Construction of a series of several self-cleaving RNA duplexes using synthetic 21-mers

AbstractTwo fragments (21-mers) containing consensus sequences for the self-cleavage domain in transcripts of satellite DNA of the newt were chemically synthesized and found to be cleaved in the presence of Mg2+. The cleaved product contained the 3′-terminal 2′,3′-cyclic phosphate. Twenty-five combinations of partially double-stranded 21-mer RNA which contained different bases within the consensus sequences and at the cleavage sites were tested for self-cleavage. It seemed that guanosine 3′-phosphate was not susceptible to transesterification at the cleavage site

Recognition of 2′-hydroxyl groups by Escherichia coli ribonuclease HI

AbstractIn order to investigate the hydrogen-bonding interactions between Escherichia coli ribonuclease HI and the 2′-hydroxyl functions of the substrate, oligonucleotide duplexes containing 2′-amino-2′-deoxyuridine or 2′-fluoro-2′-deoxyuridine at a specific site were used, and their affinities for the enzyme were determined by kinetic analyses. The results indicate that the hydroxyl groups of the nucleoside 3′-adjacent to the cleaved phosphodiester linkage and the second nucleoside 5′ to the cleaved phosphodiester act as both a proton donor and an acceptor and as a proton acceptor, respectively, in the enzyme-substrate complex. A molecular model was constructed using the interactions derived from the results

Sequence-specific cleavage of RNA by a hybrid ribonuclease H

AbstractSite-specific cleavage of the 22-, 132- and 534-base RNAs by the DNA/protein hybrid R Nase H were examined. The 22-base RNA was chemically synthesized, and 132- and 534-base RNAs were prepared by run-off transcription. The hybrid enzyme cleaves these RNAs, which contain a single target sequence, primarily at the unique phosphodiester bond within the target sequence. The hybrid enzyme performs multiple turnovers, and at a substrate/enzyme ratio of 10:1 the RNAs are almost completely cleaved by the hybrid enzyme at 37°C within 1 h. We propose that hybrid RNase H molecules with various oligodeoxyribonucleotides function as RNA restriction enzymes and are useful for structural and functional studies of RNA

Induced 2C Expression and Implantation-Competent Blastocyst-like Cysts from Primed Pluripotent Stem Cells

Soon after fertilization, the few totipotent cells of mammalian embryos diverge to form a structure called the blastocyst (BC). Although numerous cell types, including germ cells and extended-pluripotency stem cells, have been developed from pluripotent stem cells (PSCs) in vitro, generating functional BCs only from PSCs remains elusive. Here, we describe induced self-organizing 3D BC-like cysts (iBLCs) generated from mouse PSC culture. Resembling natural BCs, iBLCs have a blastocoel-like cavity and were formed with outer cells expressing trophectoderm lineage markers and with inner cells expressing pluripotency markers. iBLCs transplanted to pseudopregnant mice uteruses implanted, induced decidualization, and exhibited growth and development before resorption, demonstrating that iBLCs are implantation competent. iBLC precursor intermediates required the transcription factor Prdm14 and concomitantly activated the totipotency-related cleavage-stage MERVL reporter and 2C genes. Thus, our system may contribute to the understanding of molecular mechanisms underpinning totipotency, embryogenesis, and implantation

千葉県君津市川谷地域に露出する中部更新統柿ノ木台層から産出する冷湧水化石群集: その時空分布と共産する自生炭酸塩

金沢大学国際基幹教育院 GS教育系冷湧水性群集が房総半島の中部更新統柿ノ木台層の陸棚相から産出する．群集は，化学合成二枚貝類から排他的になり，著しく13Cに枯渇した自生炭酸塩と共産することから，AOM（嫌気的メタン酸化）に依存していたと考えられる．自生炭酸塩は巣穴壁面と巣穴周囲の堆積物中に沈殿し，巣穴からスナモグリ類の爪化石と糞化石が産出することから，これらはスナモグリ類の巣穴であると考えられる．スナモグリ類はメタン生成帯まで巣穴を堀り，海水を巣穴深部へ供給し，AOMを活性化させることによって巣穴中の硫化水素イオン濃度を上昇させた．溶存酸素濃度が高い巣穴浅部では，硫黄酸化菌が繁茂し，スナモグリ類の食糧となった．巣穴深部では，浮遊する生物源炭酸塩などを核とした針状アラゴナイトが重力方向に沈下して炭酸塩ジオペタル状構造を形成し，巣穴周囲の堆積物中では，リン酸イオン濃度の上昇により高Mgカルサイトが，また硫酸イオンの枯渇によりドロマイトが沈殿した．Cold-seep-dependent molluscan assemblages occur in the outer-shelf facies of the middle Pleistocene Kakinokidai Formation of the Kazusa Group, a forearc basin-fill sequence on the Pacific side of central Japan, in strata corresponding to the interval 707.6-667.0 ka. The assemblages consist exclusively of chemosymbiotic bivalves (lucinids, thyasirids, and solemyids) and are associated with 13C-depleted authigenic carbonates (δ13C = −61.60‰ to −10.96‰ VPDB), which suggest that their main carbon source was anaerobic oxidation of methane (AOM). Authigenic carbonate precipitates are common on burrow walls (mainly acicular aragonite) and the surrounding sediments (mainly micritic high-Mg calcite and dolomite). The burrows are cylindrical, 1.5-3.0 cm in diameter, and >1 m long. Callianassid claws and the trace fossil Palaxius (probable callianassid fecal pellets) in the burrow carbonates suggest that the burrows were produced by sediment-dwelling callianassid decapods.\nWe propose the following formation mechanism of burrows and their related authigenic carbonates. Firstly, callianassids produced deep burrows, penetrating the AOM zone and reaching the methanogenic zone. Methane then seeped into the burrows and AOM occurred in its deeper parts, promoted by a supply of seawater via callianassid activity, resulting in an increase in the concentration of hydrogen sulfide ions. Thiobacteria flourished in the shallower parts of the burrows, which were enriched in dissolved oxygen, and provided a source of food for the callianassids. In the deeper parts of the burrows, acicular aragonite precipitated around suspended carbonate nuclei and sank to the bottoms of the burrows, forming geopetal-like carbonate structures. In the surrounding sediment, high-Mg calcite precipitated in response to an increase in the concentration of phosphate ions (due to the decomposition of organic matter), and dolomite precipitated in response to decreasing concentrations of sulfate ions (caused by active AOM)