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The harbour porpoise (Phocoena phocoena), a highly mobile cetacean species of the Northern Hemisphere, inhabits basins that vary broadly in salinity, temperature, and food availability; such variation can drive divergent adaptation among local populations. To shed light on range-wide population structure and local adaptation, we generated ddRAD sequencing data spanning the entire North Atlantic and the Baltic Sea, as well as the Black Sea as an outgroup, and mapped this data to the high-quality draft genome of the species. We identified 11,978 genome-wide SNPs from 150 individuals, which we used for population genetic inferences. Our results support genetic differentiation between North Atlantic and Baltic Sea populations, with Kattegat as a transition zone. Across the North Atlantic the population differentiation is subtle from west to east, congruent with an isolation-by-distance pattern, but indicates a separation of southern North Sea harbour porpoises. We identified genomic outlier regions, i.e., scaffold regions where SNPs with high F ST across North Atlantic populations co-occur. Together with the draft genome annotation, these regions could point towards candidate genes for differential local adaptation processes among populations. Furthermore, they enable the development of a SNP panel for routine population assignment which will be useful in a conservation and management context. We identified six outlier loci putatively under positive selection, based on the population structure inferred from the complete SNP set. Our study highlights the value of genome resources in conservation and management and provides a crucial additional resource for the study of harbour porpoise evolution and phylogeny.

The harbour porpoise (Phocoena phocoena), a highly mobile cetacean species of the Northern Hemisphere, inhabits basins that vary broadly in salinity, temperature, and food availability; such variation can drive divergent adaptation among local populations. To shed light on range-wide population structure and local adaptation, we generated ddRAD sequencing data spanning the entire North Atlantic and the Baltic Sea, as well as the Black Sea as an outgroup, and mapped this data to the high-quality draft genome of the species. We identified 11,978 genome-wide SNPs from 150 individuals, which we used for population genetic inferences. Our results support genetic differentiation between North Atlantic and Baltic Sea populations, with Kattegat as a transition zone. Across the North Atlantic the population differentiation is subtle from west to east, congruent with an isolation-by-distance pattern, but indicates a separation of southern North Sea harbour porpoises. We identified genomic outlier regions, i.e., scaffold regions where SNPs with high F ST across North Atlantic populations co-occur. Together with the draft genome annotation, these regions could point towards candidate genes for differential local adaptation processes among populations. Furthermore, they enable the development of a SNP panel for routine population assignment which will be useful in a conservation and management context. We identified six outlier loci putatively under positive selection, based on the population structure inferred from the complete SNP set. Our study highlights the value of genome resources in conservation and management and provides a crucial additional resource for the study of harbour porpoise evolution and phylogeny.

Cognitive functions of individuals with psychiatric disorders differ from that of the general population. Such cognitive differences often manifest early in life as differential school performance and have a strong genetic basis. Here we measured genetic predictors of school performance in 30,982 individuals in English, Danish and mathematics via a genome-wide association study (GWAS) and studied their relationship with risk for six major psychiatric disorders. When decomposing the school performance into math and language-specific performances, we observed phenotypically and genetically a strong negative correlation between math performance and risk for most psychiatric disorders. But language performance correlated positively with risk for certain disorders, especially schizophrenia, which we replicate in an independent sample (n = 4547). We also found that the genetic variants relating to increased risk for schizophrenia and better language performance are overrepresented in individuals involved in creative professions (n = 2953) compared to the general population (n = 164,622). The findings together suggest that language ability, creativity and psychopathology might stem from overlapping genetic roots.

Cognitive functions of individuals with psychiatric disorders differ from that of the general population. Such cognitive differences often manifest early in life as differential school performance and have a strong genetic basis. Here we measured genetic predictors of school performance in 30,982 individuals in English, Danish and mathematics via a genome-wide association study (GWAS) and studied their relationship with risk for six major psychiatric disorders. When decomposing the school performance into math and language-specific performances, we observed phenotypically and genetically a strong negative correlation between math performance and risk for most psychiatric disorders. But language performance correlated positively with risk for certain disorders, especially schizophrenia, which we replicate in an independent sample (n = 4547). We also found that the genetic variants relating to increased risk for schizophrenia and better language performance are overrepresented in individuals involved in creative professions (n = 2953) compared to the general population (n = 164,622). The findings together suggest that language ability, creativity and psychopathology might stem from overlapping genetic roots.

AbstractGermline genome sequencing (GS) holds great promise for cancer prevention by identifying cancer risk and guiding prevention strategies, however research evidence is mixed regarding patient preferences for receiving GS results. The aim of this study was to discern preferences for return of results by cancer patients who have actually undergone GS. We conducted a mixed methods study with a cohort of cancer probands (n = 335) and their genetic relatives (n = 199) undergoing GS in a research setting. Both groups completed surveys when giving consent. A subset of participants (n = 40) completed semi-structured interviews. A significantly higher percentage of probands thought people would like to be informed about genetic conditions for which there is prevention or treatment that can change cancer risk compared to conditions for which there is no prevention or treatment (93% [311] versus 65% [216]; p < 0.001). Similar results were obtained for relatives (91% [180] versus 61% [121]; p < 0.001). Themes identified in the analysis of interviews were: (1) Recognised benefits of GS, (2) Balancing benefits with risks, (3) Uncertain results are perceived as unhelpful and (4) Competing obligations. While utility was an important discriminator in what was seen as valuable for this cohort, there was a variety of responses. In view of varied participant preferences regarding return of results, it is important to ensure patient understanding of test validity and identify individual choices at the time of consent to GS. The nature and value of the information, and a contextual understanding of researcher obligations should guide res

Swedish populations of Wels catfish (Silurus glanis) experienced severe declines during the 19th and 20th centuries. The main causes for the decline were loss of suitable spawning habitat and fragmentation of populations. Currently, three native and two reintroduced populations remain in Sweden. Thanks to national protection and progress in restoring habitats, population sizes have increased over the past three decades. Previous studies that used microsatellite loci have found that genetic diversity and effective population sizes in Sweden are low, while population differentiation is high. A study that used whole genome sequencing (WGS) confirmed these results for native Swedish populations (those found in the Båven, Emån, and Möckeln water systems). The current project uses the same WGS methods and expands on the previous study by considering samples from non-Swedish populations (river Garonne, France; river Ebro, Spain; hatchery, Czech Republic), as well as improving read-depth coverage and sampling from the introduced Swedish population in the Helge å water system. Both a genome-wide SNP-set and full mitochondrial sequences were used to assess genetic diversity within each population, and differentiation among them. Genetic diversity in Swedish populations is lower than in non-Swedish populations. Native Swedish populations are strongly differentiated from one another. The introduced Helge å population is strongly differentiated from Emån and Möckeln, but less so from Båven. Despite Helge å individuals having heritage that can be predominantly traced to Båven, there are clear signs of admixture with the other two native populations. Swedish populations are all strongly differentiated from the non-Swedish populations. Altogether, evidence of admixture and slightly greater genetic diversity than native Swedish populations in Helge å can at the surface be seen as promising signs. However, it remains uncertain whether these improvements are durable over multiple gene

Swedish populations of Wels catfish (Silurus glanis) experienced severe declines during the 19th and 20th centuries. The main causes for the decline were loss of suitable spawning habitat and fragmentation of populations. Currently, three native and two reintroduced populations remain in Sweden. Thanks to national protection and progress in restoring habitats, population sizes have increased over the past three decades. Previous studies that used microsatellite loci have found that genetic diversity and effective population sizes in Sweden are low, while population differentiation is high. A study that used whole genome sequencing (WGS) confirmed these results for native Swedish populations (those found in the Båven, Emån, and Möckeln water systems). The current project uses the same WGS methods and expands on the previous study by considering samples from non-Swedish populations (river Garonne, France; river Ebro, Spain; hatchery, Czech Republic), as well as improving read-depth coverage and sampling from the introduced Swedish population in the Helge å water system. Both a genome-wide SNP-set and full mitochondrial sequences were used to assess genetic diversity within each population, and differentiation among them. Genetic diversity in Swedish populations is lower than in non-Swedish populations. Native Swedish populations are strongly differentiated from one another. The introduced Helge å population is strongly differentiated from Emån and Möckeln, but less so from Båven. Despite Helge å individuals having heritage that can be predominantly traced to Båven, there are clear signs of admixture with the other two native populations. Swedish populations are all strongly differentiated from the non-Swedish populations. Altogether, evidence of admixture and slightly greater genetic diversity than native Swedish populations in Helge å can at the surface be seen as promising signs. However, it remains uncertain whether these improvements are durable over multiple gene

Despite Africa's central role in the origin of our species, our knowledge of the genomic diversity in Africa is remarkably sparse. A recent publication by Choudhury et al. underscores the scientific imperative for a broader characterisation of African genomic diversity to better understand demographic history and improve global human health.

Background: The genomic and transcriptomic landscape of widely invasive follicular thyroid carcinomas (wiFTCs) and Hurthle cell carcinoma (HCC) are poorly characterized, and subsets of these tumors lack information on genetic driver events. Objective: The aim of this study was to bridge this gap. Methods: We performed whole-genome and RNA sequencing and subsequent bioinformatic analyses of 11 wiFTCs and 2 HCCs with a particularly poor prognosis, and matched normal tissue. Results: All wiFTCs exhibited one or several mutations in established thyroid cancer genes, including TERT (n=4), NRAS (n=3), HRAS, KRAS, AKT, PTEN, PIK3CA, MUTYH, TSHR, and MEN1 (n=1 each). MutSig2CV analysis revealed recurrent somatic mutations in FAM72D (n=3, in 2 wiFTCs and in a single HCC), TP53 (n=3, in 2 wiFTCs and a single HCC), and EIF1AX (n=3), with DGCR8 (n=2) as borderline significant. The DGCR8 mutations were recurrent p.E518K missense alterations, known to cause familial multinodular goiter via disruption of microRNA (miRNA) processing. Expression analyses showed reduced DGCR8 messenger RNA expression in FTCs in general, and the 2 DGCR8 mutants displayed a distinct miRNA profile compared to DGCR8 wild-types. Copy number analyses revealed recurrent gains on chromosomes 4, 6, and 10, and fusion gene analyses revealed 27 high-quality events. Both HCCs displayed hyperploidy, which was fairly unusual in the FTC cohort. Based on the transcriptome data, tumors amassed in 2 principal clusters. Conclusion: We describe the genomic and transcriptomic landscape in wiFTCs and HCCs and identify novel recurrent mutations and copy number alterations with possible driver properties and lay the foundation for future studies.

Background: The genomic and transcriptomic landscape of widely invasive follicular thyroid carcinomas (wiFTCs) and Hurthle cell carcinoma (HCC) are poorly characterized, and subsets of these tumors lack information on genetic driver events. Objective: The aim of this study was to bridge this gap. Methods: We performed whole-genome and RNA sequencing and subsequent bioinformatic analyses of 11 wiFTCs and 2 HCCs with a particularly poor prognosis, and matched normal tissue. Results: All wiFTCs exhibited one or several mutations in established thyroid cancer genes, including TERT (n=4), NRAS (n=3), HRAS, KRAS, AKT, PTEN, PIK3CA, MUTYH, TSHR, and MEN1 (n=1 each). MutSig2CV analysis revealed recurrent somatic mutations in FAM72D (n=3, in 2 wiFTCs and in a single HCC), TP53 (n=3, in 2 wiFTCs and a single HCC), and EIF1AX (n=3), with DGCR8 (n=2) as borderline significant. The DGCR8 mutations were recurrent p.E518K missense alterations, known to cause familial multinodular goiter via disruption of microRNA (miRNA) processing. Expression analyses showed reduced DGCR8 messenger RNA expression in FTCs in general, and the 2 DGCR8 mutants displayed a distinct miRNA profile compared to DGCR8 wild-types. Copy number analyses revealed recurrent gains on chromosomes 4, 6, and 10, and fusion gene analyses revealed 27 high-quality events. Both HCCs displayed hyperploidy, which was fairly unusual in the FTC cohort. Based on the transcriptome data, tumors amassed in 2 principal clusters. Conclusion: We describe the genomic and transcriptomic landscape in wiFTCs and HCCs and identify novel recurrent mutations and copy number alterations with possible driver properties and lay the foundation for future studies.

Background: The genomic and transcriptomic landscape of widely invasive follicular thyroid carcinomas (wiFTCs) and Hurthle cell carcinoma (HCC) are poorly characterized, and subsets of these tumors lack information on genetic driver events. Objective: The aim of this study was to bridge this gap. Methods: We performed whole-genome and RNA sequencing and subsequent bioinformatic analyses of 11 wiFTCs and 2 HCCs with a particularly poor prognosis, and matched normal tissue. Results: All wiFTCs exhibited one or several mutations in established thyroid cancer genes, including TERT (n=4), NRAS (n=3), HRAS, KRAS, AKT, PTEN, PIK3CA, MUTYH, TSHR, and MEN1 (n=1 each). MutSig2CV analysis revealed recurrent somatic mutations in FAM72D (n=3, in 2 wiFTCs and in a single HCC), TP53 (n=3, in 2 wiFTCs and a single HCC), and EIF1AX (n=3), with DGCR8 (n=2) as borderline significant. The DGCR8 mutations were recurrent p.E518K missense alterations, known to cause familial multinodular goiter via disruption of microRNA (miRNA) processing. Expression analyses showed reduced DGCR8 messenger RNA expression in FTCs in general, and the 2 DGCR8 mutants displayed a distinct miRNA profile compared to DGCR8 wild-types. Copy number analyses revealed recurrent gains on chromosomes 4, 6, and 10, and fusion gene analyses revealed 27 high-quality events. Both HCCs displayed hyperploidy, which was fairly unusual in the FTC cohort. Based on the transcriptome data, tumors amassed in 2 principal clusters. Conclusion: We describe the genomic and transcriptomic landscape in wiFTCs and HCCs and identify novel recurrent mutations and copy number alterations with possible driver properties and lay the foundation for future studies.

Despite Africa's central role in the origin of our species, our knowledge of the genomic diversity in Africa is remarkably sparse. A recent publication by Choudhury et al. underscores the scientific imperative for a broader characterisation of African genomic diversity to better understand demographic history and improve global human health.

Despite Africa's central role in the origin of our species, our knowledge of the genomic diversity in Africa is remarkably sparse. A recent publication by Choudhury et al. underscores the scientific imperative for a broader characterisation of African genomic diversity to better understand demographic history and improve global human health.

Background: The genomic and transcriptomic landscape of widely invasive follicular thyroid carcinomas (wiFTCs) and Hurthle cell carcinoma (HCC) are poorly characterized, and subsets of these tumors lack information on genetic driver events. Objective: The aim of this study was to bridge this gap. Methods: We performed whole-genome and RNA sequencing and subsequent bioinformatic analyses of 11 wiFTCs and 2 HCCs with a particularly poor prognosis, and matched normal tissue. Results: All wiFTCs exhibited one or several mutations in established thyroid cancer genes, including TERT (n=4), NRAS (n=3), HRAS, KRAS, AKT, PTEN, PIK3CA, MUTYH, TSHR, and MEN1 (n=1 each). MutSig2CV analysis revealed recurrent somatic mutations in FAM72D (n=3, in 2 wiFTCs and in a single HCC), TP53 (n=3, in 2 wiFTCs and a single HCC), and EIF1AX (n=3), with DGCR8 (n=2) as borderline significant. The DGCR8 mutations were recurrent p.E518K missense alterations, known to cause familial multinodular goiter via disruption of microRNA (miRNA) processing. Expression analyses showed reduced DGCR8 messenger RNA expression in FTCs in general, and the 2 DGCR8 mutants displayed a distinct miRNA profile compared to DGCR8 wild-types. Copy number analyses revealed recurrent gains on chromosomes 4, 6, and 10, and fusion gene analyses revealed 27 high-quality events. Both HCCs displayed hyperploidy, which was fairly unusual in the FTC cohort. Based on the transcriptome data, tumors amassed in 2 principal clusters. Conclusion: We describe the genomic and transcriptomic landscape in wiFTCs and HCCs and identify novel recurrent mutations and copy number alterations with possible driver properties and lay the foundation for future studies.

Despite Africa's central role in the origin of our species, our knowledge of the genomic diversity in Africa is remarkably sparse. A recent publication by Choudhury et al. underscores the scientific imperative for a broader characterisation of African genomic diversity to better understand demographic history and improve global human health.

Three Streptomyces sp. strains with a multitude of target enzymatic activities confirmed by functional screening, namely BV129, BV286 and BV333, were subjected to genome sequencing aiming at the annotation of genes of interest, in-depth bioinformatics characterization and functional expression of the biocatalysts. A whole-genome shotgun sequencing followed by de novo genome assembly and annotation was performed revealing genomes of 6.4, 9.4 and 7.3 Mbp, respectively. Functional annotation of the proteins of interest resulted in between 2047 and 2763 putative targets. Among the various enzymatic activities that the three Streptomyces strains demonstrated to produce by functional screening, we focused our attention on transaminases (TAs) and laccases due to their high biocatalytic potential. Bioinformatics search allowed the identification of a putative TA from Streptomyces sp. BV333 as a potentially novel broad substrate scope TA and a putative laccase from Streptomyces sp. BV286 as potentially novel blue multicopper oxidase. The two sequences were cloned and overexpressed in Escherichia coli and the two novel enzymes, transaminase Sbv333-TA and laccase Sbv286-LAC, were characterized. Interestingly, both enzymes resulted to be exceptionally thermostable, Sbv333-TA showing a melting temperature (T-M = 85 degrees C) only slightly lower compared to the T-M of the most thermostable transaminases described to date (87-88 degrees C) and Sbv286-LAC being even thermoactivated at temperature gt 60 degrees C. Moreover, Sbv333-TA showed a broad substrate scope and remarkably demonstrated to be active in the transamination of beta-ketoesters, which are rarely accepted by currently known TAs. On the other hand, Sbv286-LAC showed an improved activity in the presence of the cosolvent acetonitrile. Overall, it was shown that a combination of approaches from standard microbiological and biochemical screens to genome sequencing and analysis is required to afford novel and functional b

Background: The genomic and transcriptomic landscape of widely invasive follicular thyroid carcinomas (wiFTCs) and Hurthle cell carcinoma (HCC) are poorly characterized, and subsets of these tumors lack information on genetic driver events. Objective: The aim of this study was to bridge this gap. Methods: We performed whole-genome and RNA sequencing and subsequent bioinformatic analyses of 11 wiFTCs and 2 HCCs with a particularly poor prognosis, and matched normal tissue. Results: All wiFTCs exhibited one or several mutations in established thyroid cancer genes, including TERT (n=4), NRAS (n=3), HRAS, KRAS, AKT, PTEN, PIK3CA, MUTYH, TSHR, and MEN1 (n=1 each). MutSig2CV analysis revealed recurrent somatic mutations in FAM72D (n=3, in 2 wiFTCs and in a single HCC), TP53 (n=3, in 2 wiFTCs and a single HCC), and EIF1AX (n=3), with DGCR8 (n=2) as borderline significant. The DGCR8 mutations were recurrent p.E518K missense alterations, known to cause familial multinodular goiter via disruption of microRNA (miRNA) processing. Expression analyses showed reduced DGCR8 messenger RNA expression in FTCs in general, and the 2 DGCR8 mutants displayed a distinct miRNA profile compared to DGCR8 wild-types. Copy number analyses revealed recurrent gains on chromosomes 4, 6, and 10, and fusion gene analyses revealed 27 high-quality events. Both HCCs displayed hyperploidy, which was fairly unusual in the FTC cohort. Based on the transcriptome data, tumors amassed in 2 principal clusters. Conclusion: We describe the genomic and transcriptomic landscape in wiFTCs and HCCs and identify novel recurrent mutations and copy number alterations with possible driver properties and lay the foundation for future studies.

Despite Africa's central role in the origin of our species, our knowledge of the genomic diversity in Africa is remarkably sparse. A recent publication by Choudhury et al. underscores the scientific imperative for a broader characterisation of African genomic diversity to better understand demographic history and improve global human health.

Cancers arise as a result of deleterious somatic alterations accumulating in the genome during the process of cell division. These alterations arise either via exposure to mutagens or due to errors occurring during DNA replication. In this thesis, a systematic exploration, from discovery to analyses of somatic alterations in three diverse cancers that affect dogs and humans, was undertaken. In Studies I and II, whole-exome sequencing of dogs affected by the cancers of osteosarcoma and hemangiosarcoma were done to delineate coding mutations that can contribute to their carcinogenesis. Besides, as these cancers mirror the corresponding human disease in clinical manifestation and histological features, a secondary objective was to confirm the molecular drivers found in the canines were also influencing factors in the human cancer(s). In the osteosarcoma investigations with three breeds, we found that tumors show a high frequency of somatic copy-number alterations, affecting key cancer genes. TP53 was the most frequently altered gene, akin to human osteosarcoma. The second most mutated gene, histone methyltransferase SETD2, has known epigenetic roles in multiple cancers but not in osteosarcoma. Our study highlights the strong genetic similarities between human and dog osteosarcoma, suggesting that canine disease may serve as an excellent model for developing treatment strategies in both species. In the hemangiosarcoma study in golden retrievers, putative driver alterations were identified in the tumor suppressor TP53 and in genes involved in the cell cycle regulating PI3K pathway, including PIK3CA and PIK3R1. Furthermore, we find several somatic alterations between the dog hemangiosarcoma and human angiosarcoma overlap, indicating we can use the canine model to apprise the infrequently occurring human disease. In Study III, we implemented whole-genome sequencing methodologies to define both coding and non-coding alterations in the glioblastoma cancer genome. We find the

A new Alcanivorax sp. VBW004 was isolated from a shallow hydrothermal vent in Azores Island, Portugal. In this study, we determined VBW004 was resistant to copper. This strain showed maximum tolerance of copper concentrations up to 600 μg/mL. Based on 16S rRNA gene sequencing and phylogeny revealed that this strain was more closely related to Alcanivorax borkumensis SK2. We sequenced the genome of this strain that consist of 3.8 Mb size with a G + C content of 58.4 %. In addition, digital DNA-DNA hybridizations (dDDH) and the average nucleotide identities (ANI) analysis between Alcanivorax borkumensis SK2 and Alcanivorax jadensis T9 revealed that Alcanivorax sp. VBW004 belongs to new species. Functional annotation revealed that the genome acquired multiple copper resistance encoding genes that could assist VBW004 to respond to high Cu toxicity. Our results from biosorption analysis presumed that the VBW004 is an ecologically important bacterium that could be useful for copper bioremediation.

Cancers arise as a result of deleterious somatic alterations accumulating in the genome during the process of cell division. These alterations arise either via exposure to mutagens or due to errors occurring during DNA replication. In this thesis, a systematic exploration, from discovery to analyses of somatic alterations in three diverse cancers that affect dogs and humans, was undertaken. In Studies I and II, whole-exome sequencing of dogs affected by the cancers of osteosarcoma and hemangiosarcoma were done to delineate coding mutations that can contribute to their carcinogenesis. Besides, as these cancers mirror the corresponding human disease in clinical manifestation and histological features, a secondary objective was to confirm the molecular drivers found in the canines were also influencing factors in the human cancer(s). In the osteosarcoma investigations with three breeds, we found that tumors show a high frequency of somatic copy-number alterations, affecting key cancer genes. TP53 was the most frequently altered gene, akin to human osteosarcoma. The second most mutated gene, histone methyltransferase SETD2, has known epigenetic roles in multiple cancers but not in osteosarcoma. Our study highlights the strong genetic similarities between human and dog osteosarcoma, suggesting that canine disease may serve as an excellent model for developing treatment strategies in both species. In the hemangiosarcoma study in golden retrievers, putative driver alterations were identified in the tumor suppressor TP53 and in genes involved in the cell cycle regulating PI3K pathway, including PIK3CA and PIK3R1. Furthermore, we find several somatic alterations between the dog hemangiosarcoma and human angiosarcoma overlap, indicating we can use the canine model to apprise the infrequently occurring human disease. In Study III, we implemented whole-genome sequencing methodologies to define both coding and non-coding alterations in the glioblastoma cancer genome. We find the

Cancers arise as a result of deleterious somatic alterations accumulating in the genome during the process of cell division. These alterations arise either via exposure to mutagens or due to errors occurring during DNA replication. In this thesis, a systematic exploration, from discovery to analyses of somatic alterations in three diverse cancers that affect dogs and humans, was undertaken. In Studies I and II, whole-exome sequencing of dogs affected by the cancers of osteosarcoma and hemangiosarcoma were done to delineate coding mutations that can contribute to their carcinogenesis. Besides, as these cancers mirror the corresponding human disease in clinical manifestation and histological features, a secondary objective was to confirm the molecular drivers found in the canines were also influencing factors in the human cancer(s). In the osteosarcoma investigations with three breeds, we found that tumors show a high frequency of somatic copy-number alterations, affecting key cancer genes. TP53 was the most frequently altered gene, akin to human osteosarcoma. The second most mutated gene, histone methyltransferase SETD2, has known epigenetic roles in multiple cancers but not in osteosarcoma. Our study highlights the strong genetic similarities between human and dog osteosarcoma, suggesting that canine disease may serve as an excellent model for developing treatment strategies in both species. In the hemangiosarcoma study in golden retrievers, putative driver alterations were identified in the tumor suppressor TP53 and in genes involved in the cell cycle regulating PI3K pathway, including PIK3CA and PIK3R1. Furthermore, we find several somatic alterations between the dog hemangiosarcoma and human angiosarcoma overlap, indicating we can use the canine model to apprise the infrequently occurring human disease. In Study III, we implemented whole-genome sequencing methodologies to define both coding and non-coding alterations in the glioblastoma cancer genome. We find the

Cancers arise as a result of deleterious somatic alterations accumulating in the genome during the process of cell division. These alterations arise either via exposure to mutagens or due to errors occurring during DNA replication. In this thesis, a systematic exploration, from discovery to analyses of somatic alterations in three diverse cancers that affect dogs and humans, was undertaken. In Studies I and II, whole-exome sequencing of dogs affected by the cancers of osteosarcoma and hemangiosarcoma were done to delineate coding mutations that can contribute to their carcinogenesis. Besides, as these cancers mirror the corresponding human disease in clinical manifestation and histological features, a secondary objective was to confirm the molecular drivers found in the canines were also influencing factors in the human cancer(s). In the osteosarcoma investigations with three breeds, we found that tumors show a high frequency of somatic copy-number alterations, affecting key cancer genes. TP53 was the most frequently altered gene, akin to human osteosarcoma. The second most mutated gene, histone methyltransferase SETD2, has known epigenetic roles in multiple cancers but not in osteosarcoma. Our study highlights the strong genetic similarities between human and dog osteosarcoma, suggesting that canine disease may serve as an excellent model for developing treatment strategies in both species. In the hemangiosarcoma study in golden retrievers, putative driver alterations were identified in the tumor suppressor TP53 and in genes involved in the cell cycle regulating PI3K pathway, including PIK3CA and PIK3R1. Furthermore, we find several somatic alterations between the dog hemangiosarcoma and human angiosarcoma overlap, indicating we can use the canine model to apprise the infrequently occurring human disease. In Study III, we implemented whole-genome sequencing methodologies to define both coding and non-coding alterations in the glioblastoma cancer genome. We find the

Cancers arise as a result of deleterious somatic alterations accumulating in the genome during the process of cell division. These alterations arise either via exposure to mutagens or due to errors occurring during DNA replication. In this thesis, a systematic exploration, from discovery to analyses of somatic alterations in three diverse cancers that affect dogs and humans, was undertaken. In Studies I and II, whole-exome sequencing of dogs affected by the cancers of osteosarcoma and hemangiosarcoma were done to delineate coding mutations that can contribute to their carcinogenesis. Besides, as these cancers mirror the corresponding human disease in clinical manifestation and histological features, a secondary objective was to confirm the molecular drivers found in the canines were also influencing factors in the human cancer(s). In the osteosarcoma investigations with three breeds, we found that tumors show a high frequency of somatic copy-number alterations, affecting key cancer genes. TP53 was the most frequently altered gene, akin to human osteosarcoma. The second most mutated gene, histone methyltransferase SETD2, has known epigenetic roles in multiple cancers but not in osteosarcoma. Our study highlights the strong genetic similarities between human and dog osteosarcoma, suggesting that canine disease may serve as an excellent model for developing treatment strategies in both species. In the hemangiosarcoma study in golden retrievers, putative driver alterations were identified in the tumor suppressor TP53 and in genes involved in the cell cycle regulating PI3K pathway, including PIK3CA and PIK3R1. Furthermore, we find several somatic alterations between the dog hemangiosarcoma and human angiosarcoma overlap, indicating we can use the canine model to apprise the infrequently occurring human disease. In Study III, we implemented whole-genome sequencing methodologies to define both coding and non-coding alterations in the glioblastoma cancer genome. We find the

There is still much uncertainty about the underlying mechanisms that govern antigenic drift of influenza viruses, and several theories have been proposed. These theories consider hemagglutinin (HA) to be the primary driving force, while the second key surface glycoprotein (neuraminidase, NA) and the other viral proteins have largely been ignored. This thesis focuses on all influenza A (H3N2) virus proteins, with special emphasis on NA, to better understand—and ultimately predict—the complex evolution of A(H3N2) viruses. The genetic evolution, with respect to antigenic change, was analyzed by comparing HA and NA (chapter 2) and by studying the whole-genome (chapter 3). We next optimized the enzyme-linked lectin assay (ELLA) as an NA inhibition (NI) assay for human serology (chapter 5) and rapid antigenic characterization (chapter 6). With the optimized NI ELLA, we analyzed NA from 1968 till the 2010-2011 season and subsequently compared it to HA (chapter 7). Our work clearly shows that there is antigenic evolution for NA, thus warranting the inclusion of NAs representing emerging influenza A strains in vaccines. Increasing knowledge on which mutations cause changes in the phenotype of NA can help to perform more targeted influenza surveillance. It would then be advisable to integrate genetic and antigenic NA data with sequence and antigenic data of HA, epidemiological data, and geographical data during influenza surveillance. This will facilitate consideration of NA content, and improve next generation influenza vaccines.

The deubiquitinating enzyme BAP1 is a tumor suppressor, among others involved in cholangiocarcinoma. BAP1 has many proposed molecular targets, while its Drosophila homolog is known to deubiquitinate histone H2AK119. We introduce BAP1 loss-of-function by CRISPR/Cas9 in normal human cholangiocyte organoids. We find that BAP1 controls the expression of junctional and cytoskeleton components by regulating chromatin accessibility. Consequently, we observe loss of multiple epithelial characteristics while motility increases. Importantly, restoring the catalytic activity of BAP1 in the nucleus rescues these cellular and molecular changes. We engineer human liver organoids to combine four common cholangiocarcinoma mutations (TP53, PTEN, SMAD4, and NF1). In this genetic background, BAP1 loss results in acquisition of malignant features upon xenotransplantation. Thus, control of epithelial identity through the regulation of chromatin accessibility appears to be a key aspect of BAP1's tumor suppressor function. Organoid technology combined with CRISPR/Cas9 provides an experimental platform for mechanistic studies of cancer gene function in a human context.

The deubiquitinating enzyme BAP1 is a tumor suppressor, among others involved in cholangiocarcinoma. BAP1 has many proposed molecular targets, while its Drosophila homolog is known to deubiquitinate histone H2AK119. We introduce BAP1 loss-of-function by CRISPR/Cas9 in normal human cholangiocyte organoids. We find that BAP1 controls the expression of junctional and cytoskeleton components by regulating chromatin accessibility. Consequently, we observe loss of multiple epithelial characteristics while motility increases. Importantly, restoring the catalytic activity of BAP1 in the nucleus rescues these cellular and molecular changes. We engineer human liver organoids to combine four common cholangiocarcinoma mutations (TP53, PTEN, SMAD4, and NF1). In this genetic background, BAP1 loss results in acquisition of malignant features upon xenotransplantation. Thus, control of epithelial identity through the regulation of chromatin accessibility appears to be a key aspect of BAP1's tumor suppressor function. Organoid technology combined with CRISPR/Cas9 provides an experimental platform for mechanistic studies of cancer gene function in a human context.

An important component of plant evolution is the plethora of pathways producing more than 200,000 biochemically diverse specialized metabolites with pharmacological, nutritional and ecological significance. To unravel dynamics underlying metabolic diversification, it is critical to determine lineage-specific gene family expansion in a phylogenomics framework. However, robust functional annotation is often only available for core enzymes catalyzing committed reaction steps within few model systems. In a genome informatics approach, we extracted information from early-draft gene-space assemblies and non-redundant transcriptomes to identify protein families involved in isoprenoid biosynthesis. Isoprenoids comprise terpenoids with various roles in plant-environment interaction, such as pollinator attraction or pathogen defense. Combining lines of evidence provided by synteny, sequence homology and Hidden-Markov-Modelling, we screened 17 genomes including 12 major crops and found evidence for 1,904 proteins associated with terpenoid biosynthesis. Our terpenoid genes set contains evidence for 840 core terpene-synthases and 338 triterpene-specific synthases. We further identified 190 prenyltransferases, 39 isopentenyl-diphosphate isomerases as well as 278 and 219 proteins involved in mevalonate and methylerithrol pathways, respectively. Assessing the impact of gene and genome duplication to lineage-specific terpenoid pathway expansion, we illustrated key events underlying terpenoid metabolic diversification within 250 million years of flowering plant radiation. By quantifying Angiosperm-wide versatility and phylogenetic relationships of pleiotropic gene families in terpenoid modular pathways, our analysis offers significant insight into evolutionary dynamics underlying diversification of plant secondary metabolism. Furthermore, our data provide a blueprint for future efforts to identify and more rapidly clone terpenoid biosynthetic genes from any plant species.

Background: There is tremendous potential for genome sequencing to improve clinical diagnosis and care once it becomes routinely accessible, but this will require formalizing research methods into clinical best practices in the areas of sequence data generation, analysis, interpretation and reporting. The CLARITY Challenge was designed to spur convergence in methods for diagnosing genetic disease starting from clinical case history and genome sequencing data. DNA samples were obtained from three families with heritable genetic disorders and genomic sequence data were donated by sequencing platform vendors. The challenge was to analyze and interpret these data with the goals of identifying disease-causing variants and reporting the findings in a clinically useful format. Participating contestant groups were solicited broadly, and an independent panel of judges evaluated their performance. Results: A total of 30 international groups were engaged. The entries reveal a general convergence of practices on most elements of the analysis and interpretation process. However, even given this commonality of approach, only two groups identified the consensus candidate variants in all disease cases, demonstrating a need for consistent fine-tuning of the generally accepted methods. There was greater diversity of the final clinical report content and in the patient consenting process, demonstrating that these areas require additional exploration and standardization. Conclusions: The CLARITY Challenge provides a comprehensive assessment of current practices for using genome sequencing to diagnose and report genetic diseases. There is remarkable convergence in bioinformatic techniques, but medical interpretation and reporting are areas that require further development by many groups., QC 20140819

Metastasis accounts for the poor prognosis of the majority of solid tumors. The phenotypic transition of nonmotile epithelial tumor cells to migratory and invasive “mesenchymal” cells (epithelial-to-mesenchymal transition [EMT]) enables the transit of cancer cells from the primary tumor to distant sites. There is no single marker of EMT; rather, multiple measures are required to define cell state. Thus, the multiparametric capability of high-content screening is ideally suited for the comprehensive analysis of EMT regulators. The aim of this study was to generate a platform to systematically identify functional modulators of tumor cell plasticity using the bladder cancer cell line TSU-Pr1-B1 as a model system. A platform enabling the quantification of key EMT characteristics, cell morphology and mesenchymal intermediate filament vimentin, was developed using the fluorescent whole-cell-tracking reagent CMFDA and a fluorescent promoter reporter construct, respectively. The functional effect of genome-wide modulation of protein-coding genes and miRNAs coupled with those of a collection of small-molecule kinase inhibitors on EMT was assessed using the Target Activation Bioapplication integrated in the Cellomics ArrayScan platform. Data from each of the three screens were integrated to identify a cohort of targets that were subsequently examined in a validation assay using siRNA duplexes. Identification of established regulators of EMT supports the utility of this screening approach and indicated capacity to identify novel regulators of this plasticity program. Pathway analysis coupled with interrogation of cancer-related expression profile databases and other EMT-related screens provided key evidence to prioritize further experimental investigation into the molecular regulators of EMT in cancer cells.

Background Recent advances in DNA sequencing techniques resulted in more than forty sequenced plant genomes representing a diverse set of taxa of agricultural, energy, medicinal and ecological importance. However, gene family curation is often only inferred from DNA sequence homology and lacks insights into evolutionary processes contributing to gene family dynamics. In a comparative genomics framework, we integrated multiple lines of evidence provided by gene synteny, sequence homology and protein-based Hidden Markov Modelling to extract homologous super-clusters composed of multi-domain resistance (R)-proteins of the NB-LRR type (for NUCLEOTIDE BINDING/LEUCINE-RICH REPEATS), that are involved in plant innate immunity. Results To assess the diversity of R-proteins within and between species, we screened twelve eudicot plant genomes including six major crops and found a total of 2,363 NB-LRR genes. Our curated R-proteins set shows a 50% average for tandem duplicates and a 22% fraction of gene copies retained from ancient polyploidy events (ohnologs). We provide evidence for strong positive selection acting on all identified genes and show significant differences in molecular evolution rates (Ka/Ks-ratio) among tandem- (mean = 1.59), ohnolog (mean = 1.36) and singleton (mean = 1.22) R-gene duplicates. To foster the process of gene-edited plant breeding, we report species-specific presence/absence of all 140 NB-LRR genes present in the model plant Arabidopsis and describe four distinct clusters of NB-LRR "gatekeeper" loci sharing syntenic orthologs across all analyzed genomes. Conclusion By curating a near-complete set of multi-domain R-protein clusters in an eudicot-wide scale, our analysis offers significant insight into evolutionary dynamics underlying diversification of the plant innate immune system. Furthermore, our methods provide a blueprint for future efforts to identify and more rapidly clone functional NB-LRR genes from any plant species.

Background: Chinese bayberry (Myrica rubra Sieb. and Zucc.) is a subtropical evergreen tree originating in China. It has been cultivated in southern China for several thousand years, and annual production has reached 1.1 million tons. The taste and high level of health promoting characters identified in the fruit in recent years has stimulated its extension in China and introduction to Australia. A limited number of co-dominant markers have been developed and applied in genetic diversity and identity studies. Here we report, for the first time, a survey of whole genome shotgun data to develop a large number of simple sequence repeat (SSR) markers to analyse the genetic diversity of the common cultivated Chinese bayberry and the relationship with three other Myrica species. Results: The whole genome shotgun survey of Chinese bayberry produced 9.01Gb of sequence data, about 26x coverage of the estimated genome size of 323 Mb. The genome sequences were highly heterozygous, but with little duplication. From the initial assembled scaffold covering 255 Mb sequence data, 28,602 SSRs (>= 5 repeats) were identified. Dinucleotide was the most common repeat motif with a frequency of 84.73%, followed by 13.78% trinucleotide, 1.34% tetranucleotide, 0.12% pentanucleotide and 0.04% hexanucleotide. From 600 primer pairs, 186 polymorphic SSRs were developed. Of these, 158 were used to screen 29 Chinese bayberry accessions and three other Myrica species: 91.14%, 89.87% and 46.84% SSRs could be used in Myrica adenophora, Myrica nana and Myrica cerifera, respectively. The UPGMA dendrogram tree showed that cultivated Myrica rubra is closely related to Myrica adenophora and Myrica nana, originating in southwest China, and very distantly related to Myrica cerifera, originating in America. These markers can be used in the construction of a linkage map and for genetic diversity studies in Myrica species. Conclusion: Myrica rubra has a small genome of about 323 Mb with a high level of hetero