Extending the chemistry that supports genetic information transfer in vivo: phosphorothioate DNA, phosphorothioate RNA, 2'-O-methyl RNA, and methylphosphonate DNA.

DNA and RNA are the polynucleotides known to carry genetic information in life. Chemical variants of DNA and RNA backbones have been used in structure-function and biosynthesis studies in vitro, and in antisense pharmacology, where their properties of nuclease resistance and enhanced cellular uptake are important. This study addressed the question of whether the base(s) attached to artificial backbones encodes genetic information that can be transferred in vivo. Oligonucleotides containing chemical variants of DNA or RNA were used as primers for site-specific mutagenesis of bacteriophage f1. Progeny phage were scored both genetically and physically for the inheritance of information originally encoded by bases attached to the nonstandard backbones. Four artificial backbone chemistries were tested: phosphorothioate DNA, phosphorothioate RNA, 2'-O-methyl RNA and methylphosphonate DNA. All four were found capable of faithful information transfer from their attached bases when one or three artificial positions were flanked by normal DNA. Among oligonucleotides composed entirely of nonstandard backbones, only phosphorothioate DNA supported genetic information transfer in vivo.

A biomarker that identifies senescent human cells in culture and in aging skin in vivo.

Normal somatic cells invariably enter a state of irreversibly arrested growth and altered function after a finite number of divisions. This process, termed replicative senescence, is thought to be a tumor-suppressive mechanism and an underlying cause of aging. There is ample evidence that escape from senescence, or immortality, is important for malignant transformation. By contrast, the role of replicative senescence in organismic aging is controversial. Studies on cells cultured from donors of different ages, genetic backgrounds, or species suggest that senescence occurs in vivo and that organismic lifespan and cell replicative lifespan are under common genetic control. However, senescent cells cannot be distinguished from quiescent or terminally differentiated cells in tissues. Thus, evidence that senescent cells exist and accumulate with age in vivo is lacking. We show that several human cells express a beta-galactosidase, histochemically detectable at pH 6, upon senescence in culture. This marker was expressed by senescent, but not presenescent, fibroblasts and keratinocytes but was absent from quiescent fibroblasts and terminally differentiated keratinocytes. It was also absent from immortal cells but was induced by genetic manipulations that reversed immortality. In skin samples from human donors of different age, there was an age-dependent increase in this marker in dermal fibroblasts and epidermal keratinocytes. This marker provides in situ evidence that senescent cells may exist and accumulate with age in vivo.

A human RNase E-like activity that cleaves RNA sequences involved in mRNA stability control.

We have detected an endoribonucleolytic activity in human cell extracts that processes the Escherichia coli 9S RNA and outer membrane protein A (ompA) mRNA with the same specificity as RNase E from E. coli. The human enzyme was partially purified by ion-exchange chromatography, and the active fractions contained a protein that was detected with antibodies shown to recognize E. coli RNase E. RNA containing four repeats of the destabilizing motif AUUUA and RNA from the 3' untranslated region of human c-myc mRNA were also found to be cleaved by E. coli RNase E and its human counterpart in a fashion that may suggest a role of this activity in mammalian mRNA decay. It was also found that RNA containing more than one AUUUA motif was cleaved more efficiently than RNA with only one or a mutated motif. This finding of a eukaryotic endoribonucleolytic activity corresponding to RNase E indicates an evolutionary conservation of the components of mRNA degradation systems.

WW6: an embryonic stem cell line with an inert genetic marker that can be traced in chimeras.

Mutant mice produced by gene targeting in embryonic stem (ES) cells often have a complex or embryonic lethal phenotype. In these cases, it would be helpful to identify tissues and cell types first affected in mutant embryos by following the contribution to chimeras of ES cells homozygous for the mutant allele. Although a number of strategies for following ES cell development in vivo have been reported, each has limitations that preclude its general application. In this paper, we describe ES cell lines that can be tracked to every nucleated cell type in chimeras at all developmental stages. These lines were derived from blastocysts of mice that carry an 11-Mb beta-globin transgene on chromosome 3. The transgene is readily detected by DNA in situ hybridization, providing an inert, nuclear-localized marker whose presence is not affected by transcriptional or translational controls. The "WW" series of ES lines possess the essential features of previously described ES lines, including giving rise to a preponderance of male chimeras, all of which have to date exhibited germ-line transmission. In addition, clones selected for single or double targeting events form strong chimeras, demonstrating the feasibility of using WW6 cells to identify phenotypes associated with the creation of a null mutant.

Estado actual de la investigación sobre la teoría tridimensional de la ansiedad de Lang

Este estudio ofrece una revisión de la teoría tridimensional propuesta por Lang (1968), quien mantiene que la ansiedad se manifiesta según un triple sistema de respuesta (cognitivo, fisiológico y motor o conductual) que pueden ser discordantes. Estos tres sistemas, regulados por normas diferentes, dan lugar a perfiles de respuesta diferenciales, los cuales será necesario evaluar por separado para, posteriormente, elegir el método de tratamiento más adecuado. En esta línea, el presente estudio ofrece una revisión de investigaciones que, hasta el día de hoy, han apoyado la teoría propuesta por Lang. Finalmente, este estudio revisa diferentes cuestionarios, inventarios y escalas construidos desde la teoría propuesta por Lang, revelando que, en la actualidad, la teoría tridimensional de la ansiedad continúa siendo aplicada en el ámbito clínico, de la salud y el escolar, lo que denota su actualidad, relevancia y practicidad.

Before the parliamentary elections in Ukraine. OSW Commentary No. 149, 15.10.2014

The parliamentary elections to be held in Ukraine on 26 October will bring about deep changes in the political composition of the Verkhovna Rada, Ukraine’s parliament. It is very probable that after the elections only one or two of the five parties which are represented in the parliament today will remain, and the leading positions will be taken by groupings who were still considered marginal a year ago. The Petro Poroshenko Bloc, a party which according to polls can count on victory, did not exist a year ago and today still remains in the construction phase. It is likely that around two-thirds of the newly elected deputies will be people with no parliamentary or even no political experience. On the one hand, this may be a strong impetus to revival; but on the other the lack of experience of most of the parliamentarians may be a problem. Another source of potential problems may be the process of consolidating the internally unstable political parties which have emerged during the electoral campaign. All of the parties which will count in the upcoming elections have a pro-European programme. It is probable that the numerous supporters of an anti-Western orientation (although not necessarily a pro-Russian orientation, as recent events have shown) will be represented by very few deputies elected in single-mandate constituencies. On the one hand, this homogeneity within the Rada will facilitate the country’s reforms, including work on the new constitution, while on the other it might be a subject of permanent criticism by Moscow and its Ukrainian representatives. The elections will take place in the conditions of a growing wave of social disappointment and ongoing military actions in the eastern part of the country. Nevertheless, it can be expected that the vote will be held without any major disturbances, and its course will be transparent and fair.

Improvement of the American Indian Religious Freedom Act [microform] : hearing before the Select Committee on Indian Affairs, United States Senate, One Hundredth Congress, second session, on S. 2250 to ensure that federal lands are managed in a manner that does not impair the exercise of traditional American Indian religion, May 18, 1988, Washington, DC.

CIS Microfiche Accession Numbers: CIS 89 S411-4