Methylation of the nucleobases in RNA oligonucleotides mediates duplex-hairpin conversion.

We have systematically investigated the duplex to hairpin conversion of oligoribonucleotides under the aspect of nucleobase methylation. The first part of our study refers to the self-complementary sequence rCGCGAAUUCGCGA, which forms a stable Watson-Crick base paired duplex under various buffer conditions. It is shown that this sequence is forced to adopt a hairpin conformation if one of the central 6 nt is replaced by the corresponding methylated nucleotide, such as 1-methylguanosine N(2),N(2)-dimethylguanosine, N(6),N(6)-dimethyladenosine (m(6)(2)A) or 3-methyluridine. On the other hand, the duplex structure is retained and even stabilized by replacement of a central nucleotide with N(2)-methylguanosine (m(2)G) or N(4)-methylcytidine. A borderline case is represented by N(6)-methyladenosine (m(6)A). Although generally a duplex-preserving modification, our data indicate that m(6)A in specific strand positions and at low strand concentrations is able to effectuate duplex-hairpin conversion. Our studies also include the ssu ribosomal helix 45 sequence motif, rGACCm(2)GGm(6)(2)Am(6)(2)AGGUC. In analogy, it is demonstrated that the tandem m(6)(2)A nucleobases of this oligoribonucleotide prevent duplex formation with complementary strands. Therefore, it can be concluded that nucleobase methylations at the Watson-Crick base pairing site provide the potential not only to modulate but to substantially affect RNA structure by formation of different secondary structure motifs.

Bridged cyclic oligoribonucleotides--towards models for codon-anticodon pairing.

Only three base pairs make up for stable double helices of regular A-type if both helix ends are bridged by flexible non-nucleotide linkers. These cyclic oligoribonucleotides are used as model systems for codon-anticodon pairing in order to reveal base stacking effects arising from structurally relevant bases in the direct neighbourhood of the core triplet duplex.

Flexible non-nucleotide linkers as loop replacements in short double helical RNAs.

Ethylene glycol oligomers have been studied systematically as non-nucleotide loop replacements in short hairpin oligoribonucleotides. Structural optimization concerns the length of the linkers and is based on the thermodynamic stabilities of the corresponding duplexes. The optimum linker is derived from heptakis (ethylene glycol) provided that the duplex end to be bridged comprises solely the terminal base pair; the optimum linker is derived from hexakis(ethylene glycol) if a dangling unpaired nucleotide is incorporated into the loop. Moreover, these linkers have been compared to other commonly used linker types which consist of repeating units of tris- or tetrakis(ethylene glycol) phosphate, or of 3-hydroxypropane-1-phosphate. In all cases, the correlation between linker length and duplex stability is independent of the kind of counter ions used (Na(+), Na(+)/Mg(2+), K(+)or Li(+)). Furthermore, all duplexes with non-nucleotide loop replacements are less stable than those with the corresponding standard nucleotide loop. The results corroborate that the linkers are solvent-exposed and do not specifically interfere with the terminal nucleotides at the bridged duplex end.

Synergistic effect of triiodothyronine and dexamethasone on renal tubular transport of p-aminohippurate in rats of different ages.

Postnatal maturation of the renal tubular transport of p-aminohippurate (PAH) was verified in rats both in diuresis experiand on renal cortical slices in vitro. Following treatment with various doses of triiodothyronine (T3) or dexamethasone (3 days, once a day), the increase in renal tubular transport of PAH was more distinct in young rats with immature kidney function. The synergistic effects of simultaneous treatment with both hormones indicate differences between T3 and dexamethasone in the modulation of cellular function.