Zebrafish regulatory genomic resources for disease modelling and regeneration

In the past decades, the zebrafish has become a disease model with increasing popularity owing to its advantages that include fast development, easy genetic manipulation, simplicity for imaging, and sharing conserved disease-associated genes and pathways with those of human. In parallel, studies of disease mechanisms are increasingly focusing on non-coding mutations, which require genome annotation maps of regulatory elements, such as enhancers and promoters. In line with this, genomic resources for zebrafish research are expanding, producing a variety of genomic data that help in defining regulatory elements and their conservation between zebrafish and humans. Here, we discuss recent developments in generating functional annotation maps for regulatory elements of the zebrafish genome and how this can be applied to human diseases. We highlight community-driven developments, such as DANIO-CODE, in generating a centralised and standardised catalogue of zebrafish genomics data and functional annotations; consider the advantages and limitations of current annotation maps; and offer considerations for interpreting and integrating existing maps with comparative genomics tools. We also discuss the need for developing standardised genomics protocols and bioinformatic pipelines and provide suggestions for the development of analysis and visualisation tools that will integrate various multiomic bulk sequencing data together with fast-expanding data on single-cell methods, such as single-cell assay for transposase-accessible chromatin with sequencing. Such integration tools are essential to exploit the multiomic chromatin characterisation offered by bulk genomics together with the cell-type resolution offered by emerging single-cell methods. Together, these advances will build an expansive toolkit for interrogating the mechanisms of human disease in zebrafish

Bioprospecting for Genes Encoding Hydrocarbon-Degrading Enzymes from Metagenomic Samples Isolated from Northern Adriatic Sea Sediments

Three metagenomic libraries were constructed using surface sediment samples from the northern Adriatic Sea. Two of the samples were taken from a highly polluted and an unpolluted site respectively. The third sample from a polluted site had been enriched using crude oil. The results of the metagenome analyses were incorporated in the REDPET relational database (http://redpet.bioinfo.pbf.hr/REDPET), which was generated using the previously developed MEGGASENSE platform. The database includes taxonomic data to allow the assessment of the biodiversity of metagenomic libraries and a general functional analysis of genes using hidden Markov model (HMM) profiles based on the KEGG database. A set of 22 specialised HMM-profiles was developed to detect putative genes for hydrocarbon-degrading enzymes. Use of these profiles showed that the metagenomic library generated after selection on crude oil had enriched genes for aerobic n-alkane degradation. The use of this system for bioprospecting was exemplified using potential alkB and almA genes from this library

Novi pristup konstrukciji industrijskih mikroorganizama pomoću sintetičke biologije

The recent achievement of synthesising a functioning bacterial chromosome marks a coming of age for engineering living organisms. In the future this should allow the construction of novel organisms to help solve the problems facing the human race, including health care, food, energy and environmental protection. In this minireview, the current state of the field is described and the role of synthetic biology in biotechnology in the short and medium term is discussed. It is particularly aimed at the needs of food technologists, nutritionists and other biotechnologists, who might not be aware of the potential significance of synthetic biology to the research and development in their fields. The potential of synthetic biology to produce interesting new polyketide compounds is discussed in detail.Razvojem područja sinteze funkcionalnog bakterijskog kromosoma obilježen je početak novog doba genetičkog inženjerstva. Konstrukcijom novih organizama mogli bi se riješiti neki problemi vezani uz zdravstvo, proizvodnju hrane i energenata te zaštitu okoliša. U ovom su kratkom revijalnom prikazu opisana sadašnja dostignuća na području sintetičke biologije, a raspravlja se i o njezinoj ulozi u razvoju biotehnologije. Prikaz je posebno namijenjen prehrambenim tehnolozima, nutricionistima i ostalim biotehnolozima koji možda nisu svjesni značaja što bi sintetička biologija mogla imati za njihova istraživanja. Detaljno se raspravlja o mogućem utjecaju sintetičke biologije u formiranju potpuno novih poliketida, koji se mogu upotrijebiti za proizvodnju lijekova

Integrative genomic analyses in adipocytes implicate DNA methylation in human obesity and diabetes

DNA methylation variations are prevalent in human obesity but evidence of a causative role in disease pathogenesis is limited. Here, we combine epigenome-wide association and integrative genomics to investigate the impact of adipocyte DNA methylation variations in human obesity. We discover extensive DNA methylation changes that are robustly associated with obesity (N = 190 samples, 691 loci in subcutaneous and 173 loci in visceral adipocytes, P 500 target genes, and identify putative methylation-transcription factor interactions. Through Mendelian Randomisation, we infer causal effects of methylation on obesity and obesity-induced metabolic disturbances at 59 independent loci. Targeted methylation sequencing, CRISPR-activation and gene silencing in adipocytes, further identifies regional methylation variations, underlying regulatory elements and novel cellular metabolic effects. Our results indicate DNA methylation is an important determinant of human obesity and its metabolic complications, and reveal mechanisms through which altered methylation may impact adipocyte functions

Reference-Grade Genome and Large Linear Plasmid of Streptomyces rimosus : Pushing the Limits of Nanopore Sequencing

Streptomyces rimosus ATCC 10970 is the parental strain of industrial strains used for the commercial production of the important antibiotic oxytetracycline. As an actinobacterium with a large linear chromosome containing numerous long repeat regions, high GC content, and a single giant linear plasmid (GLP), these genomes are challenging to assemble. Here, we apply a hybrid sequencing approach relying on the combination of short- and long-read next-generation sequencing platforms and whole-genome restriction analysis by using pulsed-field gel electrophoresis (PFGE) to produce a high-quality reference genome for this biotechnologically important bacterium. By using PFGE to separate and isolate plasmid DNA from chromosomal DNA, we successfully sequenced the GLP using Nanopore data alone. Using this approach, we compared the sequence of GLP in the parent strain ATCC 10970 with those found in two semi-industrial progenitor strains, R6-500 and M4018. Sequencing of the GLP of these three S. rimosus strains shed light on several rearrangements accompanied by transposase genes, suggesting that transposases play an important role in plasmid and genome plasticity in S. rimosus. The polished annotation of secondary metabolite biosynthetic pathways compared to metabolite analysis in the ATCC 10970 strain also refined our knowledge of the secondary metabolite arsenal of these strains. The proposed methodology is highly applicable to a variety of sequencing projects, as evidenced by the reliable assemblies obtained. IMPORTANCE The genomes of Streptomyces species are difficult to assemble due to long repeats, extrachromosomal elements (giant linear plasmids [GLPs]), rearrangements, and high GC content. To improve the quality of the S. rimosus ATCC 10970 genome, producer of oxytetracycline, we validated the assembly of GLPs by applying a new approach to combine pulsed-field gel electrophoresis separation and GLP isolation and sequenced the isolated GLP with Oxford Nanopore technology. By examining the sequenced plasmids of ATCC 10970 and two industrial progenitor strains, R6-500 and M4018, we identified large GLP rearrangements. Analysis of the assembled plasmid sequences shed light on the role of transposases in genome plasticity of this species. The new methodological approach developed for Nanopore sequencing is highly applicable to a variety of sequencing projects. In addition, we present the annotated reference genome sequence of ATCC 10970 with a detailed analysis of the biosynthetic gene clusters

Integrative genomic analyses in adipocytes implicate DNA methylation in human obesity and diabetes

DNA methylation variations are prevalent in human obesity but evidence of a causative role in disease pathogenesis is limited. Here, we combine epigenome-wide association and integrative genomics to investigate the impact of adipocyte DNA methylation variations in human obesity. We discover extensive DNA methylation changes that are robustly associated with obesity (N = 190 samples, 691 loci in subcutaneous and 173 loci in visceral adipocytes, P 500 target genes, and identify putative methylation-transcription factor interactions. Through Mendelian Randomisation, we infer causal effects of methylation on obesity and obesity-induced metabolic disturbances at 59 independent loci. Targeted methylation sequencing, CRISPR-activation and gene silencing in adipocytes, further identifies regional methylation variations, underlying regulatory elements and novel cellular metabolic effects. Our results indicate DNA methylation is an important determinant of human obesity and its metabolic complications, and reveal mechanisms through which altered methylation may impact adipocyte functions

Genome sequences of the oxytetracycline production strain Streptomyces rimosus R6-500 and two mutants with chromosomal rearrangements

The genome sequence of Streptomyces rimosus R6-500, an industrially improved strain which produces high titers of the important antibiotic oxytetracycline, is reported, as well as the genome sequences of two derivatives arising due to the genetic instability of the strain

Evolutionary concepts in natural products discovery:what actinomycetes have taught us

Abstract Actinomycetes are a very important source of natural products for the pharmaceutical industry and other applications. Most of the strains belong to Streptomyces or related genera, partly because they are particularly amenable to growth in the laboratory and industrial fermenters. It is unlikely that chemical synthesis can fulfil the needs of the pharmaceutical industry for novel compounds so there is a continuing need to find novel natural products. An evolutionary perspective can help this process in several ways. Genome mining attempts to identify secondary metabolite biosynthetic clusters in DNA sequences, which are likely to produce interesting chemical entities. There are often technical problems in assembling the DNA sequences of large modular clusters in genome and metagenome projects, which can be overcome partially using information about the evolution of the domain sequences. Understanding the evolutionary mechanisms of modular clusters should allow simulation of evolutionary pathways in the laboratory to generate novel compounds.</jats:p

Synthetic biology: A novel approach for the construction of industrial microorganisms

The recent achievement of synthesising a functioning bacterial chromosome marks a coming of age for engineering living organisms. In the future this should allow the construction of novel organisms to help solve the problems facing the human race, including health care, food, energy and environmental protection. In this minireview, the current state of the field is described and the role of synthetic biology in biotechnology in the short and medium term is discussed. It is particularly aimed at the needs of food technologists, nutritionists and other biotechnologists, who might not be aware of the potential significance of synthetic biology to the research and development in their fields. The potential of synthetic biology to produce interesting new polyketide compounds is discussed in detail.Croatian Science Foundation grant 09/5German Academic Exchange Service (DAAD)CNPq-TWAS (Conselho Nacional de Pesquisa do Brazil Third World Academy of Sciences, Brazil)FAPESP (Fundação de Apoio a Pesquisa do Estado de São Paulo, Brazil)FAPESP-King’s Colleg