Many different functions are regulated by circadian rhythms, including those orchestrated by discrete clock neurons within animal brains. To comprehensively characterize and assign cell identity to ...the 75 pairs of
circadian neurons, we optimized a single-cell RNA sequencing method and assayed clock neuron gene expression at different times of day. The data identify at least 17 clock neuron categories with striking spatial regulation of gene expression. Transcription factor regulation is prominent and likely contributes to the robust circadian oscillation of many transcripts, including those that encode cell-surface proteins previously shown to be important for cell recognition and synapse formation during development. The many other clock-regulated genes also constitute an important resource for future mechanistic and functional studies between clock neurons and/or for temporal signaling to circuits elsewhere in the fly brain.
Stroma is a poorly defined non-parenchymal component of virtually every organ with key roles in organ development, homeostasis, and repair. Studies of the bone marrow stroma have defined individual ...populations in the stem cell niche regulating hematopoietic regeneration and capable of initiating leukemia. Here, we use single-cell RNA sequencing (scRNA-seq) to define a cellular taxonomy of the mouse bone marrow stroma and its perturbation by malignancy. We identified seventeen stromal subsets expressing distinct hematopoietic regulatory genes spanning new fibroblastic and osteoblastic subpopulations including distinct osteoblast differentiation trajectories. Emerging acute myeloid leukemia impaired mesenchymal osteogenic differentiation and reduced regulatory molecules necessary for normal hematopoiesis. These data suggest that tissue stroma responds to malignant cells by disadvantaging normal parenchymal cells. Our taxonomy of the stromal compartment provides a comprehensive bone marrow cell census and experimental support for cancer cell crosstalk with specific stromal elements to impair normal tissue function and thereby enable emergent cancer.
Display omitted
•A single-cell bone marrow stromal cell atlas at steady-state and emergent leukemia•A portal for comparative cell and molecular analyses of bone marrow stromal cells•Distinction among putative niche cells and types of osteolineage differentiation•Leukemia remodeling of stroma to the disadvantage of normal hematopoietic cells
Molecular definition of the cell populations comprising bone marrow stroma is provided with single-cell resolution. Seventeen distinct cell subsets with new mesenchymal, pericyte, fibroblast, and endothelial subpopulations, new inferred osteolineage differentiation trajectories, and distinctions among Lepr, Nestin, and NG2-expressing HSC niche populations are defined. The changes imposed by an emergent AML indicate impaired stromal differentiation among mesenchymal cells and decreased expression of hematopoietic regulatory genes consistent with cancer cells indirectly impairing the normal hematopoietic cells with whom they must compete.
How the immune system readies for battleAlthough gene expression is tightly controlled at both the RNA and protein levels, the quantitative contribution of each step, especially during dynamic ...responses, remains largely unknown. Indeed, there has been much debate whether changes in RNA level contribute substantially to protein-level regulation. Jovanovic et al. built a genome-scale model of the temporal dynamics of differential protein expression during the stimulation of immunological dendritic cells (see the Perspective by Li and Biggin). Newly stimulated functions involved the up-regulation of specific RNAs and concomitant increases in the levels of the proteins they encode, whereas housekeeping functions were regulated posttranscriptionally at the protein level.Science, this issue 10.1126/science.1259038; see also p. 1066 Protein expression is regulated by the production and degradation of messenger RNAs (mRNAs) and proteins, but their specific relationships remain unknown. We combine measurements of protein production and degradation and mRNA dynamics so as to build a quantitative genomic model of the differential regulation of gene expression in lipopolysaccharide-stimulated mouse dendritic cells. Changes in mRNA abundance play a dominant role in determining most dynamic fold changes in protein levels. Conversely, the preexisting proteome of proteins performing basic cellular functions is remodeled primarily through changes in protein production or degradation, accounting for more than half of the absolute change in protein molecules in the cell. Thus, the proteome is regulated by transcriptional induction for newly activated cellular functions and by protein life-cycle changes for remodeling of preexisting functions.
Massively parallel DNA sequencing technologies are revolutionizing genomics by making it possible to generate billions of relatively short (approximately 100-base) sequence reads at very low cost. ...Whereas such data can be readily used for a wide range of biomedical applications, it has proven difficult to use them to generate high-quality de novo genome assemblies of large, repeat-rich vertebrate genomes. To date, the genome assemblies generated from such data have fallen far short of those obtained with the older (but much more expensive) capillary-based sequencing approach. Here, we report the development of an algorithm for genome assembly, ALLPATHS-LG, and its application to massively parallel DNA sequence data from the human and mouse genomes, generated on the Illumina platform. The resulting draft genome assemblies have good accuracy, short-range contiguity, long-range connectivity, and coverage of the genome. In particular, the base accuracy is high (≥99.95%) and the scaffold sizes (N50 size = 11.5 Mb for human and 7.2 Mb for mouse) approach those obtained with capillary-based sequencing. The combination of improved sequencing technology and improved computational methods should now make it possible to increase dramatically the de novo sequencing of large genomes. The ALLPATHS-LG program is available at http://www.broadinstitute.org/science/programs/genome-biology/crd.
Developing effective and low‐cost organic semiconductors is an opportunity for the development of organic solar cells (OPV). Herein, we report the molecular design, synthesis and characterization of ...two molecules with D–A–D–A configuration: 2-cyano-3-(5-(8-(3,4-ethylenodioxythiophen-5-yl)-2,3-diphenylquinoxalin-5-yl)thiophen-2-yl)acrylic acid (
6
) and 2-cyano-3-(5-(2,3-diphenyl-8-(thiophen-2-yl)quinoxalin-5-yl)thiophen-2-yl)acrylic acid (
7
). Moreover, we investigated the structural, theoretical and optical properties. The distribution of HOMO/LUMO orbitals and the values of the ionization potential indicate good semiconducting properties of the compounds and that they can be a bipolar material. Also, the optical study show good absorption in visible light (
λ
abs
380–550 nm). We investigate the theoretical optoelectronic properties of obtained compounds as potential materials for solar cells.
Exceptionally accurate genome reference sequences have proven to be of great value to microbial researchers. Thus, to date, about 1800 bacterial genome assemblies have been "finished" at great ...expense with the aid of manual laboratory and computational processes that typically iterate over a period of months or even years. By applying a new laboratory design and new assembly algorithm to 16 samples, we demonstrate that assemblies exceeding finished quality can be obtained from whole-genome shotgun data and automated computation. Cost and time requirements are thus dramatically reduced.
Aging is accompanied by physiological impairments, which, in insulin-responsive tissues, including the liver, predispose individuals to metabolic disease. However, the molecular mechanisms underlying ...these changes remain largely unknown. Here, we analyze genome-wide profiles of RNA and chromatin organization in the liver of young (3 months) and old (21 months) mice. Transcriptional changes suggest that derepression of the nuclear receptors PPARα, PPARγ, and LXRα in aged mouse liver leads to activation of targets regulating lipid synthesis and storage, whereas age-dependent changes in nucleosome occupancy are associated with binding sites for both known regulators (forkhead factors and nuclear receptors) and candidates associated with nuclear lamina (Hdac3 and Srf) implicated to govern metabolic function of aging liver. Winged-helix transcription factor Foxa2 and nuclear receptor corepressor Hdac3 exhibit a reciprocal binding pattern at PPARα targets contributing to gene expression changes that lead to steatosis in aged liver.
Display omitted
•An in vivo genome-wide nucleosome map in the aging liver is described•Foxa2 binds regions of decreased nucleosome occupancy at PPARα targets in old livers•Hdac3 and Srf are implicated in age-dependent metabolic dysfunction•Reciprocal binding of Foxa2 and Hdac3 at PPARα targets contributes to steatosis
Bochkis et al. use genome-wide profiling in the livers of young and old mice to observe age-dependent changes in PPARα targets as well as nucleosome occupancy linked to Foxa2 and Hdac3 binding sites. A reciprocal binding pattern of Foxa2 and Hdac3 at PPARα targets contributes to gene-expression changes that lead to steatosis in the aging liver.
There are many applications in which quantitative information about DNA mixtures with different molecular lengths is important. Gene therapy vectors are much longer than can be sequenced individually ...via short-read NGS. However, vector preparations may contain smaller DNAs that behave differently during sequencing. We have used two library preparations each for Pacific Biosystems (PacBio) and Oxford Nanopore Technologies NGS to determine their suitability for quantitative assessment of varying sized DNAs. Equimolar length standards were generated from E. coli genomic DNA. Both PacBio library preparations provided a consistent length dependence though with a complex pattern. This method is sufficiently sensitive that differences in genomic copy number between DNA from E. coli grown in exponential and stationary phase conditions could be detected. The transposase-based Oxford Nanopore library preparation provided a predictable length dependence, but the random sequence starts caused the loss of original length information. The ligation-based approach retained length information but read frequency was more variable. Modeling of E. coli versus lambda read frequency via cubic spline smoothing showed that the shorter genome could be used as a suitable internal spike-in for DNAs in the 200 bp to 10 kb range, allowing meaningful QC to be carried out with AAV preparations.
Head and neck squamous cell carcinomas (HNSCC) are an ideal immunotherapy target due to their high mutation burden and frequent infiltration with lymphocytes. Preclinical models to investigate ...targeted and combination therapies as well as defining biomarkers to guide treatment represent an important need in the field. Immunogenomics approaches have illuminated the role of mutation-derived tumor neoantigens as potential biomarkers of response to checkpoint blockade as well as representing therapeutic vaccines. Here, we aimed to define a platform for checkpoint and other immunotherapy studies using syngeneic HNSCC cell line models (MOC2 and MOC22), and evaluated the association between mutation burden, predicted neoantigen landscape, infiltrating T cell populations and responsiveness of tumors to anti-PD1 therapy. We defined dramatic hematopoietic cell transcriptomic alterations in the MOC22 anti-PD1 responsive model in both tumor and draining lymph nodes. Using a cancer immunogenomics pipeline and validation with ELISPOT and tetramer analysis, we identified the H-2Kb-restricted ICAM1
(mICAM1) as a neoantigen in MOC22. Finally, we demonstrated that mICAM1 vaccination was able to protect against MOC22 tumor development defining mICAM1 as a bona fide neoantigen. Together these data define a pre-clinical HNSCC model system that provides a foundation for future investigations into combination and novel therapeutics.
PDF
