scispace - formally typeset
Search or ask a question
Author

Luke D. Lavis

Bio: Luke D. Lavis is an academic researcher from Howard Hughes Medical Institute. The author has contributed to research in topics: Chromatin & Fluorophore. The author has an hindex of 48, co-authored 150 publications receiving 9564 citations. Previous affiliations of Luke D. Lavis include Yale University & Janelia Farm Research Campus.


Papers
More filters
Journal ArticleDOI
TL;DR: Inspired by molecular modeling, the N,N-dimethylamino substituents in tetramethylrhodamine are replaced with four-membered azetidine rings, which doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging.
Abstract: Specific labeling of biomolecules with bright fluorophores is the keystone of fluorescence microscopy. Genetically encoded self-labeling tag proteins can be coupled to synthetic dyes inside living cells, resulting in brighter reporters than fluorescent proteins. Intracellular labeling using these techniques requires cell-permeable fluorescent ligands, however, limiting utility to a small number of classic fluorophores. Here we describe a simple structural modification that improves the brightness and photostability of dyes while preserving spectral properties and cell permeability. Inspired by molecular modeling, we replaced the N,N-dimethylamino substituents in tetramethylrhodamine with four-membered azetidine rings. This addition of two carbon atoms doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging. The novel substitution is generalizable, yielding a palette of chemical dyes with improved quantum efficiencies that spans the UV and visible range.

1,140 citations

Journal ArticleDOI
TL;DR: The chemical and photophysical properties of oft-used fluorophores are examined and classic and contemporary examples in which utility has been built upon these scaffolds are highlighted.
Abstract: Small-molecule fluorescent probes embody an essential facet of chemical biology. Although numerous compounds are known, the ensemble of fluorescent probes is based on a modest collection of modular “core” dyes. The elaboration of these dyes with diverse chemical moieties is enabling the precise interrogation of biochemical and biological systems. The importance of fluorescence-based technologies in chemical biology elicits a necessity to understand the major classes of small-molecule fluorophores. Here, we examine the chemical and photophysical properties of oft-used fluorophores and highlight classic and contemporary examples in which utility has been built upon these scaffolds.

1,086 citations

Journal ArticleDOI
27 Jul 2018-Science
TL;DR: Live-cell single-molecule imaging revealed that TF LCDs interact to form local high-concentration hubs at both synthetic DNA arrays and endogenous genomic loci, suggesting that under physiological conditions, rapid, reversible, and selective multivalent LCD-LCD interactions occur between TFs and the RNA Pol II machinery to activate transcription.
Abstract: Many eukaryotic transcription factors (TFs) contain intrinsically disordered low-complexity sequence domains (LCDs), but how these LCDs drive transactivation remains unclear. We used live-cell single-molecule imaging to reveal that TF LCDs form local high-concentration interaction hubs at synthetic and endogenous genomic loci. TF LCD hubs stabilize DNA binding, recruit RNA polymerase II (RNA Pol II), and activate transcription. LCD-LCD interactions within hubs are highly dynamic, display selectivity with binding partners, and are differentially sensitive to disruption by hexanediols. Under physiological conditions, rapid and reversible LCD-LCD interactions occur between TFs and the RNA Pol II machinery without detectable phase separation. Our findings reveal fundamental mechanisms underpinning transcriptional control and suggest a framework for developing single-molecule imaging screens for drugs targeting gene regulatory interactions implicated in disease.

710 citations

Journal ArticleDOI
TL;DR: This work refined and extended the Janelia Fluor strategy, finding that incorporation of 3-substituted azetidine groups allows rational tuning of the spectral and chemical properties of rhodamine dyes with unprecedented precision.
Abstract: Pushing the frontier of fluorescence microscopy requires the design of enhanced fluorophores with finely tuned properties. We recently discovered that incorporation of four-membered azetidine rings into classic fluorophore structures elicits substantial increases in brightness and photostability, resulting in the Janelia Fluor (JF) series of dyes. We refined and extended this strategy, finding that incorporation of 3-substituted azetidine groups allows rational tuning of the spectral and chemical properties of rhodamine dyes with unprecedented precision. This strategy allowed us to establish principles for fine-tuning the properties of fluorophores and to develop a palette of new fluorescent and fluorogenic labels with excitation ranging from blue to the far-red. Our results demonstrate the versatility of these new dyes in cells, tissues and animals.

441 citations

Journal ArticleDOI
TL;DR: This review discusses the concepts and strategies of structural and functional imaging in living cells at the single-molecule level with minimal perturbations to the specimen.

414 citations


Cited by
More filters
01 May 1993
TL;DR: Comparing the results to the fastest reported vectorized Cray Y-MP and C90 algorithm shows that the current generation of parallel machines is competitive with conventional vector supercomputers even for small problems.
Abstract: Three parallel algorithms for classical molecular dynamics are presented. The first assigns each processor a fixed subset of atoms; the second assigns each a fixed subset of inter-atomic forces to compute; the third assigns each a fixed spatial region. The algorithms are suitable for molecular dynamics models which can be difficult to parallelize efficiently—those with short-range forces where the neighbors of each atom change rapidly. They can be implemented on any distributed-memory parallel machine which allows for message-passing of data between independently executing processors. The algorithms are tested on a standard Lennard-Jones benchmark problem for system sizes ranging from 500 to 100,000,000 atoms on several parallel supercomputers--the nCUBE 2, Intel iPSC/860 and Paragon, and Cray T3D. Comparing the results to the fastest reported vectorized Cray Y-MP and C90 algorithm shows that the current generation of parallel machines is competitive with conventional vector supercomputers even for small problems. For large problems, the spatial algorithm achieves parallel efficiencies of 90% and a 1840-node Intel Paragon performs up to 165 faster than a single Cray C9O processor. Trade-offs between the three algorithms and guidelines for adapting them to more complex molecular dynamics simulations are also discussed.

29,323 citations

28 Jul 2005
TL;DR: PfPMP1)与感染红细胞、树突状组胞以及胎盘的单个或多个受体作用,在黏附及免疫逃避中起关键的作�ly.
Abstract: 抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1(PfPMP1)与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用,在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员,通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

18,940 citations

01 Jan 2016
TL;DR: The principles of fluorescence spectroscopy is universally compatible with any devices to read and is available in the digital library an online access to it is set as public so you can download it instantly.
Abstract: Thank you very much for downloading principles of fluorescence spectroscopy. As you may know, people have look hundreds times for their favorite novels like this principles of fluorescence spectroscopy, but end up in malicious downloads. Rather than reading a good book with a cup of tea in the afternoon, instead they cope with some harmful bugs inside their desktop computer. principles of fluorescence spectroscopy is available in our digital library an online access to it is set as public so you can download it instantly. Our digital library spans in multiple locations, allowing you to get the most less latency time to download any of our books like this one. Kindly say, the principles of fluorescence spectroscopy is universally compatible with any devices to read.

2,960 citations

Journal ArticleDOI
22 Sep 2017-Science
TL;DR: The findings together suggest that several membrane-less organelles have been shown to exhibit a concentration threshold for assembly, a hallmark of phase separation, and represent liquid-phase condensates, which form via a biologically regulated (liquid-liquid) phase separation process.
Abstract: BACKGROUND Living cells contain distinct subcompartments to facilitate spatiotemporal regulation of biological reactions. In addition to canonical membrane-bound organelles such as secretory vesicles and endoplasmic reticulum, there are many organelles that do not have an enclosing membrane yet remain coherent structures that can compartmentalize and concentrate specific sets of molecules. Examples include assemblies in the nucleus such as the nucleolus, Cajal bodies, and nuclear speckles and also cytoplasmic structures such as stress granules, P-bodies, and germ granules. These structures play diverse roles in various biological processes and are also increasingly implicated in protein aggregation diseases. ADVANCES A number of studies have shown that membrane-less assemblies exhibit remarkable liquid-like features. As with conventional liquids, they typically adopt round morphologies and coalesce into a single droplet upon contact with one another and also wet intracellular surfaces such as the nuclear envelope. Moreover, component molecules exhibit dynamic exchange with the surrounding nucleoplasm and cytoplasm. These findings together suggest that these structures represent liquid-phase condensates, which form via a biologically regulated (liquid-liquid) phase separation process. Liquid phase condensation increasingly appears to be a fundamental mechanism for organizing intracellular space. Consistent with this concept, several membrane-less organelles have been shown to exhibit a concentration threshold for assembly, a hallmark of phase separation. At the molecular level, weak, transient interactions between molecules with multivalent domains or intrinsically disordered regions (IDRs) are a driving force for phase separation. In cells, condensation of liquid-phase assemblies can be regulated by active processes, including transcription and various posttranslational modifications. The simplest physical picture of a homogeneous liquid phase is often not enough to capture the full complexity of intracellular condensates, which frequently exhibit heterogeneous multilayered structures with partially solid-like characters. However, recent studies have shown that multiple distinct liquid phases can coexist and give rise to richly structured droplet architectures determined by the relative liquid surface tensions. Moreover, solid-like phases can emerge from metastable liquid condensates via multiple routes of potentially both kinetic and thermodynamic origins, which has important implications for the role of intracellular liquids in protein aggregation pathologies. OUTLOOK The list of intracellular assemblies driven by liquid phase condensation is growing rapidly, but our understanding of their sequence-encoded biological function and dysfunction lags behind. Moreover, unlike equilibrium phases of nonliving matter, living cells are far from equilibrium, with intracellular condensates subject to various posttranslational regulation and other adenosine triphosphate–dependent biological activity. Efforts using in vitro reconstitution, combined with traditional cell biology approaches and quantitative biophysical tools, are required to elucidate how such nonequilibrium features of living cells control intracellular phase behavior. The functional consequences of forming liquid condensates are likely multifaceted and may include facilitated reaction, sequestration of specific factors, and organization of associated intracellular structures. Liquid phase condensation is particularly interesting in the nucleus, given the growing interest in the impact of nuclear phase behavior on the flow of genetic information; nuclear condensates range from micrometer-sized bodies such as the nucleolus to submicrometer structures such as transcriptional assemblies, all of which directly interact with and regulate the genome. Deepening our understanding of these intracellular states of matter not only will shed light on the basic biology of cellular organization but also may enable therapeutic intervention in protein aggregation disease by targeting intracellular phase behavior.

2,432 citations