A Database of microRNA Expression Patterns in Xenopus laevis

MicroRNAs (miRNAs) are short, non-coding RNAs around 22 nucleotides long. They inhibit gene expression either by translational repression or by causing the degradation of the mRNAs they bind to. Many are highly conserved amongst diverse organisms and have restricted spatio-temporal expression patterns during embryonic development where they are thought to be involved in generating accuracy of developmental timing and in supporting cell fate decisions and tissue identity. We determined the expression patterns of 180 miRNAs in Xenopus laevis embryos using LNA oligonucleotides. In addition we carried out small RNA-seq on different stages of early Xenopus development, identified 44 miRNAs belonging to 29 new families and characterized the expression of 5 of these. Our analyses identified miRNA expression in many organs of the developing embryo. In particular a large number were expressed in neural tissue and in the somites. Surprisingly none of the miRNAs we have looked at show expression in the heart. Our results have been made freely available as a resource in both XenMARK and Xenbase.     

Expression Profile of the Schistosoma japonicum Degradome Reveals Differential Protease Expression Patterns and Potential Anti-schistosomal Intervention Targets

Blood fluke proteases play pivotal roles in the processes of invasion, nutrition acquisition, immune evasion, and other host-parasite interactions. Hundreds of genes encoding putative proteases have been identified in the recently published schistosome genomes. However, the expression profiles of these proteases in Schistosoma species have not yet been systematically analyzed. We retrieved and culled the redundant protease sequences of Schistosoma japonicum, Schistosoma mansoni, Echinococcus multilocularis, and Clonorchis sinensis from public databases utilizing bioinformatic approaches. The degradomes of the four parasitic organisms and Homo sapiens were then comparatively analyzed. A total of 262 S. japonicum protease sequences were obtained and the expression profiles generated using whole-genome microarray. Four main clusters of protease genes with different expression patterns were identified: proteases up-regulated in hepatic schistosomula and adult worms, egg-specific or predominantly expressed proteases, cercaria-specific or predominantly expressed proteases, and constantly expressed proteases. A subset of protease genes with different expression patterns were further validated using real-time quantitative PCR. The present study represents the most comprehensive analysis of a degradome in Schistosoma species to date. These results provide a firm foundation for future research on the specific function(s) of individual proteases and may help to refine anti-proteolytic strategies in blood flukes.     

Next Generation Sequencing Analysis Reveals Segmental Patterns of microRNA Expression in Mouse Epididymal Epithelial Cells

The functional maturation of mammalian spermatozoa is accomplished as the cells descend through the highly specialized microenvironment of the epididymis. This dynamic environment is, in turn, created by the combined secretory and absorptive activity of the surrounding epithelium and displays an extraordinary level of regionalization. Although the regulatory network responsible for spatial coordination of epididymal function remains unclear, recent evidence has highlighted a novel role for the RNA interference pathway. Indeed, as noncanonical regulators of gene expression, small noncoding RNAs have emerged as key elements of the circuitry involved in regulating epididymal function and hence sperm maturation. Herein we have employed next generation sequencing technology to profile the genome-wide miRNA signatures of mouse epididymal cells and characterize segmental patterns of expression. An impressive profile of some 370 miRNAs were detected in the mouse epididymis, with a subset of these specifically identified within the epithelial cells that line the tubule (218). A majority of the latter miRNAs (75%) were detected at equivalent levels along the entire length of the mouse epididymis. We did however identify a small cohort of miRNAs that displayed highly regionalized patterns of expression, including miR-204-5p and miR-196b-5p, which were down- and up-regulated by approximately 39- and 45-fold between the caput/caudal regions, respectively. In addition we identified 79 miRNAs (representing ~ 21% of all miRNAs) as displaying conserved expression within all regions of the mouse, rat and human epididymal tissue. These included 8/14 members of let-7 family of miRNAs that have been widely implicated in the control of androgen signaling and the repression of cell proliferation and oncogenic pathways. Overall these data provide novel insights into the sophistication of the miRNA network that regulates the function of the male reproductive tract.     

Widely Conserved Recombination Patterns among Single-Stranded DNA Viruses

The combinatorial nature of genetic recombination can potentially provide organisms with immediate access to many more positions in sequence space than can be reached by mutation alone. Recombination features particularly prominently in the evolution of a diverse range of viruses. Despite rapid progress having been made in the characterization of discrete recombination events for many species, little is currently known about either gross patterns of recombination across related virus families or the underlying processes that determine genome-wide recombination breakpoint distributions observable in nature. It has been hypothesized that the networks of coevolved molecular interactions that define the epistatic architectures of virus genomes might be damaged by recombination and therefore that selection strongly influences observable recombination patterns. For recombinants to thrive in nature, it is probably important that the portions of their genomes that they have inherited from different parents work well together. Here we describe a comparative analysis of recombination breakpoint distributions within the genomes of diverse single-stranded DNA (ssDNA) virus families. We show that whereas nonrandom breakpoint distributions in ssDNA virus genomes are partially attributable to mechanistic aspects of the recombination process, there is also a significant tendency for recombination breakpoints to fall either outside or on the peripheries of genes. In particular, we found significantly fewer recombination breakpoints within structural protein genes than within other gene types. Collectively, these results imply that natural selection acting against viruses expressing recombinant proteins is a major determinant of nonrandom recombination breakpoint distributions observable in most ssDNA virus families.Genetic recombination is a ubiquitous biological process that is both central to DNA repair pathways (10, 57) and an important evolutionary mechanism. By generating novel combinations of preexisting nucleotide polymorphisms, recombination can potentially accelerate evolution by increasing the population-wide genetic diversity upon which adaptive selection relies. Recombination can paradoxically also prevent the progressive accumulation of harmful mutations within individual genomes (18, 35, 53). Whereas its ability to defend high-fitness genomes from mutational decay possibly underlies the evolutionary value of sexuality in higher organisms, in many microbial species where pseudosexual genetic exchange is permissible among even highly divergent genomes, recombination can enable access to evolutionary innovations that would otherwise be inaccessible by mutation alone.Such interspecies recombination is fairly common in many virus families (8, 17, 27, 44, 82). It is becoming clear, however, that as with mutation events, most recombination events between distantly related genomes are maladaptive (5, 13, 38, 50, 63, 80). As genetic distances between parental genomes increase, so too does the probability of fitness defects in their recombinant offspring (16, 51). The viability of recombinants is apparently largely dependent on how severely recombination disrupts coevolved intragenome interaction networks (16, 32, 51). These networks include interacting nucleotide sequences that form secondary structures, sequence-specific protein-DNA interactions, interprotein interactions, and amino acid-amino acid interactions within protein three-dimensional folds.One virus family where such interaction networks appear to have a large impact on patterns of natural interspecies recombination are the single-stranded DNA (ssDNA) geminiviruses. As with other ssDNA viruses, recombination is very common among the species of this family (62, 84). Partially conserved recombination hot and cold spots have been detected in different genera (39, 81) and are apparently caused by both differential mechanistic predispositions of genome regions to recombination and natural selection disfavoring the survival of recombinants with disrupted intragenome interaction networks (38, 51).Genome organization and rolling circle replication (RCR)—the mechanism by which geminiviruses and many other ssDNA viruses replicate (9, 67, 79; see reference 24 for a review)—seem to have a large influence on basal recombination rates in different parts of geminivirus genomes (20, 33, 39, 61, 81). To initiate RCR, virion-strand ssDNA molecules are converted by host-mediated pathways into double-stranded “replicative-form” (RF) DNAs (34, 67). Initiated by a virus-encoded replication-associated protein (Rep) at a well-defined virion-strand replication origin (v-ori), new virion strands are synthesized on the complementary strand of RF DNAs (28, 73, 74) by host DNA polymerases. Virion-strand replication is concomitant with the displacement of old virion strands, which, once complete, yields covalently closed ssDNA molecules which are either encapsidated or converted into additional RF DNAs. Genome-wide basal recombination rates in ssDNA viruses are probably strongly influenced by the specific characteristics of host DNA polymerases that enable RCR. Interruption of RCR has been implicated directly in geminivirus recombination (40) and is most likely responsible for increased basal recombination rates both within genes transcribed in the opposite direction from that of virion-strand replication (40, 71) and at the v-ori (1, 9, 20, 69, 74).Whereas most ssDNA virus families replicate via either a rolling circle mechanism (the Nanoviridae, Microviridae, and Geminiviridae) (3, 23, 24, 31, 59, 67, 74) or a related rolling hairpin mechanism (the Parvoviridae) (25, 76), among the Circoviridae only the Circovirus genus is known to use RCR (45). Although the Gyrovirus genus (the other member of the Circoviridae) and the anelloviruses (a currently unclassified ssDNA virus group) might also use RCR, it is currently unknown whether they do or not (78). Additionally, some members of the Begomovirus genus of the Geminiviridae either have a second genome component, called DNA-B, or are associated with satellite ssDNA molecules called DNA-1 and DNA-Beta, all of which also replicate by RCR (1, 47, 68).Recombination is known to occur in the parvoviruses (19, 43, 70), microviruses (66), anelloviruses (40, 46), circoviruses (11, 26, 60), nanoviruses (30), geminivirus DNA-B components, and geminivirus satellite molecules (2, 62). Given that most, if not all, of these ssDNA replicons are evolutionarily related to and share many biological features with the geminiviruses (22, 31, 36), it is of interest to determine whether conserved recombination patterns observed in the geminiviruses (61, 81) are evident in these other groups. To date, no comparative analyses have ever been performed with different ssDNA virus families to identify, for example, possible influences of genome organization on recombination breakpoint distributions found in these viruses.Here we compare recombination frequencies and recombination breakpoint distributions in most currently described ssDNA viruses and satellite molecules and identify a number of sequence exchange patterns that are broadly conserved across this entire group.     

Conserved B-Cell Epitopes among Human Bocavirus Species Indicate Potential Diagnostic Targets

Background

Human bocavirus species 1–4 (HBoV1–4) have been associated with respiratory and enteric infections in children. However, the immunological mechanisms in response to HBoV infections are not fully understood. Though previous studies have shown cross-reactivities between HBoV species, the epitopes responsible for this phenomenon remain unknown. In this study, we used genomic and immunologic approaches to identify the reactive epitopes conserved across multiple HBoV species and explored their potential as the basis of a novel diagnostic test for HBoVs.

Methodology/Principal Findings

We generated HBoV1–3 VP2 gene fragment phage display libraries (GFPDLs) and used these libraries to analyze mouse antisera against VP2 protein of HBoV1, 2, and 3, and human sera positive for HBoVs. Using this approach, we mapped four epitope clusters of HBoVs and identified two immunodominant peptides–P1 (¹MSDTDIQDQQPDTVDAPQNT²⁰), and P2 (¹⁶²EHAYPNASHPWDEDVMPDL¹⁸⁰)–that are conserved among HBoV1–4. To confirm epitope immunogenicity, we immunized mice with the immunodominant P1 and P2 peptides identified in our screen and found that they elicited high titer antibodies in mice. These two antibodies could only recognize the VP2 of HBoV 1–4 in Western blot assays, rather than those of the two other parvoviruses human parvovirus B19 and human parvovirus 4 (PARV4). Based on our findings, we evaluated epitope-based peptide-IgM ELISAs as potential diagnostic tools for HBoVs IgM antibodies. We found that the P1+P2-IgM ELISA showed a higher sensitivity and specificity in HBoVs IgM detection than the assays using a single peptide.

Conclusions/Significance

The identification of the conserved B-cell epitopes among human bocavirus species contributes to our understanding of immunological cross-reactivities of HBoVs, and provides important insights for the development of HBoV diagnostic tools.     

Conserved microRNA function as a basis for Chinese hamster ovary cell engineering

The use of microRNAs (miRNAs) for improving the efficiency of recombinant protein production by CHO cells is gaining considerable interest for their ability to regulate entire molecular networks. Differential miRNA expression profiling and large-scale transient screening have been the prerequisite for the selection of miRNA candidates for stable manipulation, reported in CHO cells expressing a range of recombinant products. We selected a potent and well characterised tumour suppressor miRNA, miR-34a, as a high priority candidate for CHO cell engineering based on the conservation of both its sequence and function across species and cell type. Ectopic expression of miR-34a retained its functional conservation in CHO-SEAP cells by inhibiting growth by 90 % in addition to decreasing the viable cell population by 30 % when compared to controls. When the miR-34 family was stably depleted using a miRNA sponge decoy vector, the overall product yield was enhanced by ~2-fold in both fed-batch and small scale clonal batch cultures, despite having a negative impact on cell growth. These findings further strengthen the utility of miRNAs as engineering tools to modify and improve CHO cell performance.     

Resting-State Functional Connectivity Patterns Predict Chinese Word Reading Competency

Resting-state functional connectivity (RSFC) offers a novel approach to reveal the temporal synchronization of functionally related brain regions. Recent studies have identified several RSFCs whose strength was associated with reading competence in alphabetic languages. In the present study, we examined the role of intrinsic functional relations for reading a non-alphabetic language – Chinese – by correlating RSFC maps of nine Chinese reading-related seed regions and reaction time in the single-character reading task. We found that Chinese reading efficiency was positively correlated with the connection between left inferior occipital gyrus and left superior parietal lobule, between right posterior fusiform gyrus and right superior parietal lobule, and between left inferior temporal gyrus and left inferior parietal lobule. These results could not be attributed to inter-individual differences arising from the peripheral processes of the reading task such as visual input detection and articulation. The observed RSFC-reading correlation relationships are discussed in the framework of Chinese character reading, including visuospatial analyses and semantic/phonological processes.     

Gene Expression in Chicken Reveals Correlation with Structural Genomic Features and Conserved Patterns of Transcription in the Terrestrial Vertebrates

Background

The chicken is an important agricultural and avian-model species. A survey of gene expression in a range of different tissues will provide a benchmark for understanding expression levels under normal physiological conditions in birds. With expression data for birds being very scant, this benchmark is of particular interest for comparative expression analysis among various terrestrial vertebrates.

Methodology/Principal Findings

We carried out a gene expression survey in eight major chicken tissues using whole genome microarrays. A global picture of gene expression is presented for the eight tissues, and tissue specific as well as common gene expression were identified. A Gene Ontology (GO) term enrichment analysis showed that tissue-specific genes are enriched with GO terms reflecting the physiological functions of the specific tissue, and housekeeping genes are enriched with GO terms related to essential biological functions. Comparisons of structural genomic features between tissue-specific genes and housekeeping genes show that housekeeping genes are more compact. Specifically, coding sequence and particularly introns are shorter than genes that display more variation in expression between tissues, and in addition intergenic space was also shorter. Meanwhile, housekeeping genes are more likely to co-localize with other abundantly or highly expressed genes on the same chromosomal regions. Furthermore, comparisons of gene expression in a panel of five common tissues between birds, mammals and amphibians showed that the expression patterns across tissues are highly similar for orthologuous genes compared to random gene pairs within each pair-wise comparison, indicating a high degree of functional conservation in gene expression among terrestrial vertebrates.

Conclusions

The housekeeping genes identified in this study have shorter gene length, shorter coding sequence length, shorter introns, and shorter intergenic regions, there seems to be selection pressure on economy in genes with a wide tissue distribution, i.e. these genes are more compact. A comparative analysis showed that the expression patterns of orthologous genes are conserved in the terrestrial vertebrates during evolution.     

Multiscale Simulations Reveal Conserved Patterns of Lipid Interactions with Aquaporins

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Highlights? Multiscale MD simulations predict the interactions of lipids with membrane proteins ? The method is evaluated via comparison with the structure of Aqp0 in a membrane ? Simulations of aquaporins reveal a broadly conserved protein/lipid interface ? The results suggest interchange between annular and bulk lipids     

Body Mass Patterns Predict Invasions and Extinctions in Transforming Landscapes

Scale-specific patterns of resource distribution on landscapes entrain attributes of resident animal communities such that species body-mass distributions are organized into distinct aggregations. Species within each aggregation respond to resources over the same range of scale. This discontinuous pattern has predictive power: invasive species and extinct or declining species in landscapes subject to human transformation tend to be located at the edge of body-mass aggregations (P < 0.01), which may be transition zones between distinct ranges of scale. Location at scale breaks affords species great opportunity, but also potential crisis. Received 19 May 1998; accepted 24 November 1998