Ataxia-telangiectasia mutated (ATM) deficiency decreases reprogramming efficiency and leads to genomic instability in iPS cells

During cell division, one of the major features of somatic cell reprogramming by defined factors, cells are potentially exposed to DNA damage. Inactivation of the tumor suppressor gene p53 raised reprogramming efficiency but resulted in an increased number of abnormal chromosomes in established iPS cells. Ataxia-telangiectasia mutated (ATM), which is critical in the cellular response to DNA double-strand breaks, may also play an important role during reprogramming. To clarify the function of ATM in somatic cell reprogramming, we investigated reprogramming in ATM-deficient (ATM-KO) tail-tip fibroblasts (TTFs). Although reprogramming efficiency was greatly reduced in ATM-KO TTFs, ATM-KO iPS cells were successfully generated and showed the same proliferation activity as WT iPS cells. ATM-KO iPS cells had a gene expression profile similar to ES cells and WT iPS cells, and had the capacity to differentiate into all three germ layers. On the other hand, ATM-KO iPS cells accumulated abnormal genome structures upon continuous passages. Even with the abnormal karyotype, ATM-KO iPS cells retained pluripotent cell characteristics for at least 20 passages. These data indicate that ATM does participate in the reprogramming process, although its role is not essential.     

人卵子体外成熟过程中组蛋白乙酰化的动态变化模式

目的：组蛋白乙酰化与有丝分裂过程中的多个染色质相关事件有关,但是它在哺乳动物减数分裂过程中的作用仍不清楚。本研究通过观察人卵子体外成熟过程中不同阶段的组蛋白H3K9乙酰化变化模式,以探讨组蛋白乙酰化在减数分裂过程中的作用。方法：我们选择在我院进行单精子显微注射（Intracytoplasmicsperminjection,ICSI）的病人,共收集用于GV期卵子25个,MI期卵子28个用于本研究。将其中一部分直接用4％多聚甲醛固定,另一部分体外培养成熟至MII期,再用4％多聚甲醛固定。采用免疫荧光染色检测不同发育时期卵子的组蛋白H3K9乙酰化状态。结果：免疫荧光染色结果显示,GV期的卵子可检测到明显的H3K9乙酰化,MI期和MII期的卵子的H3K9乙酰化程度逐渐减弱。结论：人类卵子在成熟过程中会发生组蛋白H3K9乙酰化水平的逐渐降低,可能与减数分裂过程中特定的染色体分离、基因表达的重新编程密切相关。     

The way of the germline, its developmental cycle and epigenetic network

The long-awaited second edition of the International Symposium on "Epigenome Network, Development and Reprogramming of Germ Cells" hosted by Hiroyuki Sasaki took place at the Kyushu University School of Medicine, in Fukuoka, Japan from 22 to 24 November 2010. This meeting brought together again the crème de la crème of the Japanese research community investigating germline development, reprogramming and genetic networks as well as eminent international scientists. Novel trend concepts including the "reprogramming expressway", "canalization", "licensing", "epigenetic barrier", "flex points" and "hydroxymethylation" were introduced and discussed in the context of development, reprogramming and pluripotency.     

Proliferation Rate of Somatic Cells Affects Reprogramming Efficiency

The discovery of induced pluripotent stem (iPS) cells provides not only new approaches for cell replacement therapy, but also new ways for drug screening. However, the undefined mechanism and relatively low efficiency of reprogramming have limited the application of iPS cells. In an attempt to further optimize the reprogramming condition, we unexpectedly observed that removing c-Myc from the Oct-4, Sox-2, Klf-4, and c-Myc (OSKM) combination greatly enhanced the generation of iPS cells. The iPS cells generated without c-Myc attained salient pluripotent characteristics and were capable of producing full-term mice through tetraploid complementation. We observed that forced expression of c-Myc induced the expression of many genes involved in cell cycle control and a hyperproliferation state of the mouse embryonic fibroblasts during the early stage of reprogramming. This enhanced proliferation of mouse embryonic fibroblasts correlated negatively to the overall reprogramming efficiency. By applying small molecule inhibitors of cell proliferation at the early stage of reprogramming, we were able to improve the efficiency of iPS cell generation mediated by OSKM. Our data demonstrated that the proliferation rate of the somatic cell plays critical roles in reprogramming. Slowing down the proliferation of the original cells might be beneficial to the induction of iPS cells.     

Enriched Environment-induced Maternal Weight Loss Reprograms Metabolic Gene Expression in Mouse Offspring

The global prevalence of weight loss is increasing, especially in young women. However, the extent and mechanisms by which maternal weight loss affects the offspring is still poorly understood. Here, using an enriched environment (EE)-induced weight loss model, we show that maternal weight loss improves general health and reprograms metabolic gene expression in mouse offspring, and the epigenetic alterations can be inherited for at least two generations. EE in mothers induced weight loss and its associated physiological and metabolic changes such as decreased adiposity and improved glucose tolerance and insulin sensitivity. Relative to controls, their offspring exhibited improved general health such as reduced fat accumulation, decreased plasma and hepatic lipid levels, and improved glucose tolerance and insulin sensitivity. Maternal weight loss altered gene expression patterns in the liver of offspring with coherent down-regulation of genes involved in lipid and cholesterol biosynthesis. Epigenomic profiling of offspring livers revealed numerous changes in cytosine methylation depending on maternal weight loss, including reproducible changes in promoter methylation over several key lipid biosynthesis genes, correlated with their expression patterns. Embryo transfer studies indicated that oocyte alteration in response to maternal metabolic conditions is a strong factor in determining metabolic and epigenetic changes in offspring. Several important lipid metabolism-related genes have been identified to partially inherit methylated alleles from oocytes. Our study reveals a molecular and mechanistic basis of how maternal lifestyle modification affects metabolic changes in the offspring.     

mRNA changes during imaginal determination of the epidermal cells in the pupa ofGalleria mellonella L

Summary Changes in the mRNA population of the mesonotal epidermal cells were investigated inGalleria mellonella during the first 48 h after pupation. Total RNA was extracted and assayed by in vitro translation. The translational products were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and visualized by autoradiography. The changing banding pattern of the in vitro synthesized proteins indicates changes in the cellular pattern of mRNAs, most of which occur between 6 h and 18 h after pupal ecdysis. These changes mostly consist in the decrease or disappearance of bands. The injection of juvenile hormone (JH) immediately after pupal ecdysis does not qualitatively influence mRNA changes, but does alter their time course, for they are postponed for 6–12 h. After the injection of 20-hydroxyecdysone (20HE) the same changes can again be seen, but they are greatly accelerated. A comparison of these results with known data on the time course of reprogramming and ecdysteroid titre leads to the conclusion that the mRNA changes in the epidermal cells are a prerequisite for the renewed expression of a developmental programme. This is independent of whether, in the absence of JH, a new programme is determined or whether, under the influence of JH, the previous programme is restored. 20HE does not have any effect on the change in the developmental programme. The change seems to occur as an active and autonomous process in the epidermal cells, in accordance with a genetically fixed developmental programme.