Sequence-Based Mapping and Genome Editing Reveal Mutations in Stickleback Hps5 Cause Oculocutaneous Albinism and the casper Phenotype.

Here, we present and characterize the spontaneous X-linked recessive mutation casper, which causes oculocutaneous albinism in threespine sticklebacks (Gasterosteus aculeatus). In humans, Hermansky-Pudlak syndrome results in pigmentation defects due to disrupted formation of the melanin-containing lysosomal-related organelle (LRO), the melanosome. casper mutants display not only reduced pigmentation of melanosomes in melanophores, but also reductions in the iridescent silver color from iridophores, while the yellow pigmentation from xanthophores appears unaffected. We mapped casper using high-throughput sequencing of genomic DNA from bulked casper mutants to a region of the stickleback X chromosome (chromosome 19) near the stickleback ortholog of Hermansky-Pudlak syndrome 5 (Hps5). casper mutants have an insertion of a single nucleotide in the sixth exon of Hps5, predicted to generate an early frameshift. Genome editing using CRISPR/Cas9 induced lesions in Hps5 and phenocopied the casper mutation. Injecting single or paired Hps5 guide RNAs revealed higher incidences of genomic deletions from paired guide RNAs compared to single gRNAs. Stickleback Hps5 provides a genetic system where a hemizygous locus in XY males and a diploid locus in XX females can be used to generate an easily scored visible phenotype, facilitating quantitative studies of different genome editing approaches. Lastly, we show the ability to better visualize patterns of fluorescent transgenic reporters in Hps5 mutant fish. Thus, Hps5 mutations present an opportunity to study pigmented LROs in the emerging stickleback model system, as well as a tool to aid in assaying genome editing and visualizing enhancer activity in transgenic fish

Scattering a pulse from a chaotic cavity: Transitioning from algebraic to exponential decay

The ensemble averaged power scattered in and out of lossless chaotic cavities decays as a power law in time for large times. In the case of a pulse with a finite duration, the power scattered from a single realization of a cavity closely tracks the power law ensemble decay initially, but eventually transitions to an exponential decay. In this paper, we explore the nature of this transition in the case of coupling to a single port. We find that for a given pulse shape, the properties of the transition are universal if time is properly normalized. We define the crossover time to be the time at which the deviations from the mean of the reflected power in individual realizations become comparable to the mean reflected power. We demonstrate numerically that, for randomly chosen cavity realizations and given pulse shapes, the probability distribution function of reflected power depends only on time, normalized to this crossover time.Comment: 23 pages, 5 figure

Increasing β-catenin/Wnt3A activity levels drive mechanical strain-induced cell cycle progression through mitosis.

Mechanical force and Wnt signaling activate β-catenin-mediated transcription to promote proliferation and tissue expansion. However, it is unknown whether mechanical force and Wnt signaling act independently or synergize to activate β-catenin signaling and cell division. We show that mechanical strain induced Src-dependent phosphorylation of Y654 β-catenin and increased β-catenin-mediated transcription in mammalian MDCK epithelial cells. Under these conditions, cells accumulated in S/G2 (independent of DNA damage) but did not divide. Activating β-catenin through Casein Kinase I inhibition or Wnt3A addition increased β-catenin-mediated transcription and strain-induced accumulation of cells in S/G2. Significantly, only the combination of mechanical strain and Wnt/β-catenin activation triggered cells in S/G2 to divide. These results indicate that strain-induced Src phosphorylation of β-catenin and Wnt-dependent β-catenin stabilization synergize to increase β-catenin-mediated transcription to levels required for mitosis. Thus, local Wnt signaling may fine-tune the effects of global mechanical strain to restrict cell divisions during tissue development and homeostasis

Spatial distribution of cell-cell and cell-ECM adhesions regulates force balance while main-taining E-cadherin molecular tension in cell pairs.

Mechanical linkage between cell-cell and cell-extracellular matrix (ECM) adhesions regulates cell shape changes during embryonic development and tissue homoeostasis. We examined how the force balance between cell-cell and cell-ECM adhesions changes with cell spread area and aspect ratio in pairs of MDCK cells. We used ECM micropatterning to drive different cytoskeleton strain energy states and cell-generated traction forces and used a Förster resonance energy transfer tension biosensor to ask whether changes in forces across cell-cell junctions correlated with E-cadherin molecular tension. We found that continuous peripheral ECM adhesions resulted in increased cell-cell and cell-ECM forces with increasing spread area. In contrast, confining ECM adhesions to the distal ends of cell-cell pairs resulted in shorter junction lengths and constant cell-cell forces. Of interest, each cell within a cell pair generated higher strain energies than isolated single cells of the same spread area. Surprisingly, E-cadherin molecular tension remained constant regardless of changes in cell-cell forces and was evenly distributed along cell-cell junctions independent of cell spread area and total traction forces. Taken together, our results showed that cell pairs maintained constant E-cadherin molecular tension and regulated total forces relative to cell spread area and shape but independently of total focal adhesion area

Hygienic behaviour in the Australian stingless bees Tetragonula carbonaria and T. hockingsi

Hygienic behaviour is a natural mechanism of colony-level disease resistance to brood pathogens and has been reported in honey bees and stingless bees. A novel brood disease was recently confirmed in the Australian stingless bees Tetragonula carbonaria Smith and Tetragonula hockingsi Cockerell and there is a paucity of data available on hygienic behaviour in these species. To address this, we investigated hygienic behaviour in eight colonies of T. carbonaria and four colonies of T. hockingsi, using brood freeze-kill and pin-kill assays. Hygienic behaviour was present in both species and was rapidly expressed in both assays. In T. carbonaria, the mean time (± SE) for removal of freeze-killed and pin-killed brood was 9.1 ± 1.9 hours and 8.2 ± 0.9 hours, respectively (n=8; one trial per assay). In T. hockingsi, removal of freeze-killed and pin-killed brood was 14.1 ± 5.1 hours and 10.4 (no SE) hours, respectively. There was no significant difference (α=0.05) in time taken to complete the hygienic behaviour phases (detection, uncapping, removal or cell dismantling) between assay type or assay order in both species. However, intercolony variation was observed in both species in the assays, suggesting that like honey bees, hygienic behaviour may have a genetic component. Tetragonula carbonaria and T. hockingsi displayed significantly faster detection, uncapping, removal and cell dismantling times than any of the stingless bees or most honey bees studied previously. This may, in part, explain why stingless bees appear to suffer from relatively few brood diseases

Childhood and the politics of scale: Descaling children's geographies?

This is the post-print version of the final published paper that is available from the link below. Copyright @ 2008 SAGE Publications.The past decade has witnessed a resurgence of interest in the geographies of children's lives, and particularly in engaging the voices and activities of young people in geographical research. Much of this growing body of scholarship is characterized by a very parochial locus of interest — the neighbourhood, playground, shopping mall or journey to school. In this paper I explore some of the roots of children's geographies' preoccupation with the micro-scale and argue that it limits the relevance of research, both politically and to other areas of geography. In order to widen the scope of children's geographies, some scholars have engaged with developments in the theorization of scale. I present these arguments but also point to their limitations. As an alternative, I propose that the notion of a flat ontology might help overcome some difficulties around scalar thinking, and provide a useful means of conceptualizing sociospatiality in material and non-hierarchical terms. Bringing together flat ontology and work in children's geographies on embodied subjectivity, I argue that it is important to examine the nature and limits of children's spaces of perception and action. While these spaces are not simply `local', they seldom afford children opportunities to comment on, or intervene in, the events, processes and decisions that shape their own lives. The implications for the substance and method of children's geographies and for geographical work on scale are considered

Visible camera cryostat design and performance for the SuMIRe Prime Focus Spectrograph (PFS)

We describe the design and performance of the SuMIRe Prime Focus Spectrograph (PFS) visible camera cryostats. SuMIRe PFS is a massively multi-plexed ground-based spectrograph consisting of four identical spectrograph modules, each receiving roughly 600 fibers from a 2394 fiber robotic positioner at the prime focus. Each spectrograph module has three channels covering wavelength ranges 380~nm -- 640~nm, 640~nm -- 955~nm, and 955~nm -- 1.26~um, with the dispersed light being imaged in each channel by a f/1.07 vacuum Schmidt camera. The cameras are very large, having a clear aperture of 300~mm at the entrance window, and a mass of $\sim$280~kg. In this paper we describe the design of the visible camera cryostats and discuss various aspects of cryostat performance