| 1. |
|
|
| 2. |
|
|
| 3. |
- Beal, Jacob, et al.
(author)
-
Robust estimation of bacterial cell count from optical density
- 2020
-
In: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
-
Journal article (peer-reviewed)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
|
|
| 4. |
- Klionsky, Daniel J., et al.
(author)
-
Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
-
In: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
-
Research review (peer-reviewed)abstract
- In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.
|
|
| 5. |
- Campbell, PJ, et al.
(author)
-
Pan-cancer analysis of whole genomes
- 2020
-
In: Nature. - : Springer Science and Business Media LLC. - 1476-4687 .- 0028-0836. ; 578:7793, s. 82-
-
Journal article (peer-reviewed)abstract
- Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale1–3. Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4–5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter4; identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation5,6; analyses timings and patterns of tumour evolution7; describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity8,9; and evaluates a range of more-specialized features of cancer genomes8,10–18.
|
|
| 6. |
|
|
| 7. |
- Kanoni, Stavroula, et al.
(author)
-
Implicating genes, pleiotropy, and sexual dimorphism at blood lipid loci through multi-ancestry meta-analysis.
- 2022
-
In: Genome biology. - : Springer Science and Business Media LLC. - 1474-760X .- 1465-6906 .- 1474-7596. ; 23:1
-
Journal article (peer-reviewed)abstract
- Genetic variants within nearly 1000 loci are known to contribute to modulation of blood lipid levels. However, the biological pathways underlying these associations are frequently unknown, limiting understanding of these findings and hindering downstream translational efforts such as drug target discovery.To expand our understanding of the underlying biological pathways and mechanisms controlling blood lipid levels, we leverage a large multi-ancestry meta-analysis (N = 1,654,960) of blood lipids to prioritize putative causal genes for 2286 lipid associations using six gene prediction approaches. Using phenome-wide association (PheWAS) scans, we identify relationships of genetically predicted lipid levels to other diseases and conditions. We confirm known pleiotropic associations with cardiovascular phenotypes and determine novel associations, notably with cholelithiasis risk. We perform sex-stratified GWAS meta-analysis of lipid levels and show that 3-5% of autosomal lipid-associated loci demonstrate sex-biased effects. Finally, we report 21 novel lipid loci identified on the X chromosome. Many of the sex-biased autosomal and X chromosome lipid loci show pleiotropic associations with sex hormones, emphasizing the role of hormone regulation in lipid metabolism.Taken together, our findings provide insights into the biological mechanisms through which associated variants lead to altered lipid levels and potentially cardiovascular disease risk.
|
|
| 8. |
- Yu, ChaoQing, et al.
(author)
-
Managing nitrogen to restore water quality in China
- 2019
-
In: Nature. - : Nature Publishing Group. - 0028-0836 .- 1476-4687. ; 567:7749, s. 516-520
-
Journal article (peer-reviewed)abstract
- The nitrogen cycle has been radically changed by human activities(1). China consumes nearly one third of the world's nitrogen fertilizers. The excessive application of fertilizers(2,3) and increased nitrogen discharge from livestock, domestic and industrial sources have resulted in pervasive water pollution. Quantifying a nitrogen 'boundary'(4) in heterogeneous environments is important for the effective management of local water quality. Here we use a combination of water-quality observations and simulated nitrogen discharge from agricultural and other sources to estimate spatial patterns of nitrogen discharge into water bodies across China from 1955 to 2014. We find that the critical surface-water quality standard (1.0 milligrams of nitrogen per litre) was being exceeded in most provinces by the mid-1980s, and that current rates of anthropogenic nitrogen discharge (14.5 +/- 3.1 megatonnes of nitrogen per year) to fresh water are about 2.7 times the estimated 'safe' nitrogen discharge threshold (5.2 +/- 0.7 megatonnes of nitrogen per year). Current efforts to reduce pollution through wastewater treatment and by improving cropland nitrogen management can partially remedy this situation. Domestic wastewater treatment has helped to reduce net discharge by 0.7 +/- 0.1 megatonnes in 2014, but at high monetary and energy costs. Improved cropland nitrogen management could remove another 2.3 +/- 0.3 megatonnes of nitrogen per year-about 25 per cent of the excess discharge to fresh water. Successfully restoring a clean water environment in China will further require transformational changes to boost the national nutrient recycling rate from its current average of 36 per cent to about 87 per cent, which is a level typical of traditional Chinese agriculture. Although ambitious, such a high level of nitrogen recycling is technologically achievable at an estimated capital cost of approximately 100 billion US dollars and operating costs of 18-29 billion US dollars per year, and could provide co-benefits such as recycled wastewater for crop irrigation and improved environmental quality and ecosystem services.
|
|
| 9. |
- Zhang, Jian, et al.
(author)
-
Optimization of N-Substituted Oseltamivir Derivatives as Potent Inhibitors of Group-1 and-2 Influenza A Neuraminidases, Including a Drug-Resistant Variant
- 2018
-
In: Journal of Medicinal Chemistry. - : American Chemical Society (ACS). - 0022-2623 .- 1520-4804. ; 61:14, s. 6379-6397
-
Journal article (peer-reviewed)abstract
- On the basis of our earlier discovery of N1-selective inhibitors, the 150-cavity of influenza virus neuraminidases (NAs) could be further exploited to yield more potent oseltamivir derivatives. Among the synthesized compounds, 15b and 15c were exceptionally active against both group-1 and -2 NAs. Especially for 09N1, N2, N6, and N9 subtypes, they showed 6.80-12.47 and 1.20-3.94 times greater activity than oseltamivir carboxylate (OSC). They also showed greater inhibitory activity than OSC toward H274Y and E119V variant. In cellular assays, they exhibited greater potency than OSC toward H5N1, H5N2, H5N6, and H5N8 viruses. 15b demonstrated high metabolic stability, low cytotoxicity in vitro, and low acute toxicity in mice. Computational modeling and molecular dynamics studies provided insights into the role of R group of 15b in improving potency toward group-1 and -2 NAs. We believe the successful exploitation of the 150-cavity of NAs represents an important breakthrough in the development of more potent anti-influenza agents.
|
|
| 10. |
|
|
