Healing of erosions in rheumatoid arthritis remains elusive: results with 24 months of the anabolic agent teriparatide.

Objective: Erosion healing in rheumatoid arthritis (RA) is difficult to demonstrate. This extension study aimed to determine whether 2 years of teriparatide (TPTD) produces erosion healing. Method: Subjects in a previous 12 month randomized controlled trial of TPTD in RA were invited to receive 12 additional months of open-label TPTD. Eleven of the 24 original subjects were enrolled in the extension study, six of whom received TPTD in the final 12 months only. Subjects receiving 24 months of TPTD were assessed for reduction in erosion volume from baseline using computed tomography. We also compared erosion volumes between 12 and 24 months of TPTD. Large erosions in subjects receiving TPTD for 24 months were examined for volume change. Results: In the six patients who received 24 months of TPTD, there was no significant change in erosion volume at the metacarpophalangeal (MCP) and proximal interphalangeal (PIP) joints compared with baseline. The six subjects who received 24 months of TPTD had similar changes in erosion volume to the five who received 12 months of TPTD, in MCP (p = 0.17) and PIP (p = 0.63) joints. Assessment of large erosions in those receiving TPTD for 24 months showed no evidence of erosion healing. Conclusion: While this extension study was too small to be conclusive, we observed no evidence of reduction in erosion volume with the addition of TPTD for 24 months in subjects with RA in whom disease activity was controlled on a tumour necrosis factor inhibitor. This is consistent with our negative findings at 12 months.

Intervention strategies for microbial therapeutics in cancer immunotherapy.

Immunotherapies have drastically improved clinical outcomes in a wide range of malignancies. Nevertheless, patient responses remain highly variable, and reliable biomarkers that predict responses accurately are not yet fully understood. Compelling evidence from preclinical studies and observational data from clinical cohorts have shown that commensal microorganisms that reside in the human gastrointestinal tract, collectively termed the 'microbiome', can actively modify responses to chemotherapeutic agents and immunotherapies by influencing host immunosurveillance. Notably, microbial correlates are largely context specific, and response signatures may vary by patient population, geographic location and type of anticancer treatment. Therefore, the incongruence of beneficial microbiome signatures across studies, along with an emerging understanding of the mechanisms underlying the interactions between the microbiome, metabolome and host immune system, highlight a critical need for additional comprehensive and standardized multi-omics studies. Future research should consider key host factors, such as diet and use of medication, in both preclinical animal models and large-scale, multicenter clinical trials. In addition, there is a strong rationale to evaluate the microbiome as a tumor-extrinsic biomarker of clinical outcomes and to test the therapeutic potential of derived microbial products (e.g. defined microbial consortia), with the eventual goal of improving the efficacy of existing anticancer treatments. This review discusses the importance of the microbiome from the perspective of cancer immunotherapies, and outlines future steps that may contribute to wide-ranging clinical and translational benefits that may improve the health and quality of life of patients with cancer.

Mobile genes in the human microbiome are structured from global to individual scales.

Recent work has underscored the importance of the microbiome in human health, and has largely attributed differences in phenotype to differences in the species present among individuals. However, mobile genes can confer profoundly different phenotypes on different strains of the same species. Little is known about the function and distribution of mobile genes in the human microbiome, and in particular whether the gene pool is globally homogenous or constrained by human population structure. Here, we investigate this question by comparing the mobile genes found in the microbiomes of 81 metropolitan North Americans with those of 172 agrarian Fiji islanders using a combination of single-cell genomics and metagenomics. We find large differences in mobile gene content between the Fijian and North American microbiomes, with functional variation that mirrors known dietary differences such as the excess of plant-based starch degradation genes found in Fijian individuals. Notably, we also observed differences between the mobile gene pools of neighbouring Fijian villages, even though microbiome composition across villages is similar. Finally, we observe high rates of recombination leading to individual-specific mobile elements, suggesting that the abundance of some genes may reflect environmental selection rather than dispersal limitation. Together, these data support the hypothesis that human activities and behaviours provide selective pressures that shape mobile gene pools, and that acquisition of mobile genes is important for colonizing specific human populations.

What can comparative genomics tell us about species concepts in the genus Aspergillus?

Understanding the nature of species" boundaries is a fundamental question in evolutionary biology. The availability of genomes from several species of the genus Aspergillus allows us for the first time to examine the demarcation of fungal species at the whole-genome level. Here, we examine four case studies, two of which involve intraspecific comparisons, whereas the other two deal with interspecific genomic comparisons between closely related species. These four comparisons reveal significant variation in the nature of species boundaries across Aspergillus. For example, comparisons between A. fumigatus and Neosartorya fischeri (the teleomorph of A. fischerianus) and between A. oryzae and A. flavus suggest that measures of sequence similarity and species-specific genes are significantly higher for the A. fumigatus - N. fischeri pair. Importantly, the values obtained from the comparison between A. oryzae and A. flavus are remarkably similar to those obtained from an intra-specific comparison of A. fumigatus strains, giving support to the proposal that A. oryzae represents a distinct ecotype of A. flavus and not a distinct species. We argue that genomic data can aid Aspergillus taxonomy by serving as a source of novel and unprecedented amounts of comparative data, as a resource for the development of additional diagnostic tools, and finally as a knowledge database about the biological differences between strains and species.

Whole genome comparison of the A. fumigatus family.

The availability of the genome sequences of multiple Aspergillus spp. presents the research community with an unprecedented opportunity for discovery. The genomes of Neosartorya fischeri and Aspergillus clavatus have been sequenced in order to extend our knowledge of Aspergillus fumigatus, the primary cause of invasive aspergillosis. Through comparative genomic analysis, we hope to elucidate both obvious and subtle differences between genomes, developing new hypotheses that can be tested in the laboratory. A preliminary examination of the genomes and their predicted proteomes reveals extensive conservation between protein sequences and significant synteny, or conserved gene order. Comparative genomic analysis at the level of these closely related aspergilli should provide important insight into the evolutionary forces at play and their effect on gene content, regulation and expression.

Whole genome comparison of Aspergillus flavus and A. oryzae.

Aspergillus flavus is a plant and animal pathogen that also produces the potent carcinogen aflatoxin. Aspergillus oryzae is a closely related species that has been used for centuries in the food fermentation industry and is Generally Regarded As Safe (GRAS). Whole genome sequences for these two fungi are now complete, providing us with the opportunity to examine any genomic differences that may explain the different ecological niches of these two fungi, and perhaps to identify pathogenicity factors in A. flavus. These two fungi are very similar in genome size and number of predicted genes. The estimated genome size (36·8 Mb) and predicted number of genes (12 197) for A. flavus is similar to that of A. oryzae (36·7 Mb and 12 079, respectively). These two fungi have significantly larger genomes than Aspergillus nidulans (30·1) and Aspergillus fumigatus (29·4). The A. flavus and A. oryzae genomes are enriched in genes for secondary metabolism, but do not differ greatly from one another in the predicted number of polyketide synthases, nonribosomal peptide synthases or the number of genes coding for cytochrome P450 enzymes. A micro-scale analysis of the two fungi did show differences in DNA correspondence between the two species and in the number of transposable elements. Each species has approximately 350 unique genes. The high degree of sequence similarity between the two fungi suggests that they may be ecotypes of the same species and that A. oryzae has resulted from the domestication of A. flavus.

The sequence of the human genome.

A 2.91-billion base pair (bp) consensus sequence of the euchromatic portion of the human genome was generated by the whole-genome shotgun sequencing method. The 14.8-billion bp DNA sequence was generated over 9 months from 27,271,853 high-quality sequence reads (5.11-fold coverage of the genome) from both ends of plasmid clones made from the DNA of five individuals. Two assembly strategies-a whole-genome assembly and a regional chromosome assembly-were used, each combining sequence data from Celera and the publicly funded genome effort. The public data were shredded into 550-bp segments to create a 2.9-fold coverage of those genome regions that had been sequenced, without including biases inherent in the cloning and assembly procedure used by the publicly funded group. This brought the effective coverage in the assemblies to eightfold, reducing the number and size of gaps in the final assembly over what would be obtained with 5.11-fold coverage. The two assembly strategies yielded very similar results that largely agree with independent mapping data. The assemblies effectively cover the euchromatic regions of the human chromosomes. More than 90% of the genome is in scaffold assemblies of 100,000 bp or more, and 25% of the genome is in scaffolds of 10 million bp or larger. Analysis of the genome sequence revealed 26,588 protein-encoding transcripts for which there was strong corroborating evidence and an additional approximately 12,000 computationally derived genes with mouse matches or other weak supporting evidence. Although gene-dense clusters are obvious, almost half the genes are dispersed in low G+C sequence separated by large tracts of apparently noncoding sequence. Only 1.1% of the genome is spanned by exons, whereas 24% is in introns, with 75% of the genome being intergenic DNA. Duplications of segmental blocks, ranging in size up to chromosomal lengths, are abundant throughout the genome and reveal a complex evolutionary history. Comparative genomic analysis indicates vertebrate expansions of genes associated with neuronal function, with tissue-specific developmental regulation, and with the hemostasis and immune systems. DNA sequence comparisons between the consensus sequence and publicly funded genome data provided locations of 2.1 million single-nucleotide polymorphisms (SNPs). A random pair of human haploid genomes differed at a rate of 1 bp per 1250 on average, but there was marked heterogeneity in the level of polymorphism across the genome. Less than 1% of all SNPs resulted in variation in proteins, but the task of determining which SNPs have functional consequences remains an open challenge.

Identification and characterization of a p53 homologue in Drosophila melanogaster.

The tumor suppressor gene p53 in mammalian cells plays a critical role in safeguarding the integrity of genome. It functions as a sequence-specific transcription factor. Upon activation by a variety of cellular stresses, p53 transactivates downstream target genes, through which it regulates cell cycle and apoptosis. However, little is known about p53 in invertebrates. Here we report the identification and characterization of a Drosophila p53 homologue gene, dp53. dp53 encodes a 385-amino acid protein with significant homology to human p53 (hp53) in the region of the DNA-binding domain, and to a lesser extent the tetramerization domain. Purified dp53 DNA-binding domain protein was shown to bind to the consensus hp53-binding site by gel mobility analysis. In transient transfection assays, expression of dp53 in Schneider cells transcriptionally activated promoters that contained consensus hp53-responsive elements. Moreover, a mutant dp53 (Arg-155 to His-155), like its hp53 counterpart mutant, exerted a dominant-negative effect on transactivation. Ectopic expression of dp53 in Drosophila eye disk caused cell death and led to a rough eye phenotype. dp53 is expressed throughout the development of Drosophila with highest expression levels in early embryogenesis, which has a maternal component. Consistent with this, dp53 RNA levels were high in the nurse cells of the ovary. It appears that p53 is structurally and functionally conserved from flies to mammals. Drosophila will provide a useful genetic system to the further study of the p53 network.

The genome sequence of Drosophila melanogaster.

The fly Drosophila melanogaster is one of the most intensively studied organisms in biology and serves as a model system for the investigation of many developmental and cellular processes common to higher eukaryotes, including humans. We have determined the nucleotide sequence of nearly all of the approximately 120-megabase euchromatic portion of the Drosophila genome using a whole-genome shotgun sequencing strategy supported by extensive clone-based sequence and a high-quality bacterial artificial chromosome physical map. Efforts are under way to close the remaining gaps; however, the sequence is of sufficient accuracy and contiguity to be declared substantially complete and to support an initial analysis of genome structure and preliminary gene annotation and interpretation. The genome encodes approximately 13,600 genes, somewhat fewer than the smaller Caenorhabditis elegans genome, but with comparable functional diversity.

Comparative genomics of the eukaryotes.

A comparative analysis of the genomes of Drosophila melanogaster, Caenorhabditis elegans, and Saccharomyces cerevisiae-and the proteins they are predicted to encode-was undertaken in the context of cellular, developmental, and evolutionary processes. The nonredundant protein sets of flies and worms are similar in size and are only twice that of yeast, but different gene families are expanded in each genome, and the multidomain proteins and signaling pathways of the fly and worm are far more complex than those of yeast. The fly has orthologs to 177 of the 289 human disease genes examined and provides the foundation for rapid analysis of some of the basic processes involved in human disease.

Formation and metabolism of histone mRNAs with mutant 3' ends formed by snRNA termination signals.

Histone mRNAs are the only non-polyadenylated mRNAs, ending in a conserved 26 nt sequence which can form a stem-loop. It has not been possible to make histone mRNAs with mutant stem-loops since most mutations in the stem-loop interfere with the 3' processing reaction. The snRNA genes transcribed by RNA polymerase II form their 3' ends by transcription termination directed by a signal which is located entirely 3' of the snRNA coding sequence. Chimeric genes which express RNAs ending in a histone 3' end or in mutant histone 3' ends formed by snRNA termination signals were constructed. The mRNAs from these genes were efficiently transported from the nucleus after injection of the genes into frog oocytes. This was true even for RNAs which end in mutant stem-loops suggesting that the snRNA termination signals promote transport of the transcripts.