WD repeat-containing protein 5, a ubiquitously expressed histone methyltransferase adaptor protein, regulates smooth muscle cell-selective gene activation through interaction with pituitary homeobox 2

WD repeat-containing protein 5 (WDR5) is a common component of mammalian mixed lineage leukemia methyltransferase family members and is important for histone H3 lysine 4 methylation (H3K4me), which has been implicated in control of activation of cell lineage genes during embryogenesis. However, WDR5 has not been considered to play a specific regulatory role in epigenetic programming of cell lineage because it is ubiquitously expressed. Previous work from our laboratory showed the appearance of histone H3K4me within smooth muscle cell (SMC)-marker gene promoters during the early stages of development of SMC from multipotential embryonic cells but did not elucidate the underlying mechanisms that mediate SMC-specific and locus-selective H3K4me. Results presented herein show that knockdown of WDR5 significantly decreased SMC-marker gene expression in cultured SMC differentiation systems and in Xenopus laevis embryos in vivo. In addition, we showed that WDR5 complexes within SMC progenitor cells contained H3K4 methyltransferase enzymatic activity and that knockdown of WDR5 selectively decreased H3K4me1 and H3K4me3 enrichment within SMC-marker gene promoter loci. Moreover, we present evidence that it is recruited to these gene promoter loci through interaction with a SMC-selective pituitary homeobox 2 (Pitx2). Taken together, studies provide evidence for a novel mechanism for epigenetic control of SMC-marker gene expression during development through interaction of WDR5, homeodomain proteins, and chromatin remodeling enzymes.     

Synthesis and cytotoxicity of 17a-aza-D-homo-androster-17-one derivatives

A series of 17a-aza-d-homo-andrester-17-one derivatives, bearing hydroxyl, hydroximino, carbonyl and thiosemicarbazido groups at the position-3 or position-6 of steroidal nucleus, were prepared and evaluated in vitro against two human cell lines (Hela (human cervical carcinoma) and SMMC 7404 (human liver carcinoma)). The results showed that these compounds could exhibit a high cytotoxicity to Hela tumor cell line, especially for compounds 8 and 12, the IC₅₀ values are 15.1 and 14.0 nmol/mL, respectively. Our findings could provide new evidence showing the relationship between the chemical structure and biological activity and may be useful for the discovery of new anti-cancer drugs.     

Organization of the smallest eukaryotic spindle

In metazoans, plants, and fungi, the spindle checkpoint delays mitosis until each chromosome is attached to one or more of its own kinetochore microtubules (kMTs). Some unicellular eukaryotes, however, have been reported to have fewer kMTs than chromosomes [1-5]. If this is the case, it is unclear how the spindle checkpoint could be satisfied. In the vast majority of the previous studies, mitotic cells were chemically fixed at room temperature, but this does not?always preserve dynamic and/or small structures like spindle MTs and kinetochores [6]. Indeed, later higher-resolution studies have reversed some earlier claims [7-11]. Here we show that in Ostreococcus tauri (the smallest eukaryote known), mitosis does involve fewer spindle microtubules than chromosomes. O.?tauri cultures were enriched for mitotic cells, high-pressure frozen, and then imaged in 3D both in plastic and in a near-native ("frozen-hydrated") state through electron tomography. Mitotic cells have a distinctive intranuclear heterochromatin-free "spindle tunnel" with approximately four short and occasionally one long, incomplete (unclosed) microtubule at each end of the spindle tunnel. Because other aspects of O.?tauri's spindle checkpoint seem typical, these data suggest that O.?tauri's 20 chromosomes are physically linked and segregated as just one or a small number of groups.     

The fission yeast Schizosaccharomyces pombe has two distinct tRNase Z(L)s encoded by two different genes and differentially targeted to the nucleus and mitochondria

tRNase Z is the endonuclease that is involved in tRNA 3'-end maturation by removal of the 3'-trailer sequences from tRNA precursors. Most eukaryotes examined to date, including the budding yeast Saccharomyces cerevisiae and humans, have a single long form of tRNase Z (tRNase ZL). In contrast, the fission yeast Schizosaccharomyces pombe contains two candidate tRNase ZLs encoded by the essential genes sptrz1+ and sptrz2+. In the present study, we have expressed recombinant SpTrz1p and SpTrz2p in S. pombe. Both recombinant proteins possess precursor tRNA 3'-endonucleolytic activity in vitro. SpTrz1p localizes to the nucleus and has a simian virus 40 NLS (nuclear localization signal)-like NLS at its N-terminus, which contains four consecutive arginine and lysine residues between residues 208 and 211 that are critical for the NLS function. In contrast, SpTrz2p is a mitochondrial protein with an N-terminal MTS (mitochondrial-targeting signal). High-level overexpression of sptrz1+ has no detectable phenotypes. In contrast, strong overexpression of sptrz2+ is lethal in wild-type cells and results in morphological abnormalities, including swollen and round cells, demonstrating that the correct expression level of sptrz2+ is critical. The present study provides evidence for partitioning of tRNase Z function between two different proteins in S. pombe, although we cannot rule out specialized functions for each protein.     

Electrochemical sensor for dopamine based on a novel graphene-molecular imprinted polymers composite recognition element

A novel composite of graphene sheets/Congo red-molecular imprinted polymers (GSCR-MIPs) was synthesized through free radical polymerization (FRP) and applied as a molecular recognition element to construct dopamine (DA) electrochemical sensor. The template molecules (DA) were firstly absorbed at the GSCR surface due to their excellent affinity, and subsequently, selective copolymerization of methacrylic acid (MAA) and ethylene glycol dimethacrylate (EGDMA) was further achieved at the GSCR surface. Potential scanning was presented to extract DA molecules from the imprinted polymers film, and as a result, DA could be rapidly and completely removed by this way. With regard to the traditional MIPs, the GSCR-MIPs not only possessed a faster desorption and adsorption dynamics, but also exhibited a higher selectivity and binding capacity toward DA molecule. As a consequence, an electrochemical sensor for highly sensitive and selective detection of DA was successfully constructed as demonstration based on the synthesized GSCR-MIPs nanocomposites. Under experimental conditions, selective detection of DA in a linear concentration range of 1.0 × 10(-7)-8.3 × 10(-4)M was obtained, which revealed a lower limit of detection and wider linear response compared to some previously reported DA electrochemical MIPs sensors. The new DA electrochemical sensor based on GSCR-MIPs composites also exhibited excellent repeatability, which expressed as relative standard deviation (RSD) was about 2.50% for 30 repeated analyses of 20 μM DA.     

Ⅰ型子宫内膜癌组织中HIF-1α、Snail与E-cadherin的表达及临床意义

目的探讨I型子宫内膜癌组织中HIF-1α的表达,以及调控Snail和E-cadherin对肿瘤侵袭性生物学行为的影响。方法采用免疫组化方法,结合组织芯片技术,检测124例I型子宫内膜癌、28例内膜不典型增生、35例正常内膜组织中HIF-1α、Snail和E-cadherin的表达水平,分析三种蛋白表达之间的相关性及与临床病理因素的关系。结果 I型子宫内膜癌组织中HIF-1α、Snail和E-cadherin的表达率分别为61.3%、46.8%、36.3%,与正常内膜和不典型增生内膜组织相比,有显著统计学差异(P<0.01)。HIF-1α表达与病理分级、肿瘤肌层浸润和淋巴结转移明显相关(P<0.05)。Snail表达与FIGO分期、淋巴结转移明显相关(P<0.05)。E-cadherin的缺失与肿瘤肌层浸润和淋巴结转移显著相关(P<0.01)。I型子宫内膜癌组织中HIF-1α和Snail的表达呈明显正相关(r=0.214,P=0.017),而Snail和E-cadherin的表达存在负相关关系(r=–0.203,P=0.024)。结论 HIF-1α可能通过上调Snail的表达和抑制E-cadherin的表达在I型子宫内膜癌发生、侵袭和转移中发挥重要作用。     

Deciphering the effects of PYCR1 on cell function and its associated mechanism in hepatocellular carcinoma

Overexpression of pyrroline-5-carboxylate reductase 1 (PYCR1) has been associated with the development of certain cancers; however, no studies have specifically examined the role of PYCR1 in hepatocellular carcinoma (HCC). Based on The Cancer Genome Atlas expression array and meta-analysis conducted using the Gene Expression Omnibus database, we determined that PYCR1 was upregulated in HCC compared to adjacent nontumor tissues (P < 0.05). These data were verified using quantitative real-time polymerase chain reaction, western blotting, and immunohistochemistry analysis. Additionally, patients with low PYCR1 expression showed a higher overall survival rate than patients with high PYCR1 expression. Furthermore, PYCR1 overexpression was associated with the female sex, higher levels of alpha-fetoprotein, advanced clinical stages (III and IV), and a younger age (< 45 years old). Silencing of PYCR1 inhibited cell proliferation, invasive migration, epithelial-mesenchymal transition, and metastatic properties in HCC in vitro and in vivo. Using RNA sequencing and bioinformatics tools for data-dependent network analysis, we found binary relationships among PYCR1 and its interacting proteins in defined pathway modules. These findings indicated that PYCR1 played a multifunctional role in coordinating a variety of biological pathways involved in cell communication, cell proliferation and growth, cell migration, a mitogen-activated protein kinase cascade, ion binding, etc. The structural characteristics of key pathway components and PYCR1-interacting proteins were evaluated by molecular docking, and hotspot analysis showed that better affinities between PYCR1 and its interacting molecules were associated with the presence of arginine in the binding site. Finally, a candidate regulatory microRNA, miR-2355-5p, for PYCR1 mRNA was discovered in HCC. Overall, our study suggests that PYCR1 plays a vital role in HCC pathogenesis and may potentially serve as a molecular target for HCC treatment.