DNA sequence of human chromosome 17 and analysis of rearrangement in the human lineage.

Chromosome 17 is unusual among the human chromosomes in many respects. It is the largest human autosome with orthology to only a single mouse chromosome, mapping entirely to the distal half of mouse chromosome 11. Chromosome 17 is rich in protein-coding genes, having the second highest gene density in the genome. It is also enriched in segmental duplications, ranking third in density among the autosomes. Here we report a finished sequence for human chromosome 17, as well as a structural comparison with the finished sequence for mouse chromosome 11, the first finished mouse chromosome. Comparison of the orthologous regions reveals striking differences. In contrast to the typical pattern seen in mammalian evolution, the human sequence has undergone extensive intrachromosomal rearrangement, whereas the mouse sequence has been remarkably stable. Moreover, although the human sequence has a high density of segmental duplication, the mouse sequence has a very low density. Notably, these segmental duplications correspond closely to the sites of structural rearrangement, demonstrating a link between duplication and rearrangement. Examination of the main classes of duplicated segments provides insight into the dynamics underlying expansion of chromosome-specific, low-copy repeats in the human genome.

DNA sequence and analysis of human chromosome 8.

The International Human Genome Sequencing Consortium (IHGSC) recently completed a sequence of the human genome. As part of this project, we have focused on chromosome 8. Although some chromosomes exhibit extreme characteristics in terms of length, gene content, repeat content and fraction segmentally duplicated, chromosome 8 is distinctly typical in character, being very close to the genome median in each of these aspects. This work describes a finished sequence and gene catalogue for the chromosome, which represents just over 5% of the euchromatic human genome. A unique feature of the chromosome is a vast region of approximately 15 megabases on distal 8p that appears to have a strikingly high mutation rate, which has accelerated in the hominids relative to other sequenced mammals. This fast-evolving region contains a number of genes related to innate immunity and the nervous system, including loci that appear to be under positive selection--these include the major defensin (DEF) gene cluster and MCPH1, a gene that may have contributed to the evolution of expanded brain size in the great apes. The data from chromosome 8 should allow a better understanding of both normal and disease biology and genome evolution.

The tyrosine phosphatase PTPRD is a tumor suppressor that is frequently inactivated and mutated in glioblastoma and other human cancers.

Tyrosine phosphorylation plays a critical role in regulating cellular function and is a central feature in signaling cascades involved in oncogenesis. The regulation of tyrosine phosphorylation is coordinately controlled by kinases and phosphatases (PTPs). Whereas activation of tyrosine kinases has been shown to play vital roles in tumor development, the role of PTPs is much less well defined. Here, we show that the receptor protein tyrosine phosphatase delta (PTPRD) is frequently inactivated in glioblastoma multiforme (GBM), a deadly primary neoplasm of the brain. PTPRD is a target of deletion in GBM, often via focal intragenic loss. In GBM tumors that do not possess deletions in PTPRD, the gene is frequently subject to cancer-specific epigenetic silencing via promoter CpG island hypermethylation (37%). Sequencing of the PTPRD gene in GBM and other primary human tumors revealed that the gene is mutated in 6% of GBMs, 13% of head and neck squamous cell carcinomas, and in 9% of lung cancers. These mutations were deleterious. In total, PTPRD inactivation occurs in >50% of GBM tumors, and loss of expression predicts for poor prognosis in glioma patients. Wild-type PTPRD inhibits the growth of GBM and other tumor cells, an effect not observed with PTPRD alleles harboring cancer-specific mutations. Human astrocytes lacking PTPRD exhibited increased growth. PTPRD was found to dephosphorylate the oncoprotein STAT3. These results implicate PTPRD as a tumor suppressor on chromosome 9p that is involved in the development of GBMs and multiple human cancers.

A high-density genetic linkage map of a black spruce (Picea mariana) × red spruce (Picea rubens) interspecific hybrid.

Genetic maps provide an important genomic resource of basic and applied significance. Spruce (Picea) has a very large genome size (between 0.85 × 1010 and 2.4 × 1010 bp; 8.5-24.0 pg/1C, a mean of 17.7 pg/1C ). We have constructed a near-saturated genetic linkage map for an interspecific backcross (BC1) hybrid of black spruce (BS; Picea mariana (Mill.) B.S.P.) and red spruce (RS; Picea rubens Sarg.), using selectively amplified microsatellite polymorphic loci (SAMPL) markers. A total of 2284 SAMPL markers were resolved using 31 SAMPL-MseI selective nucleotide primer combinations. Of these, 1216 SAMPL markers showing Mendelian segregation were mapped, whereas 1068 (46.8%) SAMPL fragments showed segregation distortion at α = 0.05. Maternal, paternal, and consensus maps consistently coalesced into 12 linkage groups, corresponding to the haploid chromosome number (1n = 1x = 12) of 12 in the genus Picea. The maternal BS map consisted of 814 markers distributed over 12 linkage groups, covering 1670 cM, with a mean map distance of 2.1 cM between adjacent markers. The paternal BS × RS map consisted of 773 markers distributed over 12 linkage groups, covering 1563 cM, with a mean map distance of 2.0 cM between adjacent markers. The consensus interspecific hybrid BC1 map consisted of 1216 markers distributed over 12 linkage groups, covering 1865 cM (98% genome coverage), with a mean map distance of 1.5 cM between adjacent markers. The genetic map reported here provides an important genomic resource in Picea, Pinaceae, and conifers.

The discovery of benzanilides as c-Met receptor tyrosine kinase inhibitors by a directed screening approach.

A directed screen of a relatively small number of compounds, selected for kinase ATP pocket binding potential, yielded a novel series of hit compounds (1). Hit explosion on two binding residues identified compounds 27 and 43 as the best leads for an optimization program having reduced secondary metabolism, as measured by in vitro rat hepatocytes incubation, leading to oral bio-availability. Structure-activity relationships and molecular modeling have suggested a binding mode for the most potent inhibitor 12.

Near-saturated and complete genetic linkage map of black spruce (Picea mariana).

BACKGROUND: Genetic maps provide an important genomic resource for understanding genome organization and evolution, comparative genomics, mapping genes and quantitative trait loci, and associating genomic segments with phenotypic traits. Spruce (Picea) genomics work is quite challenging, mainly because of extremely large size and highly repetitive nature of its genome, unsequenced and poorly understood genome, and the general lack of advanced-generation pedigrees. Our goal was to construct a high-density genetic linkage map of black spruce (Picea mariana, 2n = 24), which is a predominant, transcontinental species of the North American boreal and temperate forests, with high ecological and economic importance. RESULTS: We have developed a near-saturated and complete genetic linkage map of black spruce using a three-generation outbred pedigree and amplified fragment length polymorphism (AFLP), selectively amplified microsatellite polymorphic loci (SAMPL), expressed sequence tag polymorphism (ESTP), and microsatellite (mostly cDNA based) markers. Maternal, paternal, and consensus genetic linkage maps were constructed. The maternal, paternal, and consensus maps in our study consistently coalesced into 12 linkage groups, corresponding to the haploid chromosome number (1n = 1x = 12) of 12 in the genus Picea. The maternal map had 816 and the paternal map 743 markers distributed over 12 linkage groups each. The consensus map consisted of 1,111 markers distributed over 12 linkage groups, and covered almost the entire (> 97%) black spruce genome. The mapped markers included 809 AFLPs, 255 SAMPL, 42 microsatellites, and 5 ESTPs. Total estimated length of the genetic map was 1,770 cM, with an average of one marker every 1.6 cM. The maternal, paternal and consensus genetic maps aligned almost perfectly. CONCLUSION: We have constructed the first high density to near-saturated genetic linkage map of black spruce, with greater than 97% genome coverage. Also, this is the first genetic map based on a three-generation outbred pedigree in the genus Picea. The genome length in P. mariana is likely to be about 1,800 cM. The genetic maps developed in our study can serve as a reference map for various genomics studies and applications in Picea and Pinaceae.

CancerGenes: a gene selection resource for cancer genome projects.

The genome sequence framework provided by the human genome project allows us to precisely map human genetic variations in order to study their association with disease and their direct effects on gene function. Since the description of tumor suppressor genes and oncogenes several decades ago, both germ-line variations and somatic mutations have been established to be important in cancer-in terms of risk, oncogenesis, prognosis and response to therapy. The Cancer Genome Atlas initiative proposed by the NIH is poised to elucidate the contribution of somatic mutations to cancer development and progression through the re-sequencing of a substantial fraction of the total collection of human genes-in hundreds of individual tumors and spanning several tumor types. We have developed the CancerGenes resource to simplify the process of gene selection and prioritization in large collaborative projects. CancerGenes combines gene lists annotated by experts with information from key public databases. Each gene is annotated with gene name(s), functional description, organism, chromosome number, location, Entrez Gene ID, GO terms, InterPro descriptions, gene structure, protein length, transcript count, and experimentally determined transcript control regions, as well as links to Entrez Gene, COSMIC, and iHOP gene pages and the UCSC and Ensembl genome browsers. The user-friendly interface provides for searching, sorting and intersection of gene lists. Users may view tabulated results through a web browser or may dynamically download them as a spreadsheet table. CancerGenes is available at http://cbio.mskcc.org/cancergenes.

Genomic mutation consequence calculator.

UNLABELLED: The genomic mutation consequence calculator (GMCC) is a tool that will reliably and quickly calculate the consequence of arbitrary genomic mutations. GMCC also reports supporting annotations for the specified genomic region. The particular strength of the GMCC is it works in genomic space, not simply in spliced transcript space as some similar tools do. Within gene features, GMCC can report on the effects on splice site, UTR and coding regions in all isoforms affected by the mutation. A considerable number of genomic annotations are also reported, including: genomic conservation score, known SNPs, COSMIC mutations, disease associations and others. The manual interface also offers link outs to various external databases and resources. In batch mode, GMCC returns a csv file which can easily be parsed by the end user. AUDIENCE: GMCC is intended to support the many tumor resequencing efforts, but can be useful to any study investigating genomic mutations.

Genetic variation and control of chloroplast pigment concentrations in Picea rubens, Picea mariana and their hybrids. I. Ambient and elevated [CO2] environments.

Traits related to light-energy processing have significant ecological implications for plant fitness. We studied the effects of elevated atmospheric CO(2) concentration ([CO(2)]) on chloroplast pigment traits of a red spruce (RS) (Picea rubens Sarg.)-black spruce (BS) (P. mariana (Mill.) B.S.P.) genetic complex in two experiments: (1) a comparative species' provenance experiment from across the near-northern part of the RS range; and (2) an intra- and interspecific controlled-cross experiment. Results from the provenance experiment showed that total chlorophyll (a + b) concentration was, on average, 15% higher in ambient [CO(2)] than in elevated [CO(2)] (P < 0.001). In ambient [CO(2)], BS populations averaged 11% higher total chlorophyll and carotenoid concentrations than RS populations (P < 0.001). There were significant species, CO(2), and species x CO(2) interaction effects, with chlorophyll concentration decreasing about 7 and 26% for BS and RS, respectively, in response to elevated [CO(2)]. Results from the controlled-cross experiment showed that families with a hybrid index of 25 (25% RS) had the highest total chlorophyll concentrations, and families with hybrid indices of 75 and 100 had among the lowest amounts. Initial analysis of the controlled-cross experiment supported a more additive model of inheritance; however, parental analysis showed a significant and predominant male effect for chlorophyll concentration. In ambient and elevated [CO(2)] environments, crosses with BS males had 10.6 and 17.6% higher total chlorophyll concentrations than crosses with hybrid and RS males, respectively. Our results show that chlorophyll concentration is under strong genetic control, and that these traits are positively correlated with productivity within and across species. A significant positive correlation between chlorophyll concentration and the ratio of total plant N to root dry mass was also found (r = 0.872). The almost fourfold decrease in chlorophyll concentration in RS suggests that it would be at a competitive disadvantage compared with BS in a high [CO(2)] environment.

Selfing results in inbreeding depression of growth but not of gas exchange of surviving adult black spruce trees.

In most tree species, inbreeding greatly reduces seed production, seed viability, survival and growth. In a previous large-scale quantitative analysis of a black spruce (Picea mariana (Mill.) B.S.P.) diallel experiment, selfing had large deleterious effects on growth but no impact on stable carbon isotope discrimination (an indirect measure of the ratio of net photosynthesis (A) to stomatal conductance (gwv)). It was hypothesized that selfing has no effect on carbon (C) fixation at the leaf level but impairs subsequent utilization of C. Alternatively, A and gwv may be impacted by selfing to the same extent. However, no gas exchange data were collected to test these hypotheses. We have now obtained photosynthetic gas exchange data from three selfed families and three outcrossed families (all the result of controlled pollination) from the same diallel experiment. Photosynthetic responses to intercellular CO2 concentration (A-Ci curves) were generated on four replicates per family, one block per day, over a 4-day period in July. There were no differences between selfed and outcrossed families in maximum carboxylation rate, maximum electron transport, A or gwv (both estimated at 370 ppm CO2), or the ratio A/gwv. Because selfed trees had higher mortality than outcrossed trees during the experiment, we cannot exclude the possibility that previously living selfed progeny had low A. Nevertheless, the data indicate that inbreeding can result in trees that have low productivity despite high A, supporting our hypothesis that gas exchange is similar between selfed and outcrossed progeny trees. We conclude that utilization of fixed C is modified in the surviving selfed progeny.

Light-energy processing and freezing-tolerance traits in red spruce and black spruce: species and seed-source variation.

Red spruce (Picea rubens Sarg.) and black spruce (Picea mariana (Mill.) B.S.P.) are genetically and morphologically similar but ecologically distinct species. We determined intraspecific seed-source and interspecific variation of red spruce and black spruce, from across the near-northern margins of their ranges, for several light-energy processing and freezing-tolerance adaptive traits. Before exposure to low temperature, red spruce had variable fluorescence (Fv) similar to black spruce, but higher photochemical efficiency (Fv/Fm), lower quantum yield, lower chlorophyll fluorescence (%), and higher thermal dissipation efficiency (qN), although the seed-source effect and the seed-source x species interaction were significant only for Fv/Fm. After low-temperature exposure (-40 degrees C), red spruce had significantly lower Fv/Fm, quantum yield and qN than black spruce, but higher chlorophyll fluorescence and relative fluorescence. Species, seed-source effect, and seed-source x species interaction were consistent with predictions based on genetic (e.g., geographic) origins. Multi-temperature exposures (5, -20 and -40 degrees C) often produced significant species and temperature effects, and species x temperature interactions as a result of species-specific responses to temperature exposures. The inherent physiological species-specific adaptations of red spruce and black spruce were largely consistent with a shade-tolerant, late-successional species and an early successional species, respectively. Species differences in physiological adaptations conform to a biological trade-off, probably as a result of natural selection pressure in response to light availability and prevailing temperature gradients.

Integrative genomic profiling of human prostate cancer.

Annotation of prostate cancer genomes provides a foundation for discoveries that can impact disease understanding and treatment. Concordant assessment of DNA copy number, mRNA expression, and focused exon resequencing in 218 prostate cancer tumors identified the nuclear receptor coactivator NCOA2 as an oncogene in approximately 11% of tumors. Additionally, the androgen-driven TMPRSS2-ERG fusion was associated with a previously unrecognized, prostate-specific deletion at chromosome 3p14 that implicates FOXP1, RYBP, and SHQ1 as potential cooperative tumor suppressors. DNA copy-number data from primary tumors revealed that copy-number alterations robustly define clusters of low- and high-risk disease beyond that achieved by Gleason score. The genomic and clinical outcome data from these patients are now made available as a public resource.

Subtype-specific genomic alterations define new targets for soft-tissue sarcoma therapy.

Soft-tissue sarcomas, which result in approximately 10,700 diagnoses and 3,800 deaths per year in the United States, show remarkable histologic diversity, with more than 50 recognized subtypes. However, knowledge of their genomic alterations is limited. We describe an integrative analysis of DNA sequence, copy number and mRNA expression in 207 samples encompassing seven major subtypes. Frequently mutated genes included TP53 (17% of pleomorphic liposarcomas), NF1 (10.5% of myxofibrosarcomas and 8% of pleomorphic liposarcomas) and PIK3CA (18% of myxoid/round-cell liposarcomas, or MRCs). PIK3CA mutations in MRCs were associated with Akt activation and poor clinical outcomes. In myxofibrosarcomas and pleomorphic liposarcomas, we found both point mutations and genomic deletions affecting the tumor suppressor NF1. Finally, we found that short hairpin RNA (shRNA)-based knockdown of several genes amplified in dedifferentiated liposarcoma, including CDK4 and YEATS4, decreased cell proliferation. Our study yields a detailed map of molecular alterations across diverse sarcoma subtypes and suggests potential subtype-specific targets for therapy.

Shoot water relations of mature black spruce families displaying a genotype x environment interaction in growth rate. I. Family and site effects over three growing seasons.

Pressure-volume curves were determined for black spruce (Picea mariana (Mill.) BSP) trees from four full-sib families. During the first two years, trees were measured from a plantation on a dry site. In the third year, trees were sampled from the dry site and a wet site. Diurnal measurements of shoot water potential allowed in situ shoot turgor to be estimated in addition to standard water relations traits. Over all years, Female 59 progeny displayed lower osmotic potentials at saturation (Psi(sat)) than Female 63 progeny. Genetic differences in Psi(sat) were similar on both the dry and wet sites. Modulus of elasticity (epsilon) was greater for Female 59 progeny than for Female 63 progeny, producing a compensatory effect resulting in no genetic or site differences in osmotic potential at turgor loss point (Psi(tlp)) or relative water content at turgor loss point (RWC(tlp)). Mean and predawn shoot turgor pressures (P(x) and P(pd)) were higher for Female 59 progeny than for Female 63 progeny and higher at the wet site than the dry site. Genotype x environment trends were observed; compared to Female 63 progeny, Female 59 progeny displayed 9.8 and 5.1% higher P(pd) on the dry and wet sites, respectively, and 3.4 and 9.8% greater P(pd) values in wet and dry years, respectively. Tree volume growth showed no relationship to Psi(tlp) or RWC(tlp), but was correlated with Psi(sat) and P(x); however, the strongest correlation was with P(pd) (r = 0.90).

Shoot water relations of mature black spruce families displaying a genotype x environment interaction in growth rate. II. Temporal trends and response to varying soil water conditions.

Pressure-volume curves and shoot water potentials were determined for black spruce (Picea mariana (Mill.) BSP) trees from four full-sib families at the Petawawa Research Forest, Ontario, Canada. Trees were sampled from a dry site in 1992 and from the dry site and a wet site in 1993. Modulus of elasticity (epsilon), osmotic potential at turgor loss point (Psi(tlp)) and relative water at turgor loss point (RWC(tlp)) all decreased during the growing season. Osmotic potential at saturation (Psi(sat)) and turgor displayed no general temporal trend. Across a range of environmental conditions, Female 59 progeny had equal or lower Psi(sat), and higher or similar epsilon, mean turgor pressure (P(x)) and predawn turgor pressure (P(pd)) compared with Female 63 progeny. Osmotic potential at saturation decreased as water stress increased from mild to moderate and increased as water stress increased from moderate to severe. Stable genetic differences in Psi(sat) were maintained by the same rate of osmotic adjustment from low to moderate water stress. Modulus of elasticity and RWC(tlp) decreased with decreasing water availability, whereas Psi(tlp) showed no response. The combined effects of Psi(sat) and epsilon resulted in no change in P(pd) as water stress increased from low to moderate values, but turgor declined sharply as water stress increased from moderate to high values. We conclude that drought tolerance traits strongly influence the growth of these black spruce families across sites of varying water availability.

The complete genome and proteome of Mycoplasma mobile.

Although often considered "minimal" organisms, mycoplasmas show a wide range of diversity with respect to host environment, phenotypic traits, and pathogenicity. Here we report the complete genomic sequence and proteogenomic map for the piscine mycoplasma Mycoplasma mobile, noted for its robust gliding motility. For the first time, proteomic data are used in the primary annotation of a new genome, providing validation of expression for many of the predicted proteins. Several novel features were discovered including a long repeating unit of DNA of approximately 2435 bp present in five complete copies that are shown to code for nearly identical yet uniquely expressed proteins. M. mobile has among the lowest DNA GC contents (24.9%) and most reduced set of tRNAs of any organism yet reported (28). Numerous instances of tandem duplication as well as lateral gene transfer are evident in the genome. The multiple available complete genome sequences for other motile and immotile mycoplasmas enabled us to use comparative genomic and phylogenetic methods to suggest several candidate genes that might be involved in motility. The results of these analyses leave open the possibility that gliding motility might have arisen independently more than once in the mycoplasma lineage.