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Introduction Per- and polyfluoroalkyl substances (PFAS) are a large group of fluorine-containing chemicals commonly applied in industrial and commercial products, resulting in ubiquitous presence in the environment (1,2) and attracting [...], Background: Perfluorohexane sulfonate (PFHxS) is a frequently detected per- and polyfluoroalkyl substance in most populations, including in individuals who are pregnant, a period critical for early life development. Despite epidemiological evidence of exposure, developmental toxicity, particularly at realistic human exposures, remains understudied. Objectives: We evaluated the effect of gestational exposure to human-relevant body burden of PFHxS on fetal and placental development and explored mechanisms of action combining alternative splicing (AS) and gene expression (GE) analyses. Methods: Pregnant ICR mice were exposed to 0, 0.03, and 0.3 [micro]g/kg/day from gestational day 7 to day 17 via oral gavage. Upon euthanasia, PFHxS distribution was measured using liquid chromatography-tandem mass spectrometry. Maternal and fetal phenotypes were recorded, and histopathology was examined for placenta impairment. Multiomics was adopted by combining AS and GE analyses to unveil disruptions in mRNA quality and quantity. The key metabolite transporters were validated by quantitative real-time PCR (qRT-PCR) for quantification and three-dimensional (3D) structural simulation by AlphaFold2. Targeted metabolomics based on liquid chromatography-tandem mass spectrometry was used to detect amino acid and amides levels in the placenta. Results: Pups developmentally exposed to PFHxS exhibited signs of intrauterine growth restriction (IUGR), characterized by smaller fetal weight and body length (p Discussion: The findings from this study suggest that exposure to human-relevant very-low-dose PFHxS during pregnancy in mice caused IUGR, likely via downregulating of placental amino acid transporters, thereby impairing placental amino acid transportation, resulting in impairment of placental development. Our findings confirm epidemiological findings and call for future attention on the health risk of this persistent yet ubiquitous chemical in the early developmental stage and provide a new approach for understanding gene expression from both quantitative and qualitative omics approaches in toxicological studies. https://doi.org/10.1289/EHP13217

Author(s): Matthias Längin [sup.1] [sup.2] , Tanja Mayr [sup.1] [sup.2] , Bruno Reichart [sup.2] , Sebastian Michel [sup.3] , Stefan Buchholz [sup.3] , Sonja Guethoff [sup.2] [sup.3] , Alexey Dashkevich [...], Heart transplantation is the only cure for patients with terminal cardiac failure, but the supply of allogeneic donor organs falls far short of the clinical need.sup.1-3. Xenotransplantation of genetically modified pig hearts has been discussed as a potential alternative.sup.4. Genetically multi-modified pig hearts that lack galactose-[alpha]1,3-galactose epitopes ([alpha]1,3-galactosyltransferase knockout) and express a human membrane cofactor protein (CD46) and human thrombomodulin have survived for up to 945 days after heterotopic abdominal transplantation in baboons.sup.5. This model demonstrated long-term acceptance of discordant xenografts with safe immunosuppression but did not predict their life-supporting function. Despite 25 years of extensive research, the maximum survival of a baboon after heart replacement with a porcine xenograft was only 57 days and this was achieved, to our knowledge, only once.sup.6. Here we show that [alpha]1,3-galactosyltransferase-knockout pig hearts that express human CD46 and thrombomodulin require non-ischaemic preservation with continuous perfusion and control of post-transplantation growth to ensure long-term orthotopic function of the xenograft in baboons, the most stringent preclinical xenotransplantation model. Consistent life-supporting function of xenografted hearts for up to 195 days is a milestone on the way to clinical cardiac xenotransplantation.sup.7. [alpha]1,3-galactosyltransferase-knockout pig hearts that express human CD46 and human thrombomodulin require non-ischaemic preservation with continuous perfusion and post-transplantation growth control to ensure long-term orthotopic function of the xenograft in baboons.

To determine whether MECP2 duplication syndrome is reversible, we generated a conditional MECP2 overexpression mouse model that carries two functional alleles with species-matched endogenous control elements: a human wild-type MECP2 [...], Copy number variations have been frequently associated with developmental delay, intellectual disability and autism spectrum disorders (1). MECP2 duplication syndrome is one of the most common genomic rearrangements in males (2) and is characterized by autism, intellectual disability, motor dysfunction, anxiety, epilepsy, recurrent respiratory tract infections and early death (3-5). The broad range of deficits caused by methyl-CpG-binding protein 2 (MeCP2) overexpression poses a daunting challenge to traditional biochemical-pathway-based therapeutic approaches. Accordingly, we sought strategies that directly target MeCP2 and are amenable to translation into clinical therapy. The first question that we addressed was whether the neurological dysfunction is reversible after symptoms set in. Reversal of phenotypes in adult symptomatic mice has been demonstrated in some models of monogenic loss-of-function neurological disorders (6-8), including loss of MeCP2 in Rett syndrome (9), indicating that, at least in some cases, the neuroanatomy may remain sufficiently intact so that correction of the molecular dysfunction underlying these disorders can restore healthy physiology. Given the absence of neurodegeneration in MECP2 duplication syndrome, we propose that restoration of normal MeCP2 levels in MECP2 duplication adult mice would rescue their phenotype. By generating and characterizing a conditional Mecp2-overexpressing mouse model, here we show that correction of MeCP2 levels largely reverses the behavioural, molecular and electrophysiological deficits. We also reduced MeCP2 using an antisense oligonucleotide strategy, which has greater translational potential. Antisense oligonucleotides are small, modified nucleic acids that can selectively hybridize with messenger RNA transcribed from a target gene and silence it (10,11), and have been successfully used to correct deficits in different mouse models (12-18). We find that antisense oligonucleotide treatment induces a broad phenotypic rescue in adult symptomatic transgenic MECP2 duplication mice (MECP2-TG) (19,20), and corrected MECP2 levels in lymphoblastoid cells from MECP2 duplication patients in a dose-dependent manner.

Author(s): Andrew Scott CRISPR-Cas gene editing, once considered arcane, is fast entering mainstream use in research. Most people with an interest in science have probably heard about the technique, which [...]

This essay describes possible futures that may result from accelerating technological advances and the challenges these futures present to psychotherapists. In the next 100 years, human beings will be likely to increasingly use computers and artificial intelligence and become extremely dependent on this relationship. Chip and stem cell implants may provide people with greater memory capacity, computational capacity, and skill sets. Genetic engineering, cryonics, and cloning may allow dramatic increases in the human life span. If these developments occur, they will challenge what it means to be a human being and have a soul. Although psychotherapists will acquire new tools for assessment and intervention, they will face the daunting task of addressing these changes and challenges to personhood with their clients., TELETHERAPY AND COMPUTERIZED THERAPY Predicting the future of science and society is notoriously difficult (Orrell 2007). Although psychotherapists have written about the future of psychotherapy, much of their writing addresses [...]

Cardiac muscle restitution, or true regeneration, is an unmet need in the treatment of myocardial infarction (MI), prompting a decade of study with stem cells of many kinds. Among key [...]

Concerns about energy security, the global petroleum supply and climate change have increased interest in the production of sustainable and renewable liquid transport fuels (1,2). The first-generation biofuels, ethanol and [...], Advanced biofuels produced by microorganisms have similar properties to petroleum-based fuels, and can 'drop in' to the existing transportation infrastructure. However, producing these biofuels in yields high enough to be useful requires the engineering of the microorganism's metabolism. Such engineering is not based on just one specific feedstock or host organism. Data-driven and synthetic-biology approaches can be used to optimize both the host and pathways to maximize fuel production. Despite some success, challenges still need to be met to move advanced biofuels towards commercialization, and to compete with more conventional fuels.

It is estimated that greater than 90% of human genes undergo alternative splicing of pre-mRNA (1,2) and aberrant splicing has been implicated in a number of human diseases (3). Alternative [...], Alternative splicing of pre-messenger RNA is a key feature of transcriptome expansion in eukaryotic cells, yet its regulation is poorly understood. Spliceosome assembly occurs co-transcriptionally, raising the possibility that DNA structure may directly influence alternative splicing. Supporting such an association, recent reports have identified distinct histone methylation patterns, elevated nucleosome occupancy and enriched DNA methylation at exons relative to introns. Moreover, the rate of transcription elongation has been linked to alternative splicing. Here we provide the first evidence that a DNA-binding protein, CCCTC-binding factor (CTCF), can promote inclusion of weak upstream exons by mediating local RNA polymerase II pausing both in a mammalian model system for alternative splicing, CD45, and genome-wide. We further show that CTCF binding to CD45 exon 5 is inhibited by DNA methylation, leading to reciprocal effects on exon 5 inclusion. These findings provide a mechanistic basis for developmental regulation of splicing outcome through heritable epigenetic marks.

Anyone familiar with Rett syndrome will recognize the symptoms. Mouse models--animals that carry genetic mutations similar to those that cause the condition in humans--wring their paws, walk awkwardly and learn [...]

Microalgae are considered one of the most promising feedstocks for biofuels. The productivity of these photosynthetic microorganisms in converting carbon dioxide into carbon-rich lipids, only a step or two away from biodiesel, greatly exceeds that of agricultural oleaginous crops, without competing for arable land. Worldwide, research and demonstration programs are being carded out to develop the technology needed to expand algal lipid production from a craft to a major industrial process. Although microalgae are not yet produced at large scale for bulk applications, recent advances--particularly in the methods of systems biology, genetic engineering, and biorefining--present opportunities to develop this process in a sustainable and economical way within the next 10 to 15 years. 10.1126/science.1189003

A historical overview of the evolution of rice technology in China, from traditional agriculture to usage of genetic modification, is presented. Rice technology is deemed as a complex interaction of social and material elements. Implications of wholesale transport of agricultural technology and social mechanism to developing countries and of indigenous innovation are described.

Abstract: The use of genetic engineering and biotechnology in agriculture has attracted worldwide attention over the past decade. This technology has raised highly controversial issues and considerable international debate over [...]

On a chilly late autumnal day in November 2008, Aron Geurts marched into a windowless basement animal facility to check up on his rats. There, he found five newborns in [...]

Synchronized clocks are of fundamental importance in the coordination of rhythmic behaviour among individual elements in a community or a large complex system. In physics and engineering, the Huygens paradigm [...], The engineering of genetic circuits with predictive functionality in living cells represents a defining focus of the expanding field of synthetic biology. This focus was elegantly set in motion a decade ago with the design and construction of a genetic toggle switch and an oscillator, with subsequent highlights that have included circuits capable of pattern generation, noise shaping, edge detection and event counting. Here we describe an engineered gene network with global intercellular coupling that is capable of generating synchronized oscillations in a growing population of cells. Using microfluidic devices tailored for cellular populations at differing length scales, we investigate the collective synchronization properties along with spatiotemporal waves occurring at millimetre scales. We use computational modelling to describe quantitatively the observed dependence of the period and amplitude of the bulk oscillations on the flow rate. The synchronized genetic clock sets the stage for the use of microbes in the creation of a macroscopic biosensor with an oscillatory output. Furthermore, it provides a specific model system for the generation of a mechanistic description of emergent coordinated behaviour at the colony level.

Male-male aggression (hereafter referred to as 'aggression') in the vinegar fly Drosophila melanogaster was first described almost a century ago (7). Since then, considerable progress has been made in understanding [...], Aggression is regulated by pheromones in many animal species (1-3). However, in no system have aggression pheromones, their cognate receptors and corresponding sensory neurons been identified. Here we show that 11-cis-vaccenyl acetate (cVA), a male-specific volatile pheromone, robustly promotes male-male aggression in the vinegar fly Drosophila melanogaster. The aggression-promoting effect of synthetic cVA requires olfactory sensory neurons (OSNs) expressing the receptor Or67d (4-6), as well as the receptor itself. Activation of Or67d-expressing OSNs, either by genetic manipulation of their excitability or by exposure to male pheromones in the absence of other classes of OSNs, is sufficient to promote aggression. High densities of male flies can promote aggression by the release of volatile cVA. In turn, cVA-promoted aggression can promote male fly dispersal from a food resource, in a manner dependent on Or67d-expressing OSNs. These data indicate that cVA may mediate negative-feedback control of male population density, through its effect on aggression. Identification of a pheromone--OSN pair controlling aggression in a genetic organism opens the way to unravelling the neurobiology of this evolutionarily conserved behaviour.

With the completion of genome sequences of major model organisms, increasingly sophisticated genetic tools are necessary for investigating the complex and coordinated functions of genes. Here we describe a genetic manipulation system termed 'genomic engineering' in Drosophila. Genomic engineering is a 2-step process that combines the ends-out (replacement) gene targeting with phage integrase [phi]C31-mediated DNA integration. First, through an improved and modified gene targeting method, a founder knockout line is generated by deleting the target gene and replacing it with an integration site of [phi]C31. Second, DNA integration by [phi]C31 is used to reintroduce modified target-gene DNA into the native locus in the founder knock-out line. Genomic engineering permits directed and highly efficient modifications of a chosen genomic locus into virtually any desired mutant allele. We have successfully applied the genomic engineering scheme on 6 different genes and have generated at their loci more than 70 unique alleles. cell polarity | ends-out targeting | homologous recombination | phiC31 integrase

Genetic engineering has been applied to reprogramme non-ribosomal peptide synthetases (NRPSs) to produce novel antibiotics, but little is known about what determines the efficiency of production. We explored module exchanges at nucleotide sequences encoding interpeptide linkers in dptD, a gene encoding a di-modular NRPS subunit that incorporates 3-methylglutamic acid (3m[Glu.sub.12]) and kynurenine ([Kyn.sub.13]) into daptomycin. Mutations causing amino acid substitutions, deletions or insertions in the inter-module linker had no negative effects on lipopeptide yields. Hybrid DptD subunits were generated by fusing the 3m[Glu.sub.12] module to terminal modules from calcium-dependent antibiotic (CDA) or A54145 NRPSs, and recombinants produced daptomycin analogues with [Trp.sub.13] or [lle.sub.13] at high efficiencies. A recombinant expressing DptD with a hybrid [Kyn.sub.13] module containing a di-domain from a D-Asn module caused the production of a new daptomycin analogue containing [Asn.sub.13].

The genome of Mycoplasma gallisepticum strain [R.sub.low] has been sequenced completely, but subsequent genetic studies have been limited by the lack of a replicable vector system. In this study, replicable plasmids were constructed for M. gallisepticum and Mycoplasma imitans using the oriC region upstream from the soj gene. The oriC plasmids of M. gallisepticum (pGTLori) and M. imitans (pMlori) replicated in both species, but Mycoplasma pneumoniae could not support replication of pGTLori. A 180 bp section of the oriC region of M. gallisepticum was found to be the minimal region required for plasmid replication in M. gallisepticum strain S6, the shortest oriC region defined for mycoplasmas. Targeted gene disruption of vlhA1.1 of M. gallisepticum S6 was attempted using these oriC plasmids. Constructs made in pPLoriC7 integrated into the M. gallisepticum genomic oriC region, not into the targeted gene, whereas those made in pMlori disrupted the vlhA1.2 gene, which has 97% DNA sequence identity with the vlhA1.1 gene. During in vitro passages, antimicrobial selection pressure did not influence the rate of chromosomal integration. These oriC plasmids will thus be useful for genetic studies, including inactivation or expression of selected genes, in M. gallisepticum and M. imitans, and will lead to a better understanding of their molecular biology. They are, to our knowledge, the first replicable plasmids developed for the Pneumoniae phylogenetic group of mycoplasmas.

Curcuminoids, which are produced specifically by plants of the order Zingiberales, have long been used as food additives because of their aromatic, stimulant and colouring properties and as traditional Asian medicines because of their anti-tumour, antioxidant and hepatoprotective activities. Curcuminoids are therefore attractive targets for metabolic engineering. An artificial curcuminoid biosynthetic pathway, including reactions of phenylalanine ammonia-lyase (PAL) from the yeast Rhodotorula rubra, 4-coumarate : CoA ligase (4CL) from Lithospermum erythrorhizon and curcuminoid synthase (CUS) from rice (Oryza sativa), a type III polyketide synthase, was constructed in Escherichia coli for the production of curcuminoids. Cultivation of the recombinant E. coli cells in the presence of tyrosine or phenylalanine, or both, led to production of bisdemethoxycurcumin, dicinnamoylmethane and cinnamoyl-p-coumaroylmethane. Another E. coli system carrying 4CL and CUS genes was also used for high-yield production of curcuminoids from exogenously supplemented phenylpropanoid acids: p-coumaric acid, cinnamic acid and ferulic acid. The yields of curucminoids were up to ~100 mg [l.sup.-1]. Furthermore, this system gave approximately 60 mg curcumin [l.sup.-1] from 10 g rice bran pitch, an industrial waste discharged during rice edible oil production, as a source of ferulic acid.

Alternative splicing has emerged as a promising therapeutic target in a number of human disorders. However, the discovery of compounds that target the splicing reaction has been hindered by the lack of suitable high-throughput screening assays. Conversely, the effects of known drugs on the splicing reaction are mostly unclear and not routinely assessed. We have developed a two-color fluorescent reporter for cellular assays of exon inclusion that can accommodate nearly any cassette exon and minimizes interfering effects from changes in transcription and translation. We used microtubule-associated protein tau (MAPT) exon 10, whose missplicing causes frontotemporal dementia, to test the reporter in screening libraries of known bioactive compounds. These screens yielded several compounds that alter the splicing of the exon, both in the reporter and in the endogenous MAPT mRNA. One compound, digoxin, has long been used in the treatment of heart failure, but was not known to modulate splicing. The positive compounds target different signal transduction pathways, and microarray analysis shows that each compound affects the splicing of a different set of exons in addition to MAPT exon 10. Our results identify currently prescribed cardiotonic steroids as modulators of alternative splicing and demonstrate the feasibility of screening for drugs that alter exon inclusion. digoxin | fluorescent reporter | plant steroids | MAPTI FTDP-17

Most parents would do anything within their reach for their children, including genetic modification. Some parents may want taller or faster children, smarter or healthier ones. And some parents may want children that are more comfortable in their family setting. Under special circumstances, would it be wrong for deaf or blind parents to genetically modify and select deaf or blind children? Are there good reasons not to genetically modify children in such ways? Would not prohibitions against such modifications infringe on the liberty and autonomy of prospective parents? Martin Harvey argues against such genetic dis-enhancements on a variety of moral grounds, employing the backing of a variety of traditional moral theories including deontological, teleological, and consequential arguments.

Surprisingly, some believe that it would be morally permissible to genetically un-enhance one's offspring (perhaps to share a trait of the parents or siblings). More common is the belief that it would be morally permissible to genetically enhance one's offspring, as the technology to do so becomes available. Parents believe that it would be good to provide their children with all of the usual opportunities to succeed in life (education, culture, sport). Why not provide them with enhanced traits of being bigger, faster, or smarter? However, would it be obligatory for us to avail ourselves and our future offspring of these techniques, as they become available? Are we wrong not to genetically engineer our kids to be bigger, faster, smarter? Mark Neunder argues for and defends the view that we are so obliged, as the new technology becomes available (as it clearly will).

The recA gene from Thermus thermophilus HB27 was cloned and engineered to obtain insertion (recA::kat) and deletion ([DELTA]recA) derivatives. Transcription of recA in this extreme thermophile was induced by mitomycin C, leading to the synthesis of a monocistronic mRNA. This DNA damage-mediated induction was dependent on the integrity of recA. In addition to UV sensitivity, the recA mutants of T. thermophilus showed severe pleiotropic defects, ranging from irregular nucleoid condensation and segregation to a dramatic reduction in viability during culture. An increase in the frequency of both carotenoidless and auxotrophic mutants within surviving cells of the [DELTA]recA strain indicated a high mutation rate. As RecA is not required for plasmid transformation, we have used the [alpha]-lacZ gene fragment and the ampicillin resistance gene from Escherichia coli as passenger reporters to confirm such high mutation rates. Our data support the idea that the absence of RecA results in a hypermutational phenotype in T. thermophilus. Furthermore, a direct relationship is deduced between the growth temperature and mutation rate, which finally has a deleterious effect on cell survival in the absence of RecA.

In the fall of 2000, the American Association for the Advancement of Science called on science to slow down. In a report it issued on inheritable genetic modification, AAAS took [...]

The current understanding of the relationships between cell shape, intracellular forces and signaling, nuclear shape and organization, and gene expression is in its infancy. Here we introduce a method for investigating gene-specific responses in individual cells with controlled nuclear shape and projected area. The shape of the nuclei of primary osteogenic cells were controlled on microfabricated substrata with regiospecific chemistry by confining attachment and spreading of isolated cells on adhesive islands. Gene expression and protein synthesis were altered by changing nuclear shape. Collagen I synthesis correlated directly with cell shape and nuclear shape index (NSI), where intermediate values of nuclear distension (6 < NSI < 8) promoted maximum synthesis, Osteocalcin mRNA, a bone-specific differentiation marker, was observed intra-cellularly by using reverse transcription in situ PCR at 4 days in cells constrained by the pattern and not detected in unconstrained cells of similar projected area, but different NSI. Our data supports the concept of gene expression and protein synthesis based on optimal distortion of the nucleus, possibly altering transcription factor affinity for DNA, transport to the nucleus, or nuclear matrix organization. The combination of microfabricated surfaces, reverse transcription in situ PCR, and NSI measurement is an excellent system to study how transcription factors, the nuclear matrix, and the cytoskeleton interact to control gene expression and may be useful for studying a wide variety of other cell shape/gene expression relationships. patterning | cell size | osteocalcin | reverse transcription-PCR | photolithography

The synthesis of rodlike polymer derivatives of poly(eta-benzyl alpha,L-glutamate) using genetic engineering is described. Identification and smectic ordering on the nanometer scale of these monodisperse derivatives is discussed.

Cell-based biosensing systems differ from other types of biosensors in that they provide the physiological response of the cell to the analyte as well as the bioavailability of the analyte being measured. This review focuses on the design, development, and applications of cell-based biosensing systems employing genetically engineered reporter genes.

Research describes a novel pathway designed for poly(hydroxyalkanoic acid) (PHA) synthesis from hydroxyfatty acids mediated by three sets of genes each individually coding for butyrate kinase, phosphotransbutyrylase, and PHA synthase enzymes.

Specific DNA amplification (SDA) is an efficient means of enhancing the symbiotic nitrogen fixation capacity of alfalfa plants, as revealed by an experiment conducted on Sinorhizobium meliloti strains. Strains derived from the duplication of the symbiotic region of the plant sample did not only manifest improved symbiotic properties, but exhibited increased nodulation as well. The strains also proved effective in facilitating the growth of alfalfa plants.

The contribution of HMW1 in the cytadherence of the M6 mutant was investigated via genetic manipulation. The introduction of a recombinant transposon led to the transformation of the mutant. The transformants were found to synthesize the truncated P30 from the resident allele, while the full-length P30 was synthesized from the recombinant alleles. The results further revealed that the transformants were constantly hemadsorption negative, indicating that HMW1 is essential for cytadherence.