Evolution of conserved secondary structures and their function in transcriptional regulation networks

Background: Many conserved secondary structures have been identified within conserved elements in the human genome, but only a small fraction of them are known to be functional RNAs. The evolutionary variations of these conserved secondary structures in h

Secondary structures of mRNAs and splicing of introns in eukaryotes.

After analyzing the secondary structures of 68 exon-intron-exon and the corresponding exon-exon sequence segments, it is found that about 90% of 5' and 3' terminal bases G (splicing sites) of introns are situated in the loops of secondary structures or at the ends of stems near the loops, and most of "G" s in loops are closed to the ends of loops. Approximately 92% of the connecting sites of the adjoining exons also show the similar features. About 82% of the branch point "A" s are situated in loops or at the ends of stems near the loops. Splicing sites and branch points approach each other in space because of the folding.

Cardioviral RNA structure logo analysis: entropy, correlations, and prediction

In recent years, there has been an increased number of sequenced RNAs leading to the development of new RNA databases. Thus, predicting RNA structure from multiple alignments is an important issue to understand its function. Since RNA secondary structures are often conserved in evolution, developing methods to identify covariate sites in an alignment can be essential for discovering structural elements. Structure Logo is a technique established on the basis of entropy and mutual information measured to analyze RNA sequences from an alignment. We proposed an efficient Structure Logo approach to analyze conservations and correlations in a set of Cardioviral RNA sequences. The entropy and mutual information content were measured to examine the conservations and correlations, respectively. The conserved secondary structure motifs were predicted on the basis of the conservation and correlation analyses. Our predictive motifs were similar to the ones observed in the viral RNA structure database, and the correlations between bases also corresponded to the secondary structure in the database.

Mutations stabilize small subunit ribosomal RNA in desiccation-tolerant cyanobacteria Nostoc

The ribosomal RNA molecule is an ideal model for evaluating the stability of a gene product under desiccation stress. We isolated 8 Nostoc strains that had the capacity to withstand desiccation in habitats and sequenced their 16S rRNA genes. The stabilities of 16S rRNAs secondary structures, indicated by free energy change of folding, were compared among Nostoc and other related species. The results suggested that 163 rRNA secondary structures of the desiccation-tolerant Nostoc strains were more stable than that of planktonic Nostocaceae species. The stabilizing mutations were divided into two categories: (1) those causing GC to replace other types of base pairs in stems and (2) those causing extension of stems. By mapping stabilizing mutations onto the Nostoc phylogenetic tree based on 16S rRNA gene, it was shown that most of stabilizing mutations had evolved during adaptive radiation among Nostoc spp. The evolution of 16S rRNA along the Nostoc lineage is suggested to be selectively advantageous under desiccation stress.

mRNA 很可能携带三维遗传信息

The forming mechanism of the three - dimensional structures of proteins，i.e.the mechanism of protein folding，is a basic problem in molecular biology which is still unsolved unitl now. In which a core problem is whether there is the three – dimensional genetic information that decide the three - dimensional structures of proteins. However, the research on this field has mot yet been reported. Recently，we made a comparative study on the folded structures of more than 70 mature messeneger RNAs (mRNAs) and the three - dimensional structures of the proteins encoded by them，it has been found that there exist marked correspondences between their featured structures in the following aspects: 1.The number of the structural units. An RNA molecule can form a secondary structure(stem and loop structure) by the folding and the base pairing of itself. The elementary structural unit of an RNA secondary structure is hairpin(or compound hair pin).The regular structural unit in the secondary structure of a protein is # alpha # - helix or #beta# - sheet . We have found that the hairpin number in the secondary structure of each mature mRNA is equal or approximately equal to the number of the regular secondary structural unis of the encoded protein. 2 .Turning region. Turn is a main structrual element in the secondary structure of a protein, which decides the backbone orientation of a protein molecule to some extent .Our analysis shows that the nucleotide sequence segments in an mRNA which encode the turns of the corresponding protein are overall situated in the turning regions of the mRNA secondary structure such as haipin，bulge loop or multibaranch loops. 3 .The arrangement of structural elements in space. In order to understand the backbone orientation of an RNA molecule and the arangement of its structural elements in space，we have modeled the three一dimensional structure of the mRNA molecule on SGI workstation based on its secondary structure.The result shows that the spatial arrangement of most of the nucleotide sequence segments encoding the structural elements of a protein is consistent with that of these stretural exements in the protein. For instance，the nucleotide sequences corresponding to each pleated sheet of a # beta # - sheet structure are close to each other in the mRNA secondary stucture and in the three - dimensional structure，although some of the nucleotide segments are far apart from each other in the one - dimensional sequence. For another instance，the two triplet codons of cysteines which form a disulphide bridge geneal1y are very close to each other in the mRNA folded structure. In addition，we also analyzed the locations of the codons proline - coding and the distrbution of the nucleotide sequences #alpha# - helix - coding in the folded structures of mRNAs . Some distribution laws have been found. All of these results suggest that the transfer of the genetic information from mRNA to protein not only is one – dimensional but also is three - dime ns ional. That is，there exists the genetic information that decide the three - dimensional structures of proteins. To a certain extent，we could say that the mRNA folding detemines the protein folding. Based on these results，it would be possible to predict the three - dimensional structures of proteins from the primary，secondary and tertiary structures of the m RNAs at a higher accuracy.And more important is that a new clue has been provided to uncover the“spatial coding" of the genetic information.

Phylogenetic analysis of the family Thermaceae with an emphasis on signature position and secondary structure of 16S rRNA

The sequences of the 16S rRNA genes from 38 strains of the family Thermaceae were compared by alignment analysis. The genus-specific and species-specific base substitutions or base deletions (signature positions) were found in three hypervariable regions (in the helices 6, 10 and 17). The differentiation of secondary structures of the high variable regions in the 5' end (38-497) containing several signature positions further supported the concept. Based on the comparisons of the secondary structures in the segments of 16S rRNAs, a key to the species of the family Thermaceae was proposed. (C) 2003 Published by Elsevier Science B.V. on behalf of the Federation of European Microbiological Societies.

Exploring consensus mRNA secondary (folding) structure units by stochastic sampling and folding simulation

It is extremely difficult to explore mRNA folding structure by biological experiments. In this report, we use stochastic sampling and folding simulation to test the existence of the stable secondary structural units of-mRNA, look for the folding units, and explore the probabilistic stabilization of the units. Using this method, We made simulations for all possible local optimum secondary structures of a single strand mRNA within a certain range, and searched for the common parts of the secondary structures. The consensus secondary structure units (CSSUs) extracted from the above method are mainly hairpins, with a few single strands. These CSSUs suggest that the mRNA folding units could be relatively stable and could perform specific biological function. The significance of these observations for the mRNA folding problem in general is also discussed. (c) 2004 Elsevier B.V. All rights reserved.

Bead-probe complex capture a couple of SINE and LINE family from genomes of two closely related species of East Asian cyprinid directly using magnetic separation

Background: Short and long interspersed elements (SINEs and LINEs, respectively), two types of retroposons, are active in shaping the architecture of genomes and powerful tools for studies of phylogeny and population biology. Here we developed special protocol to apply biotin-streptavidin bead system into isolation of interspersed repeated sequences rapidly and efficiently, in which SINEs and LINEs were captured directly from digested genomic DNA by hybridization to bead-probe complex in solution instead of traditional strategy including genomic library construction and screening. Results: A new couple of SINEs and LINEs that shared an almost identical 3'tail was isolated and characterized in silver carp and bighead carp of two closely related species. These SINEs (34 members), designated HAmo SINE family, were little divergent in sequence and flanked by obvious TSD indicated that HAmo SINE was very young family. The copy numbers of this family was estimated to 2 x 10(5) and 1.7 x 10(5) per haploid genome by Real-Time qPCR, respectively. The LINEs, identified as the homologs of LINE2 in other fishes, had a conserved primary sequence and secondary structures of the 3'tail region that was almost identical to that of HAmo SINE. These evidences suggest that HAmo SINEs are active and amplified recently utilizing the enzymatic machinery for retroposition of HAmoL2 through the recognition of higher-order structures of the conserved 42-tail region. We analyzed the possible structures of HAmo SINE that lead to successful amplification in genome and then deduced that HAmo SINE, SmaI SINE and FokI SINE that were similar in sequence each other, were probably generated independently and created by LINE family within the same lineage of a LINE phylogeny in the genomes of different hosts. Conclusion: The presented results show the advantage of the novel method for retroposons isolation and a pair of young SINE family and its partner LINE family in two carp fishes, which strengthened the hypotheses containing the slippage model for initiation of reverse transcription, retropositional parasitism of SINEs on LINEs, the formation of the stem loop structure in 3'tail region of some SINEs and LINEs and the mechanism of template switching in generating new SINE family.

人类基因组中保守二级结构的纯净化选择及其在转录调控网络中的作用

保守序列是一种跨物种保守的基因组序列，而且绝大多数为非蛋白编码序 列。保守序列在人类遗传疾病中发挥着重要作用。其中，一部分保守序列能够 折叠形成二级结构。已鉴定的一些保守二级结构编码一些RNA 分子，如 microRNA、RNA 编辑序列和组蛋白mRNA 3’端非翻译区茎环结构等。但是，对 于绝大部分的保守二级结构，它们的生物学功能以及作用于它们上面的进化作 用力依然是未知的。 群体的SNP 数据在分析序列上的进化作用力时非常有效。SNP 在群体中的 频率会因为受到不同的进化作用力而表现出差异，而与其是否位于基因组中的 突变热点无关。对于受纯净化选择作用的SNP，它们的频率一般会比中性SNP 具有低的新生型等位基因频率（DAF）。我们运用生物信息学的方法，在人类基 因组保守二级结构中找到746 个SNP。这746 个SNP 与基因组其它区段的SNP 在突变模式上并不存在显著差异，在保守二级结构内同样存在突变热点。通过 与侧翼序列SNP 的分布比较发现，保守二级结构上SNP 密度约为其侧翼序列的 2/3。相比于侧翼序列SNP，有更高比例的保守二级结构SNP 具有低的DAF 值。 这些结果提示，有很多保守二级结构上的SNP 因为受到纯净化选择作用而在现 代人群中被剔除了。保守二级结构与侧翼序列在SNP 密度和DAF 上的差异要高 于保守序列与非保守序列之间的差异，提示保守二级结构是受到纯净化选择作 用最为严格的一类保守序列。我们发现，在保守二级结构内部，纯净化选择作 用的强度也有差异。茎区比环区具有更低的SNP 密度，而且有更高比例的茎区 SNP 具有低的DAF 值。这个结果提示，保守二级结构上的纯净化选择力主要作 用于茎区上的位点。我们推测，这可能是茎区上的突变往往比环区的突变对二级结构的造成更大的影响导致的。 我们通过寻找保守二级结构与转录因子SOX2、OCT4、NANOG、SUZ12 和C-MYC 结合位点之间的重叠，还分析了保守二级结构在转录调控网络中的作用。结果 显示，很多保守二级结构是作为转录因子的结合位点调控了许多与发育相关的 转录因子编码基因的表达。转录因子与保守二级结构之间的结合模式非常复杂， 可以有多个转录因子结合到同一个保守二级结构上，也可以是一个转录因子结 合到自身编码基因相关的保守二级结构上。不同的转录因子和保守二级结构结 合可以主导靶基因的特异模式，当绝大多数相关的保守二级结构与SUZ12 结合 时，基因表达受到抑制，而当绝大多数相关的保守二级结构不与SUZ12 结合时， 基因表达受到激活。在转录调控网络中，约有30%的保守二级结构是作为启动 子来调控基因的表达。因为转录因子SOX2、OCT4、NANOG、SUZ12 和C-MYC 仅仅 只结合到很小一部分保守二级结构上，提示可能还有更多的转录因子会结合到 保守二级结构上。因此，保守二级结构介导的转录调控网络要比目前已知的复 杂得多。

Adaptive recognition of small molecules by nucleic acid aptamers through a label-free approach

We report a novel label-free method for the investigation of the adaptive recognition of small molecules by nucleic acid aptamers using capillary electrophoresis analysis. Cocaine and argininamide were chosen as model molecules, and the two corresponding DNA aptamers were used. These single-strand DNAs folded into their specific secondary structures, which were mainly responsible for the binding of the target molecules with high affinity and specificity. For molecular recognition, the nucleic acid structures then underwent additional conformational changes, while keeping the target molecules stabilized by intermolecular hydrogen bonds. The intrinsic chemical and physical properties of the target molecules enabled them to act as indicators for adaptive binding. Thus any labeling or modification of the aptamers or target molecules were made obsolete. This label-free method for aptamer-based molecular recognition was also successfully applied to biological fluids and therefore indicates that this approach is a promising tool for bioanalysis.

Maximins S, a novel group of antimicrobial peptides from toad Bombina maxima

Amphibian skin secretions are rich in antimicrobial peptides acting as important components of innate defense system against invading microorganisms. A novel type of peptide, designated as maximin S, was deduced by random sequencing of 793 clones from a constructed Bombina maxima skin cDNA library. The putative primary structures of maximin S peptides can be grouped into five species, in which maximin S I has 14 amino acid residues and the rest of maximin S peptides (S2-S5) all have 18 amino acid residues. Unlike most of the amphibian antimicrobial peptides so far identified, the newly characterized four maximin S precursors are composed of maximin S I and different combinations of tandem repeated maximin S2-S5 linked by internal peptides. Except maximin S I, the predicted secondary structures of maximin S2-S5 show a similar amphipathic alpha-helical structure. MALDI-TOF mass spectrometry analysis of partially isolated skin secretions of the toad indicates that most of the deduced maximin S peptides are expressed. Two deduced maximin S peptides (S1, S4) were synthesized and their antimicrobial activities were tested. Maximin S4 only had an antibiotic activity against mycoplasma and had no antibacterial or antifungal activity toward tested strains. Maximin S1 had no activity under the same conditions. (C) 2004 Elsevier Inc. All rights reserved.

蛋白质的二级结构和mRNA 的二级结构的相关性研究

利用置信度为95 的特征字研究了蛋白质二级结构以及其对应mRNA 二级结构, 发现 蛋白质二级结构和mRNA 二级结构有明显的相关性. 规则二级结构A2螺旋, B2折叠以及包含 有Turn 的边界明显倾向于mRNA 二级结构的茎区, 而避免出现在环区.

Is there a genetic relationship between mRNA and the encoded protein at three-dimensional level?

A comparative study on the structures of some mRNAs and their encoded proteins shows an intriguing correlation between the two foldings. Non-random distribution of codons in the secondary structures of mRNAs is also shown, which appears to be in accordance with the conformational properties of amino acids in protein structures to some extent. These results seem to suggest that there may be a kind of genetic relationship between mRNA and protein at three-dimensional level.

Structural electrochemical study of hemoglobin by in situ circular dichroism thin layer spectroelectrochemistry

Secondary and tertiary or quaternary structural changes in hemoglobin (HB) during an electroreduction process were studied by in situ circular dichroism (CD) spectroelectrochemistry with a long optical path thin-layer cell. By means of singular value decomposition least-squares analysis, CD spectra in the far-UV region give two similar a components with different CD intensity, indicating slight denaturation in the secondary structures due to the electric field effect. CD spectra in the Soret band show a R --> T transition of two quaternary structural components induced by electroreduction of the heme, which changes the redox states of the center ion from Fe3+ to Fe2+ and the coordination number from 6 to 5. The double logarithmic analysis shows that electroreduction of hemoglobin follows a chemical reaction with R --> T transition. Some parameters in the electrochemical process were obtained: formal potential, E-0t = -0.167 V; electrochemical kinetic overpotential, DeltaE(0) = -0.32 V; standard electrochemical reaction rate constant, k(0) = 1.79 x 10(-5) cm s(-1); product of electron transfer coefficient and electron number, alphan=0.14; and the equilibrium constant of R --> T transition, K-c = 9.0.