=0.092),其余各时间点B1、B2、B3均低于A组(均P0.05)。A组6 h小时后纤维状肌动蛋白(F-actin)明显解聚,分布明显减少,应力纤维排列紊乱或者消失;B1组、B2组、B3组6 h小时后,与A组相比,F-actin的分布明显较多,应力纤维排列较为整齐。结论:LPS刺激人肺毛细血管内皮细胞有明显损伤效应,加入MT2A后细胞相关炎性因子释放量、细胞骨架损伤情况明显减轻,表明一定浓度的MT2A对LPS介导的肺毛细血管损伤有明显保护作用。