The coming of age for Piwi proteins

Piwi proteins, a subfamily of Argonaute (Ago) proteins, have recently been shown to bind endogenous small RNAs. However, differences between Ago proteins (which bind microRNAs and small interfering RNAs) and Piwi proteins and Piwi-interacting RNAs (piRNAs) suggest novel functions for Piwi proteins. Here, we highlight the recent progress in understanding Piwi function and the implications for germline and stem cell development.     

Computational immunology: The coming of age

The explosive growth in biotechnology combined with major advances in information technology has the potential to radically transform immunology in the postgenomics era. Not only do we now have ready access to vast quantities of existing data, but new data with relevance to immunology are being accumulated at an exponential rate. Resources for computational immunology include biological databases and methods for data extraction, comparison, analysis and interpretation. Publicly accessible biological databases of relevance to immunologists number in the hundreds and are growing daily. The ability to efficiently extract and analyse information from these databases is vital for efficient immunology research. Most importantly, a new generation of computational immunology tools enables modelling of peptide transport by the transporter associated with antigen processing (TAP), modelling of antibody binding sites, identification of allergenic motifs and modelling of T-cell receptor serial triggering.     

Enrichment of genomic DNA for polymorphism detection in a non-model highly polyploid crop plant

Large polyploid genomes of non-model species remain challenging targets for DNA polymorphism discovery despite the increasing throughput and continued reductions in cost of sequencing with new technologies. For these species especially, there remains a requirement to enrich genomic DNA to discover polymorphisms in regions of interest because of large genome size and to provide the sequence depth to enable estimation of copy number. Various methods of enriching DNA have been utilised, but some recent methods enable the efficient sampling of large regions (e.g. the exome). We have utilised one of these methods, solution-based hybridization (Agilent SureSelect), to capture regions of the genome of two sugarcane genotypes (one Saccharum officinarum and one Saccharum hybrid) based mainly on gene sequences from the close relative Sorghum bicolor. The capture probes span approximately 5.8?megabases (Mb). The enrichment over whole-genome shotgun sequencing was 10-11-fold for the two genotypes tested. This level of enrichment has important consequences for detecting single nucleotide polymorphisms (SNPs) from a single lane of Illumina (Genome Analyzer) sequence reads. The detection of polymorphisms was enabled by the depth of sequence at or near probe sites and enabled the detection of 270?000-280?000 SNPs within each genotype from a single lane of sequence using stringent detection parameters. The SNPs were present in 13?000-16?000 targeted genes, which would enable mapping of a large number of these chosen genes. SNP validation from 454 sequencing and between-genotype confirmations gave an 87%-91% validation rate.     

Linkage mapping and comparative genomics using next-generation RAD sequencing of a non-model organism

Restriction-site associated DNA (RAD) sequencing is a powerful new method for targeted sequencing across the genomes of many individuals. This approach has broad potential for genetic analysis of non-model organisms including genotype-phenotype association mapping, phylogeography, population genetics and scaffolding genome assemblies through linkage mapping. We constructed a RAD library using genomic DNA from a Plutella xylostella (diamondback moth) backcross that segregated for resistance to the insecticide spinosad. Sequencing of 24 individuals was performed on a single Illumina GAIIx lane (51 base paired-end reads). Taking advantage of the lack of crossing over in homologous chromosomes in female Lepidoptera, 3,177 maternally inherited RAD alleles were assigned to the 31 chromosomes, enabling identification of the spinosad resistance and W/Z sex chromosomes. Paired-end reads for each RAD allele were assembled into contigs and compared to the genome of Bombyx mori (n = 28) using BLAST, revealing 28 homologous matches plus 3 expected fusion/breakage events which account for the difference in chromosome number. A genome-wide linkage map (1292 cM) was inferred with 2,878 segregating RAD alleles inherited from the backcross father, producing chromosome and location specific sequenced RAD markers. Here we have used RAD sequencing to construct a genetic linkage map de novo for an organism that has no previous genome data. Comparative analysis of P. xyloxtella linkage groups with B. mori chromosomes shows for the first time, genetic synteny appears common beyond the Macrolepidoptera. RAD sequencing is a powerful system capable of rapidly generating chromosome specific data for non-model organisms.     

Biomolecularmodeling and simulation: a field coming of age

We assess the progress in biomolecular modeling and simulation, focusing on structure prediction and dynamics, by presenting the field’s history, metrics for its rise in popularity, early expressed expectations, and current significant applications. The increases in computational power combined with improvements in algorithms and force fields have led to considerable success, especially in protein folding, specificity of ligand/biomolecule interactions, and interpretation of complex experimental phenomena (e.g. NMR relaxation, protein-folding kinetics and multiple conformational states) through the generation of structural hypotheses and pathway mechanisms. Although far from a general automated tool, structure prediction is notable for proteins and RNA that preceded the experiment, especially by knowledge-based approaches. Thus, despite early unrealistic expectations and the realization that computer technology alone will not quickly bridge the gap between experimental and theoretical time frames, ongoing improvements to enhance the accuracy and scope of modeling and simulation are propelling the field onto a productive trajectory to become full partner with experiment and a field on its own right.     

The coming of age of tumour immunotherapy

Compared with the earlier incidence of acute infectious diseases, the introduction of vaccines has been one of the major public health success achievements. In contrast, vaccine development to control some persisting infections such as HIV remains a major challenge. There are many similarities with this task and that of controlling tumours by immunotherapy. Generating CTL responses by using pulsed dendritic cells has become a popular approach and has led to success with the mouse model. With viral antigens, priming with DNA plasmids and boosting with a chimeric live vector results in high levels of CTL activity, and is worth trying with cancer. A recent review highlights three other difficulties posed by tumours: epitope stability, maiming or killing of CTL by the tumour, and accessibility of the tumour vasculature to immune components. The new ability to label CTL by staining with specific tetrameric peptide/MHC complexes offers the possibility of effectively studying this third aspect. Our increased knowledge of tumour-associated antigens, viral or otherwise, and our growing ability to manipulate the immune system, offers hope that control of at least some human tumours may be within reach.     

Single molecule biology: coming of age

Cellular heterogeneity and stochastic fluctuation play key roles in biological processes. Single molecule approaches have the key advantage of avoiding ensemble averaging, allowing the observation of transient intermediates and heterogeneity (both static and dynamic). Thus they have revolutionised the way many biological questions are addressed. The challenge ahead is to develop integrated approaches such as the combination of single molecule imaging with single molecule manipulation to probe the dynamics of gene regulatory and cell signalling networks in living cells.     

The coming of age of virus-like particle vaccines

Virus-like particles are supra-molecular assemblages, usually icosahedral or rod-like structures. They incorporate key immunologic features of viruses which include repetitive surfaces, particulate structures and induction of innate immunity through activation of pathogen-associated molecular-pattern recognition receptors. They carry no replicative genetic information and can be produced recombinantly in large scale. Virus-like particles thus represent a safe and effective vaccine platform for inducing potent B- and T-cell responses. In addition to being effective vaccines against the corresponding virus from which they are derived, virus-like particles can also be used to present foreign epitopes to the immune system. This can be achieved by genetic fusion or chemical conjugation. This technological innovation has greatly broadened the scope of their use, from immunizing against microbial pathogens to immunotherapy for chronic diseases. Towards this end, virus-like particles have been used to induce autoantibodies to disease-associated self-molecules involved in chronic diseases, such as hypertension and Alzheimer's disease. The recognition of the potent immunogenicity and commercial potential for virus-like particles has greatly accelerated research and development activities. During the last decade, two prophylactic virus-like particle vaccines have been registered for human use, while another 12 vaccines entered clinical development.