Search

According to the literature, the autoantigen La is involved in Cap-independent translation. It was proposed that one prerequisite for this function is the formation of a protein dimer. However, structural analyses argue against La protein dimers. Noteworthy to mention, these structural analyses were performed under reducing conditions. Here we describe that La protein can undergo redox-dependent structural changes. The oxidized form of La protein can form dimers, oligomers and even polymers stabilized by disulfide bridges. The primary sequence of La protein contains three cysteine residues. Only after mutation of all three cysteine residues to alanine La protein becomes insensitive to oxidation, indicating that all three cysteines are involved in redox-dependent structural changes. Biophysical analyses of the secondary structure of La protein support the redox-dependent conformational changes. Moreover, we identified monoclonal anti-La antibodies (anti-La mAbs) that react with either the reduced or oxidized form of La protein. Differential reactivities to the reduced and oxidized form of La protein were also found in anti-La sera of autoimmune patients.

Already decades ago, we and many other groups showed a nucleo-cytoplasmic translocation of La protein in cultured cells. This shuttling of La protein was seen after e.g. UV irradiation, virus infections, hydrogen peroxide exposure, and Fenton reaction based on iron or copper ions. In common, all these conditions are somehow related to oxidative stress. Unfortunately, all these harsh conditions could also cause an artificial release of La protein. Even until today, the shut-tling and a cytoplasmic function of La/SS-B are controversially discussed. Moreover, the driving mechanism for the shuttling of La protein remains unclear. Recently we showed that La protein undergoes redox dependent conformational changes. Moreover, we developed anti-La mono-clonal antibodies (anti-La mAbs) which are specific for either the reduced form of La protein or the oxidized form. Using these tools, here we show that redox dependent conformational changes are the driving force for the shuttling of La protein. Moreover, we show that transloca-tion of La protein to the cytoplasm can be triggered in a ligand/receptor dependent manner un-der physiological conditions. We show that ligands of toll-like receptors lead to a redox de-pendent shuttling of La protein. The shuttling of La protein depends on the redox status of the respective cell type. Endothelial cells are usually resistant to the shuttling of La protein while dendritic cells are highly sensitive. However, deprivation of intracellular reducing agents in endothelial cells turns endothelial cells sensitive to a redox dependent shuttling of La protein.

Since the first description of the nuclear autoantigens in the late sixties and early seventies of the last century we and many other groups have tried with difficulty to establish monoclonal anti-bodies (mabs) against nuclear antigens including to the autoantigen La/SS-B. To date, only a few anti-La mabs have been derived by conventional hybridoma technology. However, these pre-viously described anti-La mabs are not bona fide autoantibodies as they recognize either human La specific- or cryptic or post-translationally modified epitopes which are not accessible on na-tive mouse La protein. Herein we present a series of novel murine anti-La mabs including truly autoreactive ones. These mabs were elicited from a human La transgenic animal through adop-tive transfer of T cells from non-transgenic mice immunized with human La antigen. Detailed epitope and paratope analyses experimentally confirm the hypothesis that somatic hypermuta-tions occurring during T cell dependent maturation can lead to autoreactivity to the nuclear au-toantigen La/SS-B.

The article deals with the creation and functioning of a mediation service. The purpose, functions, basic principles of the mediation service, as well as the necessity and importance of creating such a service in a preschool educational organization as a way to protect the rights and freedoms of minors. The author not only describes his point of view, but also provides a regulatory framework for the creation and functioning of a mediation service., В статье рассматривается вопрос создания и функционирования службы медиации. Цель, функции, основные принципы службы медиации, а так же необходимость и значимость создания такой службы в дошкольной образовательной организации как способа защиты прав и свобод несовершеннолетних. Автор не только описывает свою точку зрение, но и дает нормативно-правовую опору создания и функционирования службы медиации.

Forensic DNA phenotyping is gaining interest as the number of applications increases within the forensic genetics community. The possibility of providing investigative leads in addition to conventional DNA profiling for human identification provides new insights into otherwise “cold” police investigations. The ability of reporting on the bio-geographical ancestry (BGA), appearance characteristics and age based on DNA obtained from a crime scene sample of an unknown donor makes the exploration of such markers and the development of new methods meaningful for criminal investigations. The VISible Attributes through GEnomics (VISAGE) Consortium aims to disseminate and broaden the use of predictive markers and develop fully optimized and validated prototypes for forensic casework implementation. Here, the first VISAGE appearance and ancestry tool development, performance and validation is reported. A total of 153 SNPs (96.84 % assay conversion rate) were successfully incorporated into a single multiplex reaction using the AmpliSeq™ design pipeline, and applied for massively parallel sequencing with the Ion S5 platform. A collaborative effort involving six VISAGE laboratory partners was devised to perform all validation tests. An extensive validation plan was carefully organized to explore the assay's overall performance with optimum and low-input samples, as well as with challenging and casework mock samples. In addition, forensic validation studies such as concordance and mixture tests recurring to the Coriell sample set with known genotypes were performed. Finally, inhibitor tolerance and specificity were also evaluated. Results showed a robust, highly sensitive assay with good overall concordance between laboratories.