Comparison of histone modifications in in vivo and in vitro fertilization mouse embryos

Histone modifications are thought to play important roles in various cellular functions. In this article, the distribution patterns of acetylation on histone H4, methylation on histone H3 lysine 9, and phosphorylation on histone H3 serine 10 were examined in in vivo and in vitro fertilization (IVF) preimplantation mouse embryos by using indirect immunofluorescence and scanning confocal microscopy. We desired to know whether the IVF, which has been widely used as a routine assisted reproductive technology in animal and human, was safe at the epigenetic level. As results, we found that there was no difference in these histone modification patterns in in vivo and IVF mouse embryos from zygote to blastocyst stage. Moreover, these histone modifications had different distributions at all examined stages, but they were consistent with the mouse embryo developmental stages.     

SUMO Modification Regulates BLM and RAD51 Interaction at Damaged Replication Forks

The gene mutated in Bloom''s syndrome, BLM, is important in the repair of damaged replication forks, and it has both pro- and anti-recombinogenic roles in homologous recombination (HR). At damaged forks, BLM interacts with RAD51 recombinase, the essential enzyme in HR that catalyzes homology-dependent strand invasion. We have previously shown that defects in BLM modification by the small ubiquitin-related modifier (SUMO) cause increased γ-H2AX foci. Because the increased γ-H2AX could result from defective repair of spontaneous DNA damage, we hypothesized that SUMO modification regulates BLM''s function in HR repair at damaged forks. To test this hypothesis, we treated cells that stably expressed a normal BLM (BLM+) or a SUMO-mutant BLM (SM-BLM) with hydroxyurea (HU) and examined the effects of stalled replication forks on RAD51 and its DNA repair functions. HU treatment generated excess γ-H2AX in SM-BLM compared to BLM+ cells, consistent with a defect in replication-fork repair. SM-BLM cells accumulated increased numbers of DNA breaks and were hypersensitive to DNA damage. Importantly, HU treatment failed to induce sister-chromatid exchanges in SM-BLM cells compared to BLM+ cells, indicating a specific defect in HR repair and suggesting that RAD51 function could be compromised. Consistent with this hypothesis, RAD51 localization to HU-induced repair foci was impaired in SM-BLM cells. These data suggested that RAD51 might interact noncovalently with SUMO. We found that in vitro RAD51 interacts noncovalently with SUMO and that it interacts more efficiently with SUMO-modified BLM compared to unmodified BLM. These data suggest that SUMOylation controls the switch between BLM''s pro- and anti-recombinogenic roles in HR. In the absence of BLM SUMOylation, BLM perturbs RAD51 localization at damaged replication forks and inhibits fork repair by HR. Conversely, BLM SUMOylation relieves its inhibitory effects on HR, and it promotes RAD51 function.     

两株海洋真菌的鉴定及其次级代谢产物抑制烟草花叶病毒及抗肿瘤活性

摘要：【目的】通过对2株活性海洋真菌发酵产物提取物抑制烟草花叶病毒和抗肿瘤活性进行研究,为进一步得到活性纯品化合物作为抗病毒及抗肿瘤的先导化合物奠定基础。【方法】菌株发酵产物的粗提物是通过甲醇浸取并在真空条件下蒸干得到的。粗提物中溶于水的部分为水溶性部分,不溶于水的部分为脂溶性部分。通过间接酶联免疫法检测样品抑制烟草花叶病毒的活性,通过四甲基偶氮唑盐微量酶反应比色法（MTT法）检测样品抗肿瘤活性,通过形态及ITS rDNA序列法进行菌株鉴定。【结果】两株海洋真菌抑制烟草花叶病毒活性和抗肿瘤的活性均较高。分子鉴定结果显示,两株真菌分别与Penicillium oxalicum 和 Neosartorya fischeri 的同源性极高。菌株0312F1发酵液的水溶性部分具有抗病毒及抗肿瘤活性,菌株1008F1发酵液的脂溶性部分具有抑制烟草花叶病毒活性,而水溶性部分具有抗肿瘤活性。【结论】菌株0312F1和菌株1008F1发酵液的提取物抑制烟草花叶病毒的活性部位不同,而抗肿瘤活性部位相同。菌株0312F1发酵液提取物的水溶性活性部位对肝癌细胞BEL-7404的抑制效果比对胃癌细胞SGC-7901的抑制效果明显,而菌株1008F1发酵液提取物的水溶性活性部位对胃癌细胞SGC-7901的抑制效果比对肝癌细胞BEL-7404的抑制效果明显。     

Searching the Cytochrome P450 Enzymes for the Metabolism of Meranzin Hydrate: A Prospective Antidepressant Originating from Chaihu-Shugan-San

Meranzin hydrate (MH), an absorbed bioactive compound from the Traditional Chinese Medicine (TCM) Chaihu-Shugan-San (CSS), was first isolated in our laboratory and was found to possess anti-depression activity. However, the role of cytochrome P450s (CYPs) in the metabolism of MH was unclear. In this study, we screened the CYPs for the metabolism of MH in vitro by human liver microsomes (HLMs) or human recombinant CYPs. MH inhibited the enzyme activities of CYP1A2 and CYP2C19 in a concentration-dependent manner in the HLMs. The K_m and V_max values of MH were 10.3±1.3 µM and 99.1±3.3 nmol/mg protein/min, respectively, for the HLMs; 8.0±1.6 µM and 112.4±5.7 nmol/nmol P450/min, respectively, for CYP1A2; and 25.9±6.6 µM and 134.3±12.4 nmol/nmol P450/min, respectively, for CYP2C19. Other human CYP isoforms including CYP2A6, CYP2C9, CYP2D6, CYP2E1 and CYP3A4 showed minimal or no effect on MH metabolism. The results suggested that MH was simultaneously a substrate and an inhibitor of CYP1A2 and CYP2C9, and MH had the potential to perpetrate drug-drug interactions with other CYP1A2 and CYP2C19 substrates.     

Construction of a cross-species cell landscape at single-cell level

Individual cells are basic units of life. Despite extensive efforts to characterize the cellular heterogeneity of different organisms, cross-species comparisons of landscape dynamics have not been achieved. Here, we applied single-cell RNA sequencing (scRNA-seq) to map organism-level cell landscapes at multiple life stages for mice, zebrafish and Drosophila. By integrating the comprehensive dataset of > 2.6 million single cells, we constructed a cross-species cell landscape and identified signatures and common pathways that changed throughout the life span. We identified structural inflammation and mitochondrial dysfunction as the most common hallmarks of organism aging, and found that pharmacological activation of mitochondrial metabolism alleviated aging phenotypes in mice. The cross-species cell landscape with other published datasets were stored in an integrated online portal—Cell Landscape. Our work provides a valuable resource for studying lineage development, maturation and aging.     

Role of hypoxia in the hallmarks of human cancer

Hypoxia has been recognized as one of the fundamentally important features of solid tumors and plays a critical role in various cellular and physiologic events, including cell proliferation, survival, angiogenesis, immunosurveillance, metabolism, as well as tumor invasion and metastasis. These responses to hypoxia are at least partially orchestrated by activation of the hypoxia‐inducible factors (HIFs). HIF‐1 is a key regulator of the response of mammalian cells to oxygen deprivation and plays critical roles in the adaptation of tumor cells to a hypoxic microenvironment. Hypoxia and overexpression of HIF‐1 have been associated with radiation therapy and chemotherapy resistance, an increased risk of invasion and metastasis, and a poor clinical prognosis of solid tumors. The discovery of HIF‐1 signaling has led to a rapidly increasing understanding of the complex mechanisms involved in tumor hypoxia and has helped greatly in screening novel anticancer agents. In this review, we will first introduce the cellular responses to hypoxia and HIF‐1 signaling pathway in hypoxia, and then summarize the multifaceted role of hypoxia in the hallmarks of human cancers. J. Cell. Biochem. 107: 1053–1062, 2009. © 2009 Wiley‐Liss, Inc.     

Lactic Acid Production from Pretreated Hydrolysates of Corn Stover by a Newly Developed Bacillus coagulans Strain

An inhibitor-tolerance strain, Bacillus coagulans GKN316, was developed through atmospheric and room temperature plasma (ARTP) mutation and evolution experiment in condensed dilute-acid hydrolysate (CDH) of corn stover. The fermentabilities of other hydrolysates with B. coagulans GKN316 and the parental strain B. coagulans NL01 were assessed. When using condensed acid-catalyzed steam-exploded hydrolysate (CASEH), condensed acid-catalyzed liquid hot water hydrolysate (CALH) and condensed acid-catalyzed sulfite hydrolysate (CASH) as substrates, the concentration of lactic acid reached 45.39, 16.83, and 18.71 g/L by B. coagulans GKN316, respectively. But for B. coagulans NL01, only CASEH could be directly fermented to produce 15.47 g/L lactic acid. The individual inhibitory effect of furfural, 5-hydroxymethylfurfural (HMF), vanillin, syringaldehyde and p-hydroxybenzaldehyde (pHBal) on xylose utilization by B. coagulans GKN316 was also studied. The strain B. coagulans GKN316 could effectively convert these toxic inhibitors to the less toxic corresponding alcohols in situ. These results suggested that B. coagulans GKN316 was well suited to production of lactic acid from undetoxified lignocellulosic hydrolysates.     

氧化亚铁硫杆菌亚铁氧化系统的研究进展

氧化亚铁硫杆菌(Acidithiobacillus ferrooxidans)为无机化能自养菌,革兰氏阴性,能在极端酸性环境中生长.由于在生物冶金中的应用及特殊的生理学效应,该菌受到研究者的广泛关注.A.ferrooxidans能氧化亚铁、元素硫及还原态硫化物获得电子,并通过一系列电子载体将电子传递给氧生成水,同时释放能量供生命活动需要.目前对A.ferrooxidans电子传递系统的研究主要集中于亚铁氧化电子传递系统,已发现多种与亚铁氧化电子传递相关电子载体和操纵子,如电子载体铜蓝蛋白(Rustocyanin,Rus)、细胞色素C(Cytochrome C,Cyc)、细胞色素C氧化酶(Cytochrome Coxidase,Cox)、亚铁氧化酶(Iro)、细胞色素bc1复合物(cytochrome bc1 complex,bc1)等,以及rus操纵子和pet操纵子.综述了近年来有关A.ferrooxidans 亚铁氧化电子传递链相关蛋白载体,rus和pet操纵子结构与功能及表达调控等方面的研究进展.     

t-PA cDNA在CHO细胞中的高效稳定表达

我们曾报道t-PA mRNA非翻译区序列对其表达有明显的抑制作用,在此基础上,通过对5′-UTR及3′-UTR的改造,使t-PA在COS-7细胞中的表达水平提高30倍左右。将t-PA表达质粒用电击法转染中国仓鼠卵巢细胞二氢叶酸还原酶缺陷株(CHO-dhfr),经过混合加压及筛选,在CHO细胞中高效表达了t-PA,表达水平达到5000～6000 IU/10~6细胞/24hr。重组t-PA具有与天然t-PA相同的分子量及酶活性。经过8个月连续传代,表达水平未下降,表明细胞株是稳定的,其主要指标均符合工程细胞株的要求。