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The development of a novel method, technique or technology can revolutionize the way in which we perform experiments and facilitate our understanding of fundamental issues in our field of research. For example, genome editing using the CRISPR-Cas system can lend itself to numerous cellular applications, and the development of three-dimensional cultures will advance our understanding of key developmental issues, such as tissue morphogenesis and organogenesis. The articles in this series highlight recent technological and technical advances that were developed to tackle important biological issues, and discuss the wealth of knowledge that using them has produced.
CRISPR-based genetic screens are providing new insights into the consequences of deficiencies in DNA damage response and repair pathways. These include insights into the regulation of homologous recombination and of replication stress and their crosstalk with other repair pathways, into novel cancer therapies and into the basis of cancer-drug resistance.
Mammalian RNA polymerase II transcribes protein-coding genes and non-coding transcription units, including long non-coding RNAs (lncRNAs). Studies applying recently developed nascent transcriptomics technology have revealed differences in transcription initiation and termination between lncRNAs and protein-coding genes, bearing relevance to genomic stress and DNA damage.
Recent technological breakthroughs in mapping and visualizing chromatin contacts have considerably improved our understanding of 3D genome organization and function. This Review discusses the features, strengths and limitations of various methods of genome organization analysis, including sequencing-based techniques, microscopy-based techniques and computational and modelling approaches.
CRISPR–Cas systems have revolutionized genome editing, and the CRISPR–Cas toolkit has been expanding to include single-base editing enzymes, targeting RNA and fusing inactive Cas proteins to effectors that regulate various nuclear processes. Consequently, CRISPR–Cas systems are being tested for gene and cell therapies.
Metabolites can actively regulate biological processes and may directly modulate phenotype. The current challenge of metabolomics is to provide a platform for the discovery of such bioactive metabolites and — in combination with other omics technologies — to determine their biological functions.
Spatial proteomics improves our understanding of protein function by revealing the subcellular localizations of proteins and their movement between compartments. This Review discusses spatial proteomics approaches, their successful application in cell biology and ways to improve integration of spatial proteomics data.
Taking advantage of genetic engineering, synthetic biology allows control and design of new cell functions. Recent advances in the development of genetic tools and the assembly of progressively more sophisticated gene circuits have made ‘designer cells’ a reality, with applications ranging from industry and biotechnology to medicine.
Fluorescence nanoscopy enables the optical imaging of cellular components with resolutions at the nanometre scale. With the growing availability of super-resolution microscopes, nanoscopy methods are being increasingly applied. Quantitative, multicolour, live-cell nanoscopy and the corresponding labelling strategies are under continuous development.
The three-dimensional (3D) organization of eukaryote chromosomes regulates genome function and nuclear processes such as DNA replication, transcription and DNA-damage repair. Experimental and computational methodologies for 3D genome analysis have been rapidly expanding, with a focus on high-throughput chromatin conformation capture techniques and on data analysis.
CRISPR–Cas9-based genome editing tools have been developed recently to study non-coding transcriptional regulatory elements, enabling the characterization of enhancers in their endogenous context. The applications, current limitations and future development of such CRISPR–Cas9 tools are discussed, with emphasis on identifying and characterizing enhancer elements in a high-throughput manner.
Metabolomics has been utilized extensively for the identification of single metabolites and their use as biomarkers. Owing to recent technical advances, it is now possible to use metabolomics to better understand whole metabolic pathways and to more precisely pinpoint the involvement of metabolites in physiology and pathology.
The CRISPR–Cas9 system is a powerful, sequence-specific tool that was initially developed for gene and genome editing. The recent adoption of nuclease-deactivated Cas9 (dCas9) has enabled expansion of the use of the system to multiplexed and inducible transcription regulation, genome-wide screens and cell fate engineering.
Lipids tailor membrane identities and function as molecular hubs in all cellular processes. The development of pioneering technologies, including affinity-purification lipidomics and the liposome microarray-based assay (LiMA), will enable researchers to decipher protein–lipid interactions and enhance our understanding of how lipids modulate protein function and structure.
Ribosome profiling has the power to interrogate —in vivoand on a global scale — what is being translated, how this translation is regulated and where in the cell the translation of specific sets of proteins occurs.
The unique physical and chemical properties of microfluidic devices have underpinned notable recent advances in molecular cell biology research and will facilitate a new era of biological enquiry of increased precision.
Advances in mass spectrometry-based proteomics are enabling the multidimensional analysis of protein properties such as abundance, localization, post-translational modifications and interactions for thousands of proteins. Complemented by new tools for data analysis and integration, these advances are transforming our understanding of various biological processes.
Considerable progress has been made in the past few years in our ability to visualize the structure of G protein-coupled receptors (GPCRs) and their signalling complexes. This is due to a series of technical improvements in areas such as protein engineering, lipidic cubic phase-based crystallization and microfocus synchrotron beamlines.
To achieve effective visual communication of complex data, it is important that biologists identify the needs of their audience — whether they are peers or an outreach audience. This guide to the available wealth of resources, ranging from software tools to professional illustrators, should help researchers to generate better figures and presentations tailored to the needs of any audience.
Recent advances in three-dimensional (3D) culture techniques have increased our understanding of the cellular mechanisms that drive epithelial tissue development, the genetic regulation of cell behaviours in epithelial tissues and the role of microenvironmental factors in normal development and disease. 3D culture can be used to build complex organs and to advance therapeutic approaches.
RNAi is used for genome-wide functional screens in cultured cells and animals. New experimental and bioinformatics approaches, including the combination of RNAi with genome-editing strategies, has improved the efficacy of RNAi screening and follow-up experiments, and enhanced our understanding of gene function and regulatory networks.
The optogenetic toolkit has rapidly expanded to include various proteins and cellular functions, such as cell signalling, that can be controlled by light. The practical considerations in using and deciding between optogenetic systems, such as systems that use light-oxygen-voltage (LOV) domains, phytochrome proteins, CRYPTOCHROME 2 (CRY2) and the fluorescent protein Dronpa, are now well defined.
Large-scale methodologies to facilitate the systematic measurement of protein abundance, translation level, turnover rate, post-translational modification, localization and interaction with other proteins are beginning to enable dynamic assessments of proteomes at the single-cell level.
Advances over the past decade in the development of imaging probes, microscopy techniques and image analysis have enabled researchers to gain a deeper knowledge and understanding of the dynamic processes of embryonic differentiation, patterning and morphogenesis through quantitative whole-animal imaging studies with high spatiotemporal resolution.