How research institutions can foster innovation.

Carrying out research means being innovative, which requires novelty. Novelty is an important source of scientific breakthroughs and has great technological impact. Research institutions stand to benefit from fostering innovation. Here, we outline what academic institutions can do to help their scientists become more innovative.

Historic nucleic acids isolated by Friedrich Miescher contain RNA besides DNA.

One hundred fifty years ago, Friedrich Miescher discovered DNA when he isolated "Nuclein"-as he named it-from nuclei of human pus cells. Miescher recognized his isolate as a new type of molecule equal in importance to proteins. He realised that it is an acid of large molecular weight and high phosphorus content. Subsequently, he discovered Nuclein also in the nuclei of other cell types, realised that it chemically defines the nucleus, and speculated on its role in proliferation, heredity and fertilisation. While now universally recognised as the discoverer of DNA, whether Miescher also discovered RNA has not yet been addressed. To determine whether his isolation also yielded RNA, we first reproduced his historic protocols. Our resulting modern Nuclein contained a significant percentage of RNA. Encouraged by this result, we then analysed a sample of Nuclein isolated by Miescher from salmon sperm. Assuming that the RNA present in this sample had degraded to nucleobases, we tested for the presence of uracil in the historic Nuclein. Detection of significant levels of uracil by LC-UV-MS demonstrates that Miescher isolated both forms of nucleic acid-DNA and RNA-and underlines the fundamental nature of his discovery for the field of molecular genetics.

How We Forgot Who Discovered DNA: Why It Matters How You Communicate Your Results.

One hundred and fifty years ago, a hopeful young researcher reported a recent discovery he had made. Working in the bowels of a medieval castle in the German city of Tübingen, he had isolated a then entirely new type of molecule. This was the birth of a field that would fundamentally change the course of biology, medicine, and beyond. His discovery: DNA. His name: Friedrich Miescher. In this article, the authors try to find answers to the question why-despite the fact that virtually everyone nowadays knows DNA-hardly anyone remembers the man who discovered it. In the history of science, the discovery of DNA was a seminal moment. Why then did it not enter into public memory? Ground-breaking discoveries can occur in a historical context that is not ready to appreciate them. But that's not all that decides who is remembered and who is forgotten. Scientific pioneers sometimes fail to publicize their findings in a way that ensures that they receive the attention they merit. As discussed here, their personalities and habits may cause discoveries to be "overwritten" by more recent researchers, resulting in distorted cultural memories no longer reflecting the initial event.

Interdisciplinary Communication Needs to Become a Core Scientific Skill.

As scientific research has advanced so too has the complexity of the questions addressed. Cross-disciplinary collaborations are often the most efficient route to managing that complexity and require effective communication across boundaries. To continue driving science forward and be able to tackle global challenges, the art of good interdisciplinary communication needs to become a core skill in a scientist's portfolio.

Evolution of the vertebrate beaded filament protein, Bfsp2; comparing the in vitro assembly properties of a "tailed" zebrafish Bfsp2 to its "tailless" human orthologue.

In bony fishes, Bfsp2 orthologues are predicted to possess a C-terminal tail domain, which is absent from avian, amphibian and mammalian Bfsp2 sequences. These sequences, are however, not conserved between fish species and therefore questions whether they have a functional role. For other intermediate filament proteins, the C-terminal tail domain is important for both filament assembly and regulating interactions between filaments. We confirm that zebrafish has a single Bfsp2 gene by radiation mapping. Two transcripts (bfsp2α and bfsp2ß) are produced by alternative splicing of the last exon. Using a polyclonal antibody specific to a tridecameric peptide in the C-terminal tail domain common to both zebrafish Bfsp2 splice variants, we have confirmed its expression in zebrafish lens fibre cells. We have also determined the in vitro assembly properties of zebrafish Bfsp2α and conclude that the C-terminal sequences are required to regulate not only the diameter and uniformity of the in vitro assembly filaments, but also their filament-filament associations in vitro. Therefore we conclude zebrafish Bfsp2α is a functional orthologue conforming more closely to the conventional domain structure of intermediate filament proteins. Data mining of the genome databases suggest that the loss of this tail domain could occur in several stages leading eventually to completely tailless orthologues, such as human BFSP2.

Investigating the genetics of visual processing, function and behaviour in zebrafish.

Over the past three decades, the zebrafish has been proven to be an excellent model to investigate the genetic control of vertebrate embryonic development, and it is now also increasingly used to study behaviour and adult physiology. Moreover, mutagenesis approaches have resulted in large collections of mutants with phenotypes that resemble human pathologies, suggesting that these lines can be used to model diseases and screen drug candidates. With the recent development of new methods for gene targeting and manipulating or monitoring gene expression, the range of genetic modifications now possible in zebrafish is increasing rapidly. Combined with the classical strengths of the zebrafish as a model organism, these advances are set to substantially expand the type of biological questions that can be addressed in this species. In this review, we outline how the potential of the zebrafish can be harvested in the context of eye development and visual function. We review recent technological advances used to study the formation of the eyes and visual areas of the brain, visual processing on the cellular, subcellular and molecular level, and the genetics of visual behaviour in vertebrates.

Subfunctionalization of duplicated zebrafish pax6 genes by cis-regulatory divergence.

Gene duplication is a major driver of evolutionary divergence. In most vertebrates a single PAX6 gene encodes a transcription factor required for eye, brain, olfactory system, and pancreas development. In zebrafish, following a postulated whole-genome duplication event in an ancestral teleost, duplicates pax6a and pax6b jointly fulfill these roles. Mapping of the homozygously viable eye mutant sunrise identified a homeodomain missense change in pax6b, leading to loss of target binding. The mild phenotype emphasizes role-sharing between the co-orthologues. Meticulous mapping of isolated BACs identified perturbed synteny relationships around the duplicates. This highlights the functional conservation of pax6 downstream (3') control sequences, which in most vertebrates reside within the introns of a ubiquitously expressed neighbour gene, ELP4, whose pax6a-linked exons have been lost in zebrafish. Reporter transgenic studies in both mouse and zebrafish, combined with analysis of vertebrate sequence conservation, reveal loss and retention of specific cis-regulatory elements, correlating strongly with the diverged expression of co-orthologues, and providing clear evidence for evolution by subfunctionalization.

The GTP-binding protein Septin 7 is critical for dendrite branching and dendritic-spine morphology.

Septins, a highly conserved family of GTP-binding proteins, were originally identified in a genetic screen for S. cerevisiae mutants defective in cytokinesis [1, 2]. In yeast, septins maintain the compartmentalization of the yeast plasma membrane during cell division by forming rings at the cortex of the bud neck, and these rings establish a lateral diffusion barrier. In contrast, very little is known about the functions of septins in mammalian cells [3, 4] including postmitotic neurons [5-7]. Here, we show that Septin 7 (Sept7) localizes at the bases of filopodia and at branch points in developing hippocampal neurons. Upon downregulation of Sept7, dendritic branching is impaired. In mature neurons, Sept7 is found at the bases of dendritic spines where it associates with the plasma membrane. Mature Sept7-deficient neurons display elongated spines. Furthermore, Sept5 and Sept11 colocalize with and coimmunoprecipitate with Sept7, thereby arguing for the existence of a Septin5/7/11 complex. Taken together, our findings show an important role for Sept7 in regulating dendritic branching and dendritic-spine morphology. Our observations concur with data from yeast, in which downregulation of septins yields elongated buds, suggesting a conserved function for septins from yeast to mammals.

A putative nuclear function for mammalian Staufen.

In addition to its role in rRNA processing and ribosome assembly, the nucleolus plays a part in the assembly of non-ribosomal ribonucleoprotein particles (RNPs) that are destined for cytoplasmic RNA delivery. Recent evidence indicates that mammalian Staufen2, a brain-specific RNA-binding protein involved in RNA localization, can--at least transiently--enter the nucleolus. Therefore, the assembly of Staufen2 into transport-competent RNPs might occur in the nucleus before their export into the cytoplasm. This could provide new insights into the mechanisms of subcellular RNA localization.

Dynamic interaction between P-bodies and transport ribonucleoprotein particles in dendrites of mature hippocampal neurons.

The dendritic localization of mRNAs and their subsequent translation at stimulated synapses contributes to the experience-dependent remodeling of synapses and thereby to the establishment of long-term memory. Localized mRNAs are transported in a translationally silent manner to distal dendrites in specific ribonucleoprotein particles (RNPs), termed transport RNPs. A recent study suggested that processing bodies (P-bodies), which have recently been identified as sites of RNA degradation and translational control in eukaryotic cells, may participate in the translational control of dendritically localized mRNAs in Drosophila neurons. This study raised the interesting question of whether dendritic transport RNPs are distinct from P-bodies or whether those structures share significant overlap in their molecular composition in mammalian neurons. Here, we show that P-body and transport RNP markers do not colocalize and are not transported together in the same particles in dendrites of mammalian neurons. Detailed time-lapse videomicroscopy analyses reveal, however, that both P-bodies and transport RNPs can interact in a dynamic manner via docking. Docking is a frequent event involving as much as 50% of all dendritic P-bodies. Chemically induced neuronal activity results in a 60% decrease in the number of P-bodies in dendrites, suggesting that P-bodies disassemble after synaptic stimulation. Our data lend support to the exciting hypothesis that dendritically localized mRNAs might be stored in P-bodies and be released and possibly translated when synapses become activated.

High-efficiency transfection of short hairpin RNAs-encoding plasmids into primary hippocampal neurons.

The transfection of expression constructs encoding a variety of transgenes is a widely used method to study gene function in cultured cells. Especially when the efficiency of the knock-down of target proteins via small interfering RNAs (siRNAs) is to be determined by quantitative Western blotting, large proportions of untransfected cells compromise the analysis. Achieving high transfection efficiencies in postmitotic cells, such as neurons, poses a particular problem in that these cells cannot be selected for the expression of the transgene following transfection. It is therefore important to develop protocols that allow for the highly efficient transfection of these cells. In the present study, we identify three important parameters that prove especially useful for chronically difficult to transfect short hairpin RNA (shRNA)-encoding plasmids: the amount and quality of the plasmid DNA used and the use of new nucleofection programs. Combining those changes increases the rate of transfected cells from less than 5% to up to approximately 80%. Importantly, these high transfection efficiencies can be obtained while maintaining good cell viability and normal cellular development. Taken together, these improvements allow for a detailed biochemical and phenotypical analysis of neurons that have been nucleoporated with a wide variety of shRNAs.

Discovering DNA: Friedrich Miescher and the early years of nucleic acid research.

In the winter of 1868/9 the young Swiss doctor Friedrich Miescher, working in the laboratory of Felix Hoppe-Seyler at the University of Tübingen, performed experiments on the chemical composition of leukocytes that lead to the discovery of DNA. In his experiments, Miescher noticed a precipitate of an unknown substance, which he characterised further. Its properties during the isolation procedure and its resistance to protease digestion indicated that the novel substance was not a protein or lipid. Analyses of its elementary composition revealed that, unlike proteins, it contained large amounts of phosphorous and, as Miescher confirmed later, lacked sulphur. Miescher recognised that he had discovered a novel molecule. Since he had isolated it from the cells' nuclei he named it nuclein, a name preserved in today's designation deoxyribonucleic acid. In subsequent work Miescher showed that nuclein was a characteristic component of all nuclei and hypothesised that it would prove to be inextricably linked to the function of this organelle. He suggested that its abundance in tissues might be related to their physiological status with increases in "nuclear substances" preceding cell division. Miescher even speculated that it might have a role in the transmission of hereditary traits, but subsequently rejected the idea. This article reviews the events and circumstances leading to Miescher's discovery of DNA and places them within their historic context. It also tries to elucidate why it was Miescher who discovered DNA and why his name is not universally associated with this molecule today.

Perplexing bodies: The putative roles of P-bodies in neurons.

Processing bodies (P-bodies) have recently come to the fore as important cellular sites of mRNA degradation and translational silencing. Despite these central functions in the control of gene expression, the roles of P-bodies have only been characterized in a limited number of cell types and physiological contexts. Neurons are highly plastic cells that undergo dynamic changes as new connections are made or existing ones modified. This neuronal plasticity relies, in part, on the local synthesis of proteins from localized mRNAs. A strict control of the translation and turnover of these localized mRNAs, both in terms of which proteins are synthesized and when and where they are produced, is a key prerequisite for this process to be synapse-specific. Despite recent advances, the molecular mechanisms mediating this control remain largely elusive. The discovery of P-bodies in neuronal dendrites near synapses and their response to stimuli involved in neuronal plasticity raises the interesting hypothesis that P-bodies might be a component of the cellular machinery that controls neuronal plasticity and thereby processes such as learning and memory.

Human pathologies associated with defective RNA transport and localization in the nervous system.

RNA localization is emerging as an important process to restrict certain proteins to specific subcellular domains and thus spatially control the expression of genes within cells. It is used, for instance, to compartmentalize the developing embryo during early embryogenesis. The localization of RNA also plays important roles later during development, such as in asymmetric cell divisions, cell migration and the outgrowth and pathfinding of axons and dendrites. In differentiated cells, it serves to subdivide the cell into functionally distinct compartments. For example, in mature neurons it is believed to contribute to the plastic changes of individual synapses underlying learning and memory. In this review, we highlight the importance of subcellular RNA localization for the function of the nervous system and neurological diseases associated with defective RNA localization and translation. These diseases include fragile X mental retardation syndrome, spinocerebellar ataxia and spinal muscular atrophy.

Dendritic localization of the translational repressor Pumilio 2 and its contribution to dendritic stress granules.

Pumilio (Pum) protein acts as a translational inhibitor in several organisms including yeast, Drosophila, Xenopus, and mammals. Two Pumilio genes, Pum1 and Pum2, have been identified in mammals, but their function in neurons has not been identified. In this study, we found that Pum2 mRNA is expressed during neuronal development and that the protein is found in discrete particles in both the cell body and the dendritic compartment of fully polarized neurons. This finding indicates that Pum2 is a novel candidate of dendritically localized ribonucleoparticles (RNPs). During metabolic stress, Pum2 is present in stress granules (SGs), which are subsequently detected in the somatodendritic domain. It remains excluded from processing bodies under all conditions. When overexpressed in neurons and fibroblasts, Pum2 induces the formation of SGs that also contain T-cell intracellular antigen 1 (TIA-1)-related protein, eukaryotic initiation factor 4E, poly(A)-binding protein, TIA-1, and other RNA-binding proteins including Staufen1 and Barentsz. This induction of SGs is dependent on the RNA-binding domain and a glutamine-rich region in the N terminus of Pum2. This glutamine-rich region behaves in a similar manner as TIA-1 and prion protein, two molecules with known roles in protein aggregation. Pum2 downregulation in neurons via RNA interference (RNAi) interferes with the formation of SGs during metabolic stress. Cotransfection with an RNAi-resistant portion of the Pum2 mRNA restores SG formation. These results suggest a role for Pum2 in dendritic RNPs and SG formation in mammalian neurons.

Large-scale mapping of mutations affecting zebrafish development.

BACKGROUND: Large-scale mutagenesis screens in the zebrafish employing the mutagen ENU have isolated several hundred mutant loci that represent putative developmental control genes. In order to realize the potential of such screens, systematic genetic mapping of the mutations is necessary. Here we report on a large-scale effort to map the mutations generated in mutagenesis screening at the Max Planck Institute for Developmental Biology by genome scanning with microsatellite markers. RESULTS: We have selected a set of microsatellite markers and developed methods and scoring criteria suitable for efficient, high-throughput genome scanning. We have used these methods to successfully obtain a rough map position for 319 mutant loci from the Tübingen I mutagenesis screen and subsequent screening of the mutant collection. For 277 of these the corresponding gene is not yet identified. Mapping was successful for 80 % of the tested loci. By comparing 21 mutation and gene positions of cloned mutations we have validated the correctness of our linkage group assignments and estimated the standard error of our map positions to be approximately 6 cM. CONCLUSION: By obtaining rough map positions for over 300 zebrafish loci with developmental phenotypes, we have generated a dataset that will be useful not only for cloning of the affected genes, but also to suggest allelism of mutations with similar phenotypes that will be identified in future screens. Furthermore this work validates the usefulness of our methodology for rapid, systematic and inexpensive microsatellite mapping of zebrafish mutations.

Living autobiographically: Concepts of aging and artistic expression in painting and modern dance.

This article discusses the ways in which artists have incorporated or failed to incorporate the aging process of their bodies into their art. Using Russian ballet dancer Mikhail Baryshnikov and the French painter Claude Monet as cases in point, we explore situations in which physical changes brought about by aging compromises artists' ability to engage with their artistic medium. Connecting Monet's oeuvre and Baryshnikov's dance performances to life writing accounts, we draw on John Paul Eakin's concept of "living autobiographically": In this vein, life writing research does not only have to take into account concepts of identity as they emerge from life writing narratives, but it also needs to explore the somatic, corporeal and material dimensions of these narratives.