Evolution of secondary structure in the family of 7SL-like RNAs

Primate and rodent genomes are populated with hundreds of thousands copies of Alu and B1 elements dispersed by retroposition, i.e., by genomic reintegration of their reverse transcribed RNAs. These, as well as primate BC200 and rodent 4.5S RNAs, are ancestrally related to the terminal portions of 7SL RNA sequence. The secondary structure of 7SL RNA (an integral component of the signal recognition particle) is conserved from prokaryotes to distant eukaryotic species. Yet only in primates and rodents did this molecule give rise to retroposing Alu and B1 RNAs and to apparently functional BC200 and 4.5S RNAs. To understand this transition and the underlying molecular events, we examined, by comparative analysis, the evolution of RNA structure in this family of molecules derived from 7SL RNA.RNA sequences of different simian (mostly human) and prosimian Alu subfamilies as well as rodent B1 repeats were derived from their genomic consensus sequences taken from the literature and our unpublished results (prosimian and New World Monkey). RNA secondary structures were determined by enzymatic studies (new data on 4.5S RNA are presented) and/or energy minimization analyses followed by phylogenetic comparison. Although, with the exception of 4.5S RNA, all 7SL-derived RNA species maintain the cruciform structure of their progenitor, the details of 7SL RNA folding domains are modified to a different extent in various RNA groups. Novel motifs found in retropositionally active RNAs are conserved among Alu and B1 subfamilies in different genomes. In RNAs that do not proliferate by retroposition these motifs are modified further. This indicates structural adaptation of 7SL-like RNA molecules to novel functions, presumably mediated by specific interactions with proteins; these functions were either useful for the host or served the selfish propagation of RNA templates within the host genome.Abbreviations FAM fossil Alu element - FLAM free left Alu monomer - FRAM free right Alu monomer - L-Alu left Alu subunit - R-Alu right Alu subunit Correspondence to: D. LabudaDedicated to Dr. Robert Cedergren on the occasion of his 25th anniversary at the University of Montreal     

Most human Alu and murine B1 repeats are unique

Alus and B1s are short interspersed repeat elements (SINEs) indirectly derived from the 7SL RNA gene. While most researchers recognize that there exists extensive variability between individual elements, the extent of this variability has never been systematically tested. We examined all Alu elements over 200 nucleotides and all B1 elements over 100 nucleotides in the human and mouse genomes, and analyzed the number of copies of each element at various stringencies from 22 nucleotides to full length. Over 98% of 923,277 Alus and 365,377 B1s examined were unique when queried at full length. When the criterion was reduced to half the length of the repeat, 97% of the Alus and 73% of the B1s were still found to be a single copy. All single and multi-copy sequences have been mapped and documented. Access to the data is possible using the AluPlus website http://www.ibr.hawaii.edu.     

Expression of B7-H1 and B7-DC on the airway epithelium is enhanced by double-stranded RNA

Viral infection in the airway provokes various immune responses, including Th1 and Th2 responses, which are partly initiated by double-stranded RNA (dsRNA), a viral product for its replication. B7-H1 (PD-L1) and B7-DC (PD-L2) are B7-family molecules that bind to programmed death-1 (PD-1) on lymphocytes and are implicated in peripheral tolerance. We investigated the effect of dsRNA on the expression of B7-H1 and B7-DC on airway epithelial cell lines. B7-H1 and B7-DC were constitutively expressed on the cells, and their expression was profoundly upregulated by stimulation with an analog of viral dsRNA, polyinosinic-polycytidylic acid. B7-H1 and B7-DC were also upregulated by stimulation with IFN-gamma, IL-13, and the supernatant from T cell clones. A relatively high concentration of dexamethasone (1 microM) was required to suppress the upregulation of B7-H1 or B7-DC. These results suggest that epithelial B7-H1 and B7-DC play a role in virus-associated immune responses in the airways.     

Selection for fitness versus selection for robustness in RNA secondary structure folding

I investigate the competition between two quasispecies residing on two disparate neutral networks. Under the assumption that the two neutral networks have different topologies and fitness levels, it is the mutation rate that determines which quasispecies will eventually be driven to extinction. For small mutation rates, I find that the quasispecies residing on the neutral network with the lower replication rate will disappear. For higher mutation rates, however, the faster replicating sequences may be outcompeted by the slower replicating ones if the connection density on the second neutral network is sufficiently high. The analytical results are in excellent agreement with flow-reactor simulations of replicating RNA sequences.     

Successive waves of fixation of B1 variants in rodent lineage history

Summary A new method of analyzing phylogenetic relations among members of sequence family (Quentin 1988) discriminates at least six possible B1 subfamilies in the mouse genome. Several additional and independent observations suggest that these grouping have evolutionary significance, and that successive waves of fixation of new variants occur during rodent lineage history. We have reason to believe that, in a genome, the founder sequences of different families of retroposons are in competition with regard to the amplification/fixation process.     

正在崛起的RNA纳米技术

RNA纳米技术得益于纽约大学西曼(Nadrian C.Seeman)教授开创的DNA纳米技术,RNA是由腺嘌呤(A)、尿嘧啶(U)、鸟嘌呤(G)和胞嘧啶(C)构成的一种核糖核酸高分子,与DNA的Watson-Crick碱基配对(A-T,G-C)的双螺旋链的结构不完全一样,RNA的二级结构里经常出现一些非传统的碱基配对如环环相互作用,这些非传统配对促使RNA分子折叠成刚性结构。本文综述了正在崛起的RNA纳米技术,列举了一些著名的实验,如郭培宣(Peixuan Guo)等从自然界的phi29噬菌体中发现的pRNA纳米马达是由六个小RNA分子构成的六环结构,Jaeger等发展了RNA构造术(RNA-tectonics),根据已知的RNA分子的碱基和非传统配对,他们设计利用小RNA分子构造二聚体、一维线性多聚体、和二维网状的七巧板迷宫(jigsaw puzzle)等图案,用tRNA分子或设计用几条RNA分子来构建多面体如立方体和八面体等立体结构等。RNA纳米技术正在崛起,它将在医学、生物技术、合成生物学和纳米技术领域扮演重要的角色。     

Evolution of B2 repeats: the muroid explosion

B2 repeats are a group of short interspersed elements (SINEs) specific for rodent genomes. Copy numbers were determined for different rodent genera. All the Muroid (rat, mouse, deer mouse, hamster, gerbil) rodent genomes analyzed exhibited 80,000–100,000 copies per haploid genome, whereas the squirrel genome contains only 2,500 copies, and fewer than 100 (if any) copies were observed for the Hystricognath rodents (guinea pig and nutria). These findings demonstrate that there was an explosion of amplification of B2 elements within muroid rodents. The similar copy number of B2 elements within the different muroid species could be explained by formation of a high proportion of the B2 elements prior to the divergence of the different muroid species. However, the 3-end of the B2 sequence is unique between murid and cricetid rodents suggesting that the majority of elements amplified after the divergence of these species. Also consistent with recent amplification of these elements in parallel within the muroid genomes is the finding that within mouse and rat there are distinct subfamilies of B2 repeats. The pattern of consistent parallel amplification of B2 elements in muroid species contrasts with the sporadic nature of ID repeat amplification in the same genomes. The consensus of the young mouse subfamily of elements corresponds to the B2 RNA that is preferentially transcribed in embryonic, tumor, and normal liver cells. The subfamily is young based on both its low divergence from the subfamily consensus sequence and the finding that the most recent B2 element insertions in the mouse genome are members of this subfamily.     

ExpaRNA-P: simultaneous exact pattern matching and folding of RNAs

Background

Identifying sequence-structure motifs common to two RNAs can speed up the comparison of structural RNAs substantially. The core algorithm of the existent approach ExpaRNA solves this problem for a priori known input structures. However, such structures are rarely known; moreover, predicting them computationally is no rescue, since single sequence structure prediction is highly unreliable.

Results

The novel algorithm ExpaRNA-P computes exactly matching sequence-structure motifs in entire Boltzmann-distributed structure ensembles of two RNAs; thereby we match and fold RNAs simultaneously, analogous to the well-known “simultaneous alignment and folding” of RNAs. While this implies much higher flexibility compared to ExpaRNA, ExpaRNA-P has the same very low complexity (quadratic in time and space), which is enabled by its novel structure ensemble-based sparsification. Furthermore, we devise a generalized chaining algorithm to compute compatible subsets of ExpaRNA-P’s sequence-structure motifs. Resulting in the very fast RNA alignment approach ExpLoc-P, we utilize the best chain as anchor constraints for the sequence-structure alignment tool LocARNA. ExpLoc-P is benchmarked in several variants and versus state-of-the-art approaches. In particular, we formally introduce and evaluate strict and relaxed variants of the problem; the latter makes the approach sensitive to compensatory mutations. Across a benchmark set of typical non-coding RNAs, ExpLoc-P has similar accuracy to LocARNA but is four times faster (in both variants), while it achieves a speed-up over 30-fold for the longest benchmark sequences (≈400nt). Finally, different ExpLoc-P variants enable tailoring of the method to specific application scenarios. ExpaRNA-P and ExpLoc-P are distributed as part of the LocARNA package. The source code is freely available at http://www.bioinf.uni-freiburg.de/Software/ExpaRNA-P.

Conclusions

ExpaRNA-P’s novel ensemble-based sparsification reduces its complexity to quadratic time and space. Thereby, ExpaRNA-P significantly speeds up sequence-structure alignment while maintaining the alignment quality. Different ExpaRNA-P variants support a wide range of applications.

Electronic supplementary material

The online version of this article (doi:10.1186/s12859-014-0404-0) contains supplementary material, which is available to authorized users.     

The p53 target protein Wig-1 binds hnRNP A2/B1 and RNA Helicase A via RNA

The p53-induced Wig-1 gene encodes a double stranded RNA-binding zinc finger protein. We generated Saos-2 osteosarcoma cells expressing tetracycline-inducible Flag-tagged human Wig-1. Induction of Wig-1 expression by doxycycline inhibited cell growth in a long-term assay but did not cause any changes in cell cycle distribution nor increased fraction of apoptotic cells. Using co-immunoprecipitation and mass spectrometry, we identified two Wig-1-binding proteins, hnRNP A2/B1 and RNA Helicase A, both of which are involved in RNA processing. The binding was dependent on the presence of RNA. Our results establish a link between the p53 tumor suppressor and RNA processing via hnRNPA2/B1 and RNA Helicase A.     

Recruitment of 7SL RNA to assembling HIV‐1 virus‐like particles

Retroviruses incorporate specific host cell RNAs into virions. In particular, the host noncoding 7SL RNA is highly abundant in all examined retroviruses compared with its cellular levels or relative to common mRNAs such as actin. Using live cell imaging techniques, we have determined that the 7SL RNA does not arrive with the HIV‐1 RNA genome. Instead, it is recruited contemporaneously with assembly of the protein HIV‐1 Gag at the plasma membrane. Further, we demonstrate that complexes of 7SL RNA and Gag can be immunoprecipitated from both cytosolic and plasma membrane fractions. This indicates that 7SL RNAs likely interact with Gag prior to high‐order Gag multimerization at the plasma membrane. Thus, the interactions between Gag and the host RNA 7SL occur independent of the interactions between Gag and the host endosomal sorting complex required for transport (ESCRT) proteins, which are recruited temporarily at late stages of assembly. The interactions of 7SL and Gag are also independent of interactions of Gag and the HIV‐1 genome which are seen on the plasma membrane prior to assembly of Gag.      

RNA2D3D: A program for Generating,Viewing, and Comparing 3-Dimensional Models of RNA

Abstract

Using primary and secondary structure information of an RNA molecule, the program RNA2D3D automatically and rapidly produces a first-order approximation of a 3-dimensional conformation consistent with this information. Applicable to structures of arbitrary branching complexity and pseudoknot content, it features efficient interactive graphical editing for the removal of any overlaps introduced by the initial generating procedure and for making conformational changes favorable to targeted features and subsequent refinement. With emphasis on fast exploration of alternative 3D conformations, one may interactively add or delete base-pairs, adjacent stems can be coaxially stacked or unstacked, single strands can be shaped to accommodate special constraints, and arbitrary subsets can be defined and manipulated as rigid bodies. Compaction, whereby base stacking within stems is optimally extended into connecting single strands, is also available as a means of strategically making the structures more compact and revealing folding motifs. Subsequent refinement of the first-order approximation, of modifications, and for the imposing of tertiary constraints is assisted with standard energy refinement techniques. Previously determined coordinates for any part of the molecule are readily incorporated, and any part of the modeled structure can be output as a PDB or XYZ file. Illustrative applications in the areas of ribozymes, viral kissing loops, viral internal ribosome entry sites, and nanobiology are presented.     

Improving the prediction of RNA secondary structure by detecting and assessing conserved stems

The prediction of RNA secondary structure can be facilitated by incorporating with comparative analysis of homologous sequences. However, most of existing comparative methods are vulnerable to alignment errors and thus are of low accuracy in practical application. Here we improve the prediction of RNA secondary structure by detecting and assessing conserved stems shared by all sequences in the alignment. Our method can be summarized by: 1) we detect possible stems in single RNA sequence using the so-called position matrix with which some possibly paired positions can be uncovered; 2) we detect conserved stems across multiple RNA sequences by multiplying the position matrices; 3) we assess the conserved stems using the Signal-to-Noise; 4) we compute the optimized secondary structure by incorporating the so-called reliable conserved stems with predictions by RNAalifold program. We tested our method on data sets of RNA alignments with known secondary structures. The accuracy, measured as sensitivity and specificity, of our method is greater than predictions by RNAalifold.     

Comparative analysis of Stowaway-like miniature inverted repeat transposable elements in wheat group 7 chromosomes: Abundance,composition, and evolution

Miniature inverted repeat transposable elements (MITEs) are the most ubiquitous transposable elements in eukaryotic genomes; they play a prominent role in sequence divergence and genome evolution. There are many well-characterized Stowaway-like MITE families in wheat, but their distribution, abundance, and composition at the chromosome level are still not well understood. In this study, we systematically investigated the Stowaway-like MITEs in wheat group 7 chromosomes based on the survey sequences of isolated wheat chromosomes, to compare them at the chromosome level and to reveal their evolutionary role on wheat polyploidization. In summary, 2026 MITEs were identified, of which 587, 714, and 725 were distributed on 7A, 7B, and 7D chromosomes, respectively. There are more MITEs present on 7D, compared to 7A and 7B, suggesting A and B subgenomes eliminated some repetitive elements during two hybridization processes. Furthermore, some chromosome/arm-specific MITEs were also identified, providing information on the function and evolution of MITEs in wheat genomes. The sequence diversity of the MITE insertions was also investigated. This study for the first time investigated the abundance and composition of MITEs at the chromosome level, which will be beneficial to improve our understanding of the distribution of wheat MITEs and their evolutionary role in polyploidization.