Generation of rat-mouse chimeras by introducing single cells of rat inner cell masses into mouse blastocysts

正In the field of developmental biology and regenerative medicine,mammalian interspecific chimeras have been proved very useful for investigating early embryonic development and the immune system establishment,and extended to a promising potential for human organ generation(Rossant et al.,1982).In the early     

跨膜Ca~(2+)梯度对肌质网Ca~3+-ATP酶调节的特异性

我们曾报道跨膜Ca~(2+)梯度可通过膜脂影响肌质网Ca~(2+)-ATP 酶的构象和活性。本文就跨膜Ca~(2+)梯度对肌质网Ca~(2+)-ATP 酶的调节是否具有特异性作进一步研究。结果表明这种特异性表现在两方面:一是跨膜Ca~(2+)梯度对肌质网Ca~(2+)-ATP 酶功能的调节不能归结于跨膜Ca~(2+)浓度梯度所导致的膜电位的作用,离子载体FCCP 可消除跨膜电位但并不影响肌质网Ca~(2+)-ATP 酶的活力;二是其它二价金属离子如Sr~(2+)的跨膜梯度对肌质网Ca~(2+)-ATP 酶活力基本无影响。荧光偏振系列探剂n-AS 测定的结果表明跨膜Ca~(2+)与Sr~(2+)梯度对嵌有Ca~(2+)-ATP 酶的脂酶体的中部流动性的影响有较大差异。而Ca~(2+)-ATP 酶的Ca~(2+)结合位点正处于脂双层中部,这进一步提示膜脂参与了跨膜Ca~(2+)梯度对Ca~(2+)-ATP 酶的调节作用。     

Discovery of potent,low-absorbable sodium-dependent glucose cotransporter 1 (SGLT1) inhibitor SGL5213 for type 2 diabetes treatment

A new series of C-phenyl d-glucitol derivatives was designed and synthesized, and their SGLT1 inhibitory potency and absorbability were evaluated. We also investigated whether kidney drug retention could be avoided by creating molecules with different excretion pathways. To achieve a class of molecules with low absorption and that were excreted in bile, optimized synthesis was performed to bring the ClogP value and the topological polar surface area to within the appropriate ranges. Compounds 34d and 34j were poorly absorbed, but the absorbed compounds were mainly excreted in bile. Thus, smaller amounts of persistent residue in the kidneys were observed. Since 34d exerted a glucose-lowering effect at a dose of 0.3?mg/kg (p.o.) in SD rats, this compound (SGL5213) could be a clinical candidate for the treatment of type 2 diabetes.     

New SIRT2 inhibitors: Histidine-based bleomycin spin-off

Bleomycin is considered to exert its antitumor activity via DNA cleavage mediated by activated oxygen generated from the iron complex in its chelator moiety. Spin-offs from this moiety, HPH-1Trt and HPH-2Trt, with anti-cancer activities were recently synthesized. In this paper, we developed inhibitors of nicotinamide adenine dinucleotide-dependent deacetylase isoform 2 of Sirtuin protein (SIRT2), based on HPH-1Trt/HPH-2Trt, and aimed to generate new anti-cancer drugs. HPH-1Trt and HPH-2Trt had in vitro anti-SIRT2 inhibitory activity with 50% inhibitory concentration (IC₅₀) values of 5.5 and 8.8?μM, respectively. A structural portion of HPH-1Trt/HPH-2Trt, a tritylhistidine derivative TH-1, had stronger activity (IC₅₀?=?1.7?μM), and thus, fourteen derivatives of TH-1 were synthesized. Among them, TH-3 had the strongest activity (IC₅₀?=?1.3?μM). Selective binding of TH-3 in the pocket of SIRT2 protein was confirmed with a molecular docking study. Furthermore, TH-3 strongly lowered viability of the breast cancer cell line MCF7 with an IC₅₀ of 0.71?μM. A structure-activity relationship study using cell lines suggested that the mechanism of TH-3 to suppress MCF7 cells involves not only SIRT2 inhibition, but also another function. This compound may be a new candidate anti-cancer drug.     

Homozygous loss-of-function mutations in FSIP2 cause male infertility with asthenoteratospermia

<正>Male infertility, as a major issue of human reproduction health,prevents successful natural conception. Asthenoteratospermia mainly presents one or multiple anomalies in head, neck and tail of spermatozoa, and impairs sperm function and motility(Coutton et al., 2015). Recurrent abnormalities of the fibrous sheath     

Generation of two transgenic amphioxus lines using the Tol2 transposon system

正Amphioxus or lancelets are regarded as a promising model animals for studying developmental mechanisms in chordates, and the evolution of vertebrate characters, because of their important phylogenetic position and their genomic and anatomical simplicity(Bertrand and Escriva, 2011; Holland and Yu, 2004). Although several recent advances have made amphioxus more tractable as     

Mitochondrial genome regulates mitotic fidelity by maintaining centrosomal homeostasis

Centrosomes direct spindle morphogenesis to assemble a bipolar mitotic apparatus to enable error-free chromosome segregation and preclude chromosomal instability (CIN). Amplified centrosomes, a hallmark of cancer cells, set the stage for CIN, which underlies malignant transformation and evolution of aggressive phenotypes. Several studies report CIN and a tumorigenic and/or aggressive transformation in mitochondrial DNA (mtDNA)-depleted cells. Although several nuclear-encoded proteins are implicated in centrosome duplication and spindle organization, the involvement of mtDNA encoded proteins in centrosome amplification (CA) remains elusive. Here we show that disruption of mitochondrial function by depletion of mtDNA induces robust CA and mitotic aberrations in osteosarcoma cells. We found that overexpression of Aurora A, Polo-like kinase 4 (PLK4), and Cyclin E was associated with emergence of amplified centrosomes. Supernumerary centrosomes in rho0 (mtDNA-depleted) cells resulted in multipolar mitoses bearing “real” centrosomes with paired centrioles at the multiple poles. This abnormal phenotype was recapitulated by inhibition of respiratory complex I in parental cells, suggesting a role for electron transport chain (ETC) in maintaining numeral centrosomal homeostasis. Furthermore, rho0 cells displayed a decreased proliferative capacity owing to a G₂/M arrest. Downregulation of nuclear-encoded p53 in rho0 cells underscores the importance of mitochondrial and nuclear genome crosstalk and may perhaps underlie the observed mitotic aberrations. By contrast, repletion of wild-type mtDNA in rho0 cells (cybrid) demonstrated a much lesser extent of CA and spindle multipolarity, suggesting partial restoration of centrosomal homeostasis. Our study provides compelling evidence to implicate the role of mitochondria in regulation of centrosome duplication, spindle architecture, and spindle pole integrity.     

Identifying normal embryos from reciprocal translocation carriers by whole chromosome haplotyping

正Reciprocal translocation is a chromosomal structural abnormality that arises when two non-homologous chromosomes rearrange and attach with each other, an incidence that occurs in about 1/500to 1/625 newborns (Mackie and Scriven, 2002). This event typically does not lead to any significant loss of genetic material, thus reciprocal translocation carriers do not exhibit any severe abnormal     

愈伤组织形成过程中钙离子与激素诱导效应的关系

实验研究了在胡萝卜、烟草愈伤组织形成过程中,激素诱导作用与钙离子的关系。结果表明:在含有正常浓度的激素和Ca~(2+)的培养基中,0.1—1mmol/L Ca~(2+)螯合剂EGTA抑制愈伤组织鲜重增长78.0%—88.4%; 10—50μmol/L尼群地平及10—60μmol/L异博定等细胞膜钙通道阻断剂分别抑制愈伤组织鲜重增长19.1%—81.9%及17.6%—70.3%。除去上述Ca~(2+)螯合剂及Ca~(2+)通道阻断剂后,受抑制的外植体基本上可恢复生长。在无激素培养基中,10—30μmol/L Ca~(2+)载体A_(23187)可使外植体膨大,使外植体脱分化细胞增多,并出现分生细胞团,初步说明A_(23187)诱导的Ca~(2+)内流可以部分地模拟激素的作用。以膜显示剂氯四环素探测胞内Ca~(2+)分布时发现分裂细胞、脱分化细胞、分生细胞团及愈伤组织区域的细胞荧光较强。以上事实说明在愈伤组织形成中激素诱导效应与细胞内Ca~(2+)有密切关系。     

Control of flowering and inflorescence architecture in tomato by synergistic interactions between ALOG transcription factors

正Inflorescences are flower-bearing shoots that originate from pools of stem cells in shoot apical meristems (SAM).Inflorescence architecture is determined by a process of meristem maturation,during which stem cell fate switches from a vegetative to a reproductive growth program.A major factor in plant reproductive success in nature and yield in agriculture is the number of branches and flowers on inflorescences (Kobayashi and Weigel,2007;     

The N6-adenine methylation in yeast genome profiled by single-molecule technology

正The most common and abundant DNA modification is 5-methylcytosine(5mC),which has been well-established as an epigenetic mark regulating gene expression in eukaryotes(Jones,2012).Another DNA modification N~6-methyldeoxyadenosine(6mA),previously reported as a widespread DNA methylation in prokaryotes,     

Amelioration of mechanism-based inactivation of CYP3A4 by a H-PGDS inhibitor

This work describes the rational amelioration of mechanism-based inactivation (MBI) of Cytochrome P450 (CYP) 3A4 in a human hematopoietic prostaglandin D synthase (hH-PGDS) inhibitor (cpd 1). We utilized metabolism reports in order to check if patterns in the metabolism of 1 and similar compounds by CYP3A4 could be deciphered. Then we used structure based design, first modifying the CYP3A4 crystal structure (pdb code: 4NY4) by adding an oxyferryl moiety to the heme, followed by validating the modified structure to obtain the 1′ and 4 position oxidation products of midazolam and then recapitulating the metabolism patterns deciphered previously for 1 and analogs. We checked if the pattern deciphered could lead to a putative reactive moiety. Finally we used the docking pose of 1 into this model of the modified CYP3A4 crystal structure to guide transformation of 1 into MBI-free H-PGDS inhibitors.     

Structure-based design and biological evaluation of inhibitors of the pseudomonas aeruginosa heme oxygenase (pa-HemO)

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大麦根细胞质膜Ca~(2+)-ATP酶和Ca~(2+)转运系统的特性

用大麦质膜微囊研究细胞质膜 Ca~(2+)转运过程,发现质膜 Ca~(2+)—ATP酶在反应系统中不存在Mg~(2+)时可正常表现活性。跨膜Ca~(2+)转运按其对Mg~(2+)的需求可分为两个过程,一个是不需Mg~(2+)的、具高Ca~(2+)亲和力和较低的转运能力;另一个则是需Mg~(2+)的、具低Ca~(2+)亲和力和较高的转运能力。前者的动力学特征与Ca~(2+)—ATP酶相近,而后者则相差很大。据此推测,大麦根细胞质膜上除Ca~(2+)—ATP酶外,还存在另一个不同的Ca~(2+)转运系统。由两者分别承担的Ca~(2+)转运过程在细胞钙信使系统中可能起着不同的作用。     

Functional non-homologous end joining patterns triggered by CRISPR/Cas9 in human cells

正CRISPR/Cas9-mediated genome engineering technologies are now widely applied in various organisms,including mouse and human cells(Cong et al.,2013;Mali et al.,2013;Yang et al.,2013;Hsu et al.,2014).The most widely used customized CRISPR/Cas9(Sp Cas9)is derived from Streptococcus pyogenes(Cong et al.,2013).The CRISPR/Cas9 system creates site-specific double-     

Poly(ADP-ribosyl)ation associated changes in CTCF-chromatin binding and gene expression in breast cells

CTCF is an evolutionarily conserved and ubiquitously expressed architectural protein regulating a plethora of cellular functions via different molecular mechanisms. CTCF can undergo a number of post-translational modifications which change its properties and functions. One such modifications linked to cancer is poly(ADP-ribosyl)ation (PARylation). The highly PARylated CTCF form has an apparent molecular mass of 180?kDa (referred to as CTCF180), which can be distinguished from hypo- and non-PARylated CTCF with the apparent molecular mass of 130?kDa (referred to as CTCF130). The existing data accumulated so far have been mainly related to CTCF130. However, the properties of CTCF180 are not well understood despite its abundance in a number of primary tissues. In this study we performed ChIP-seq and RNA-seq analyses in human breast cells 226LDM, which display predominantly CTCF130 when proliferating, but CTCF180 upon cell cycle arrest. We observed that in the arrested cells the majority of sites lost CTCF, whereas fewer sites gained CTCF or remain bound (i.e. common sites). The classical CTCF binding motif was found in the lost and common, but not in the gained sites. The changes in CTCF occupancies in the lost and common sites were associated with increased chromatin densities and altered expression from the neighboring genes. Based on these results we propose a model integrating the CTCF130/180 transition with CTCF-DNA binding and gene expression changes. This study also issues an important cautionary note concerning the design and interpretation of any experiments using cells and tissues where CTCF180 may be present.     

Synthesis, in vitro and in silico evaluation of novel trans-stilbene analogues as potential COX-2 inhibitors

25 new trans-stilbene and trans-stilbazole derivatives were investigated using in vitro and in silico techniques. The selectivity and potency of the compounds were assessed using commercial ELISA test. The obtained results were incorporated into 2D QSAR assay. The most promising compound 4-nitro-3′,4′,5′-trihydroxy-trans-stilbene (N1) was synthetized and its potency and selectivity were confirmed. N1 was classified as preferential COX-2 inhibitor. Its ability to inhibit COX-2 in MCF-7 cell line was established and its cytotoxicity by MTT test was assessed. The compound was more cytotoxic than celecoxib within studied concentration range. Finally, the investigated trans-stilbene was docked into COX-1 and COX-2 active sites using “CDOCKER” protocol.     

Generating a reporter mouse line marking medium spiny neurons in the developing striatum driven by Arpp21 cis-regulatory elements

正The striatum, as the primary input nucleus in the basal ganglion,plays an important role in neural circuits crucial for the control of critical motivation, motor planning and procedural learning(Kreitzer and Malenka, 2008). Most cells in the striatum are GABAergic, including a large population (90%-95%) of medium spiny neurons (MSNs) and a small population of interneurons.