NanoJ a high-performance open-source super-resolution microscopy toolbox, bioRxiv, 2018-10-02
Super-resolution microscopy has become essential for the study of nanoscale biological processes. This type of imaging often requires the use of specialised image analysis tools to process a large volume of recorded data and extract quantitative information. In recent years, our team has built an open-source image analysis framework for super-resolution microscopy designed to combine high performance and ease of use. We named it NanoJ - a reference to the popular ImageJ software it was developed for. In this paper, we highlight the current capabilities of NanoJ for several essential processing steps spatio-temporal alignment of raw data (NanoJ-Core), super-resolution image reconstruction (NanoJ-SRRF), image quality assessment (NanoJ-SQUIRREL), structural modelling (NanoJ-VirusMapper) and control of the sample environment (NanoJ-Fluidics). We expect to expand NanoJ in the future through the development of new tools designed to improve quantitative data analysis and measure the reliability of fluorescent microscopy studies.
biorxiv bioinformatics 100-200-users 2018Quiescent cells actively replenish CENP-A nucleosomes to maintain centromere identity and proliferative potential, bioRxiv, 2018-10-02
SummaryCentromeres provide a robust model for epigenetic inheritance as they are specified by sequence-independent mechanisms involving the histone H3-variant CENP-A. Prevailing models indicate that the high intrinsic stability of CENP-A nucleosomes maintains centromere identity indefinitely. Here, we demonstrate that CENP-A is not stable at centromeres, but is instead gradually and continuously incorporated in quiescent cells including G0-arrested tissue culture cells and prophase I-arrested oocytes. Quiescent CENP-A incorporation involves the canonical CENP-A deposition machinery, but displays distinct requirements from cell cycle-dependent deposition. We demonstrate that Plk1 is required specifically for G1 CENP-A deposition, whereas transcription promotes CENP-A incorporation in quiescent oocytes. Preventing CENP-A deposition during quiescence results in significantly reduced CENP-A levels and perturbs chromosome segregation following the resumption of cell division. In contrast to quiescent cells, terminally differentiated cells fail to maintain CENP-A levels. Our work reveals that quiescent cells actively maintain centromere identity providing an indicator of proliferative potential.
biorxiv cell-biology 0-100-users 2018There is no single functional atlas even for a single individual Parcellation of the human brain is state dependent, bioRxiv, 2018-10-02
AbstractThe goal of human brain mapping has long been to delineate the functional subunits in the brain and elucidate the functional role of each of these brain regions. Recent work has focused on whole-brain parcellation of functional Magnetic Resonance Imaging (fMRI) data to identify these subunits and create a functional atlas. Functional connectivity approaches to understand the brain at the network level require such an atlas to assess connections between parcels and extract network properties. While no single functional atlas has emerged as the dominant atlas to date, there remains an underlying assumption that such an atlas exists. Using fMRI data from a highly sampled subject as well as two independent replication data sets, we demonstrate that functional parcellations based on fMRI connectivity data reconfigure substantially and in a meaningful manner, according to brain state.
biorxiv neuroscience 200-500-users 2018A spatial atlas of inhibitory cell types in mouse hippocampus, bioRxiv, 2018-10-01
Understanding the function of a tissue requires knowing the spatial organization of its constituent cell types. In the cerebral cortex, single-cell RNA sequencing (scRNA-seq) has revealed the genome-wide expression patterns that define its many, closely related cell types, but cannot reveal their spatial arrangement. Here we introduce probabilistic cell typing by in situ sequencing (pciSeq), an approach that leverages prior scRNA-seq classification to identify cell types using multiplexed in situ RNA detection. We applied this method to map the inhibitory neurons of hippocampal area CA1, a cell system critical for memory function, for which ground truth is available from extensive prior work identifying the laminar organization of subtly differing cell types. Our method confidently identified 16 interneuron classes, in a spatial arrangement closely matching ground truth. This method will allow identifying the spatial organization of fine cell types across the brain and other tissues.
biorxiv neuroscience 100-200-users 2018A mouse tissue atlas of small non-coding RNA, bioRxiv, 2018-09-29
SUMMARYSmall non-coding RNAs (ncRNAs) play a vital role in a broad range of biological processes both in health and disease. A comprehensive quantitative reference of small ncRNA expression would significantly advance our understanding of ncRNA roles in shaping tissue functions. Here, we systematically profiled the levels of five ncRNA classes (miRNA, snoRNA, snRNA, scaRNA and tRNA fragments) across eleven mouse tissues by deep sequencing. Using fourteen biological replicates spanning both sexes, we identified that ~ 30% of small ncRNAs are distributed across the body in a tissue-specific manner with some are also being sexually dimorphic. We found that miRNAs are subject to “arm switching” between healthy tissues and that tRNA fragments are retained within tissues in both a gene- and a tissue-specific manner. Out of eleven profiled tissues we confirmed that brain contains the largest number of unique small ncRNA transcripts, some of which were previously annotated while others are identified for the first time in this study. Furthermore, by combining these findings with single-cell ATAC-seq data, we were able to connect identified brain-specific ncRNA with their cell types of origin. These results yield the most comprehensive characterization of specific and ubiquitous small RNAs in individual murine tissues to date, and we expect that this data will be a resource for the further identification of ncRNAs involved in tissue-function in health and dysfunction in disease.HIGHLIGHTS<jatslist list-type=simple><jatslist-item>-An atlas of tissue levels of multiple small ncRNA classes generated from 14 biological replicates of both sexes across 11 tissues<jatslist-item><jatslist-item>-Distinct distribution patterns of miRNA arms and tRNA fragments across tissues suggest the existence of tissue-specific mechanisms of ncRNA cleavage and retention<jatslist-item><jatslist-item>-miRNA expression is sex specific in healthy tissues<jatslist-item><jatslist-item>-Small RNA-seq and scATAC-seq data integration produce a detailed map of cell-type specific ncRNA profiles in the mouse brain<jatslist-item>
biorxiv genomics 0-100-users 2018Pathogen Detection and Microbiome Analysis of Infected Wheat Using a Portable DNA Sequencer, bioRxiv, 2018-09-29
ABSTRACTFungal diseases of plants are responsible for major losses in agriculture, highlighting the need for rapid and accurate identification of plant pathogens. Disease outcomes are often defined not only by the main pathogen but are influenced by diverse microbial communities known as the microbiome at sites of infection. Here we present the first use of whole genome shot-gun sequencing with a portable DNA sequencing device as a method for the detection of fungal pathogens from wheat (Triticum aestivum) in a standard molecular biology laboratory. The data revealed that our method is robust and applicable to the diagnosis of fungal diseases including wheat stripe rust (caused by Puccinia striiformis f. sp. tritici), septoria tritici blotch (caused by Zymoseptoria tritici) and yellow leaf spot (caused by Pyrenophora tritici repentis). We also identified the bacterial genus Pseudomonas co-present with Puccinia and Zymoseptoria but not Pyrenophora infections. One limitation of the method is the over-representation of redundant wheat genome sequences from samples. This could be addressed by long-range amplicon-based sequencing approaches in future studies, which specifically target non-host organisms. Our work outlines a new approach for detection of a broad range of plant pathogens and associated microbes using a portable sequencer in a standard laboratory, providing the basis for future development of an on-site disease monitoring system.
biorxiv plant-biology 0-100-users 2018