Prediction and statistics of pseudoknots in RNA structures using exactly clustered stochastic simulations

Ab initio RNA secondary structure predictions have long dismissed helices interior to loops, so-called pseudoknots, despite their structural importance. Here, we report that many pseudoknots can be predicted through long time scales RNA folding simulations, which follow the stochastic closing and opening of individual RNA helices. The numerical efficacy of these stochastic simulations relies on an O(n^2) clustering algorithm which computes time averages over a continously updated set of n reference structures. Applying this exact stochastic clustering approach, we typically obtain a 5- to 100-fold simulation speed-up for RNA sequences up to 400 bases, while the effective acceleration can be as high as 100,000-fold for short multistable molecules (<150 bases). We performed extensive folding statistics on random and natural RNA sequences, and found that pseudoknots are unevenly distributed amongst RNAstructures and account for up to 30% of base pairs in G+C rich RNA sequences (Online RNA folding kinetics server including pseudoknots : http://kinefold.u-strasbg.fr/ ).Comment: 6 pages, 5 figure

The fetal profile line:a proposal for a sonographic reference line to classify forehead and mandible anomalies in the second and third trimester

Objectives To test the fetal profile (FP) line, defined as the line that passes through the anterior border of the mandible and the nasion, as a reference line for forehead and mandible anomalies. Methods Volumes of 248 normal and 24 pathological fetuses (1636 and 1937?weeks gestation, respectively) were analysed retrospectively. When the FP line passes anteriorly, across or posteriorly to the frontal bone, this was defined as negative, zero or positive, respectively. When the FP line was positive the distance (F distance) between the FP line and the frontal bone was measured. Results No cases with a negative FP line were found in the normal fetuses. Before 27?weeks gestation the FP line was always zero except in one case. After 27?weeks gestation the FP line was positive in up to 25% (F distance (mean, range): 2.8, 2.13.6?mm). The FP line correctly identified 13 cases with retrognathia, 5 cases with frontal bossing and 3 cases with a sloping forehead. Conclusion Although large prospective studies are needed, the FP line may be a useful tool to detect second trimester profile anomalies such as retrognathia, sloping forehead and frontal bossing with the possibility of quantifying the latter. (c) 2012 John Wiley & Sons, Ltd

Predicting RNA pseudoknot folding thermodynamics

Based on the experimentally determined atomic coordinates for RNA helices and the self-avoiding walks of the P (phosphate) and C(4) (carbon) atoms in the diamond lattice for the polynucleotide loop conformations, we derive a set of conformational entropy parameters for RNA pseudoknots. Based on the entropy parameters, we develop a folding thermodynamics model that enables us to compute the sequence-specific RNA pseudoknot folding free energy landscape and thermodynamics. The model is validated through extensive experimental tests both for the native structures and for the folding thermodynamics. The model predicts strong sequence-dependent helix-loop competitions in the pseudoknot stability and the resultant conformational switches between different hairpin and pseudoknot structures. For instance, for the pseudoknot domain of human telomerase RNA, a native-like and a misfolded hairpin intermediates are found to coexist on the (equilibrium) folding pathways, and the interplay between the stabilities of these intermediates causes the conformational switch that may underlie a human telomerase disease

Stem-loop structures can effectively substitute for an RNA pseudoknot in-1 ribosomal frameshifting

Biophysical Structural ChemistrySupramolecular & Biomaterials Chemistr

Structural parameters affecting the kinetics of RNA hairpin formation

There is little experimental knowledge on the sequence dependent rate of hairpin formation in RNA. We have therefore designed RNA sequences that can fold into either of two mutually exclusive hairpins and have determined the ratio of folding of the two conformations, using structure probing. This folding ratio reflects their respective folding rates. Changing one of the two loop sequences from a purine- to a pyrimidine-rich loop did increase its folding rate, which corresponds well with similar observations in DNA hairpins. However, neither changing one of the loops from a regular non-GNRA tetra-loop into a stable GNRA tetra-loop, nor increasing the loop size from 4 to 6 nt did affect the folding rate. The folding kinetics of these RNAs have also been simulated with the program ‘Kinfold’. These simulations were in agreement with the experimental results if the additional stabilization energies for stable tetra-loops were not taken into account. Despite the high stability of the stable tetra-loops, they apparently do not affect folding kinetics of these RNA hairpins. These results show that it is possible to experimentally determine relative folding rates of hairpins and to use these data to improve the computer-assisted simulation of the folding kinetics of stem–loop structures

Five pseudoknots are present at the 204 nucleotides long 3' noncoding region of tobacco mosak virus RNA

Biophysical Structural Chemistr

Structural motifs in the RNA encoded by the early nodulation gene enod40 of soybean

Biophysical Structural ChemistryAnimal science

The 3′-untranslated region of the α2C-adrenergic receptor mRNA impedes translation of the receptor message

We report that tyro subtypes of alpha(2)-adrenergic receptors (alpha(2A/D)- and alpha(2C)-AR) are ectopically expressed with dramatically different efficiencies and that this difference is due to a 288-nucleotide (nt) segment in the 3'-untranslated region (3'-UTR) of the alpha(2C)-AR mRNA that impairs translational processing, NIH-3T3 fibroblasts mere transfected with receptor constructs (coding region plus 552 nt, alpha(2C)-AR; coding region plus 1140 nt, alpha(2A/D)-AR) and a vector conferring G418 resistance. Transcription was driven by the murine sarcoma virus promoter element, and the receptor gene segment was upstream of an SV40 polyadenylation cassette. Drug-resistant transfectants were evaluated for expression of receptor mRNA and protein, 90% of the NIH-3T3 alpha(2C)-AR transfectants expressed receptor mRNA, but only 14% of the clonal cell Lines expressed receptor protein. In contrast, 90% of the NIH-3T3 alpha(2A/D)-AR transfectants expressed receptor protein (200-5000 fmol/mg). Similar results were obtained following transfection of DDT1MF-2 cells with the two receptor constructs. The role of the 3'-UTR of the alpha(2C)-AR in mRNA processing was determined by generating new constructs in which the 3'-UTR. was progressively truncated from 552 to 470, 182, 143, or 74 nt 3' to the stop codon. Truncation of the 3'-UTR resulted in the expression of receptor protein in the G418-resistant transfectants (nt 74, 100%; nt 143, 80%; nt 182, 50%). The level of mRNA in the transfectants expressing the receptor protein was not greater than that in nonexpressing clones, and the differences in protein expression did not reflect altered mRNA stability in the truncated construct. The alpha(2C)-AR mRNA with the longer 3'-UTR underwent translational initiation as it was found in the polysome fraction, indicating that the lack of receptor protein was due to impaired translational elongation or termination. These data suggest that translational efficiency is a hey mechanism for regulating alpha(2C)-AR expression and associated signaling events.Biophysical Structural Chemistr

Elongation factor Tu3 from the kirromycin-producer Streptomyces ramocissimus is resistant to three classes of EF-Tu-specific inhibitors

Microbial BiotechnologyBiophysical Structural Chemistr