Tumour evolution inferred by single-cell sequencing

Genomic analysis provides insights into the role of copy number variation in disease, but most methods are not designed to resolve mixed populations of cells. In tumours, where genetic heterogeneity is common, very important information may be lost that would be useful for reconstructing evolutionary history. Here we show that with flow-sorted nuclei, whole genome amplification and next generation sequencing we can accurately quantify genomic copy number within an individual nucleus. We apply single-nucleus sequencing to investigate tumour population structure and evolution in two human breast cancer cases. Analysis of 100 single cells from a polygenomic tumour revealed three distinct clonal subpopulations that probably represent sequential clonal expansions. Additional analysis of 100 single cells from a monogenomic primary tumour and its liver metastasis indicated that a single clonal expansion formed the primary tumour and seeded the metastasis. In both primary tumours, we also identified an unexpectedly abundant subpopulation of genetically diverse 'pseudodiploid' cells that do not travel to the metastatic site. In contrast to gradual models of tumour progression, our data indicate that tumours grow by punctuated clonal expansions with few persistent intermediates.     

The extent and patterns of usage of Agent Orange and other herbicides in Vietnam

Herbicides including Agent Orange were sprayed by United States forces for military purposes during the Vietnam War (1961-1971) at a rate more than an order of magnitude greater than for similar domestic weed control. In 1974, the US National Academy of Sciences published estimates of the extent and distribution of herbicides sprayed. Here we present revised estimates, developed using more-complete data. The spray inventory is expanded by more than seven million litres, in particular with heavily dioxin-contaminated herbicides. Estimates for the amount of dioxin sprayed are almost doubled. Hamlet census data reveal that millions of Vietnamese were likely to have been sprayed upon directly. Our identification of specific military herbicide targets has led to a more coherent understanding of spraying. Common errors in earlier interpretations of the spray data are also discussed.     

Tubular and glomerular nephropathy induced in rabbits by injection of a heterologous antitubular basement membrane serum]

A nephropathy affecting both tubules and glomeruli has been induced in Rabbits by the injection of an heterologous immune serum directed against Rabbit tubular basement membrane. It differs from experimental antiglomerular basement membrane nephropathy by the fixation of antibodies and C3 on tubular basement membrane and by the occurrence of glucosuria.     

Regulatory networks define phenotypic classes of human stem cell lines

Stem cells are defined as self-renewing cell populations that can differentiate into multiple distinct cell types. However, hundreds of different human cell lines from embryonic, fetal and adult sources have been called stem cells, even though they range from pluripotent cells-typified by embryonic stem cells, which are capable of virtually unlimited proliferation and differentiation-to adult stem cell lines, which can generate a far more limited repertoire of differentiated cell types. The rapid increase in reports of new sources of stem cells and their anticipated value to regenerative medicine has highlighted the need for a general, reproducible method for classification of these cells. We report here the creation and analysis of a database of global gene expression profiles (which we call the 'stem cell matrix') that enables the classification of cultured human stem cells in the context of a wide variety of pluripotent, multipotent and differentiated cell types. Using an unsupervised clustering method to categorize a collection of approximately 150 cell samples, we discovered that pluripotent stem cell lines group together, whereas other cell types, including brain-derived neural stem cell lines, are very diverse. Using further bioinformatic analysis we uncovered a protein-protein network (PluriNet) that is shared by the pluripotent cells (embryonic stem cells, embryonal carcinomas and induced pluripotent cells). Analysis of published data showed that the PluriNet seems to be a common characteristic of pluripotent cells, including mouse embryonic stem and induced pluripotent cells and human oocytes. Our results offer a new strategy for classifying stem cells and support the idea that pluripotency and self-renewal are under tight control by specific molecular networks.     

Water stress and hybridization between Quercus gambelii and Quercus grisea

The historical and environmental forces involved in determining the geographic location of plant hybrid zones have long been of interest. Often hybrid zones appear to be intimately tied to the environment, yet because many abiotic factors covary it is often difficult to understand which are truly the most important in maintaining the position and structure of a hybrid zone. This study uses empirical data and modeling analyses to examine whether abiotic factors are responsible for the location and structure of an oak ( Quercus ) hybrid zone and, if so, which factors are the most important. Specifically, leaf trait measurements and ecological niche models were used to test the hypothesis that water availability plays a primary role in promoting and maintaining the location of hybridization between 2 species. Leaf trait analyses and ecological niche models both supported the hypothesis that water availability determines the location and structure of the hybrid zone. These findings lend support for the general importance of environmental factors in determining hybrid zone location and structure. Furthermore, they demonstrate how functional trait analyses and predictive ecological niche models can be used in future hybrid zone research.     

Outbred embryos rescue inbred half-siblings in mixed-paternity broods of live-bearing females

Females commonly mate with more than one male, and polyandry has been shown to increase reproductive success in many species. Insemination by multiple males shifts the arena for sexual selection from the external environment to the female reproductive tract, where sperm competition or female choice of sperm could bias fertilization against sperm from genetically inferior or genetically incompatible males. Evidence that polyandry can be a strategy for avoiding incompatibility comes from studies showing that inbreeding cost is reduced in some egg-laying species by postcopulatory mechanisms that favour fertilization by sperm from unrelated males. In viviparous (live-bearing) species, inbreeding not only reduces offspring genetic quality but might also disrupt feto-maternal interactions that are crucial for normal embryonic development. Here we show that polyandry in viviparous pseudoscorpions reduces inbreeding cost not through paternity-biasing mechanisms favouring outbred offspring, but rather because outbred embryos exert a rescuing effect on inbred half-siblings in mixed-paternity broods. The benefits of polyandry may thus be more complex for live-bearing females than for females that lay eggs.     

Estimating root biomass and distribution after fire in a Great Basin woodland using cores and pits

Quantifying root biomass is critical to an estimation and understanding of ecosystem net primary production, biomass partitioning, and belowground competition. We compared 2 methods for determining root biomass: a new soil-coring technique and traditional excavation of quantitative pits. We conducted the study in an existing Joint Fire Sciences demonstration area in the central Great Basin. This area is representative of a shrub (sagebrush) ecosystem exhibiting tree (pinyon and juniper) encroachment. The demonstration area had a prescribed burn implemented 4 years prior to our study, and we sampled both control and burned plots. The samples were stratified across 3 microsites (interspace, under shrub, and under tree) and 4 soil depths (0–8, 8–23, 23–38, and 38–52 cm) to determine the effects of plant life form and burning on root biomass. We found that estimates of total root biomass were similar between quantitative pits and our soil cores. However, cores tended to show a more even distribution of root biomass across all microsites and depths than did pits. Overall, results indicated that root biomass differs significantly among microsites and soil depths and that the amount of root biomass at a given depth differs among microsites. Burning reduced root biomass in our study by 23% and altered the spatial distribution of root mass.     

Maternal Rnf12/RLIM is required for imprinted X-chromosome inactivation in mice

Two forms of X-chromosome inactivation (XCI) ensure the selective silencing of female sex chromosomes during mouse embryogenesis. Imprinted XCI begins with the detection of Xist RNA expression on the paternal X?chromosome (Xp) at about the four-cell stage of embryonic development. In the embryonic tissues of the inner cell mass, a random form of XCI occurs in blastocysts that inactivates either Xp or the maternal X?chromosome (Xm). Both forms of XCI require the non-coding Xist RNA that coats the inactive X?chromosome from which it is expressed. Xist has crucial functions in the silencing of X-linked genes, including Rnf12 (refs 3, 4) encoding the ubiquitin ligase RLIM (RING finger LIM-domain-interacting protein). Here we show, by targeting a conditional knockout of Rnf12 to oocytes where RLIM accumulates to high levels, that the maternal transmission of the mutant X?chromosome (Δm) leads to lethality in female embryos as a result of defective imprinted XCI. We provide evidence that in Δm female embryos the initial formation of Xist clouds and Xp silencing are inhibited. In contrast, embryonic stem cells lacking RLIM are able to form Xist clouds and silence at least some X-linked genes during random XCI. These results assign crucial functions to the maternal deposit of Rnf12/RLIM for the initiation of imprinted XCI.