Entwicklung von RNA‐basierten Impfstoffen

mRNA‐based vaccines Despite substantial improvements in the development of influenza vaccines, the production and availability of these vaccines remain suboptimal. mRNA‐based vaccines provide an alternative because they can be produced in a fast, flexible and scalable manner. This new vaccine format induces B‐ as well as T‐cell responses and generates memory cells. It confers balanced, long lasting and protective immunity against influenza. Even after thermic stress the efficacy of the vaccines remains fully functional and therefore demonstrates the stability of this new class of drug substances. Since these mRNA based‐vaccines elicit long lived, humoral and cellular immune responses, this new vaccine platform is a technology to produce not only prophylactic vaccines against infection diseases but also to develop effective tumor immunotherapeutics.     

RNA‐Schalter

Riboswitches Synthetic Biology intends to develop novel biological systems such as artificial metabolic pathways. With these, microorganisms are employed to make useful substances such as drugs, biofuels or other chemicals from cheap and renewable resources. Besides genes, regulators are essential parts of genetic circuits. Riboswitches represent a new class of such regulators. They can be developed with customized features using ligand binding RNAs.     

Gene Correction by RNA‐DNA Oligonucleotides

An oligonucleotide composed of a contiguous stretch of RNA and DNA residues has been developed to facilitate the correction of single‐base mutations of episomal and chromosomal targets in mammalian cells. The design of the oligonucleotide exploited the highly recombinogenic RNA‐DNA hybrids and featured hairpin capped ends avoiding destruction by cellular helicases or exonucleases. The RNA‐DNA oligonucleotide (RDO) was designed to correct a point mutation in the tyrosinase gene and caused a permanent gene correction in mouse albino melanocytes, determined by clonal analysis at the level of genomic sequence, protein and phenotypic change. Recently, we demonstrated correction of the tyrosinase gene using the same RDO in vivo, as detected by dark pigmentation of several hairs and DOPA staining of hair follicles in the treated skin of albino mice. Such RDOs might hold a promise as a therapeutic method for the treatment of skin diseases. However, the frequency of gene correction varies among different cells, indicating that cellular activities, such as recombination and repair, may be important for gene conversion by RDOs. As this technology becomes more widely utilized in the scientific community, it will be important to understand the mechanism and to optimize the design of RDOs to improve their efficiency and general applicability.     

Norovirus RNA‐dependent RNA polymerase: A computational study of metal‐binding preferences

Norovirus (NV) RNA‐dependent RNA polymerase (RdRP) is essential for replicating the genome of the virus, which makes this enzyme a key target for the development of antiviral agents against NV gastroenteritis. In this work, a complex of NV RdRP bound to manganese ions and an RNA primer‐template duplex was investigated using X‐ray crystallography and hybrid quantum chemical/molecular mechanical simulations. Experimentally, the complex crystallized in a tetragonal crystal form. The nature of the primer/template duplex binding in the resulting structure indicates that the complex is a closed back‐tracked state of the enzyme, in which the ‐end of the primer occupies the position expected for the post‐incorporated nucleotide before translocation. Computationally, it is found that the complex can accept a range of divalent metal cations without marked distortions in the active site structure. The highest binding energy is for copper, followed closely by manganese and iron, and then by zinc, nickel, and cobalt. Proteins 2017; 85:1435–1445. © 2017 Wiley Periodicals, Inc.     

The DNA‐ and RNA‐binding protein FACTOR of DNA METHYLATION 1 requires XH domain‐mediated complex formation for its function in RNA‐directed DNA methylation

Studies have identified a sub‐group of SGS3‐LIKE proteins including FDM1–5 and IDN2 as key components of RNA‐directed DNA methylation pathway (RdDM). Although FDM1 and IDN2 bind RNAs with 5′ overhangs, their functions in the RdDM pathway remain to be examined. Here we show that FDM1 interacts with itself and with IDN2. Gel filtration suggests that FDM1 may exist as a homodimer in a heterotetramer complex in vivo. The XH domain of FDM1 mediates the FDM1–FDM1 and FDM1–IDN2 interactions. Deletion of the XH domain disrupts FDM1 complex formation and results in loss‐of‐function of FDM1. These results demonstrate that XH domain‐mediated complex formation of FDM1 is required for its function in RdDM. In addition, FDM1 binds unmethylated but not methylated DNAs through its coiled‐coil domain. RNAs with 5′ overhangs does not compete with DNA for binding by FDM1, indicating that FDM1 may bind DNA and RNA simultaneously. These results provide insight into how FDM1 functions in RdDM.