Chemical genetics: tailoring tools for cell biology

Chemical genetics is a research approach that uses small molecules as probes to study protein functions in cells or whole organisms. Here, I review the parallels between classical genetic and chemical-genetic approaches and discuss the merits of small molecules to dissect dynamic cellular processes. I then consider the pros and cons of different screening approaches and specify strategies aimed at identifying and validating cellular target proteins. Finally, I highlight the impact of chemical genetics on our current understanding of cell biology and its potential for the future.     

不同结构类型糖类物质的荧光标记策略

为阐明糖链结构与功能的关系,寻找高灵敏度和高分离度的微量分析方法对糖 类物质的分析至关重要.近几年来,荧光标记方法作为糖类物质的色谱定量及辅助结 构分析的最佳选择,日益受到人们的广泛关注.荧光标记法分为柱前标记和柱后标记 两种,且灵敏度高、分离度好、有多种标记试剂可供选择.本文根据不同糖类物质的 结构特点,分别对中性糖和氨基糖以及含有唾液酸、糖醛酸、硫酸基的酸性糖的荧光 标记方法及其应用进行了系统的综述,并对近年来报道的荧光标记试剂的特点和反应 机理进行了比较和总结,以期为糖类物质微量分析方法的研究提供参考.     

Chemical biology

A selection of interesting papers that were published in the two months before our press date in major journals most likely to report significant results in chemical biology.     

Chemical biology

A selection of interesting papers that were published in the two months before our press date in major journals most likely to report significant results in chemical biology.     

Chemical biology

A selection of interesting papers that were published in the two months before our press date in major journals most likely to report significant results in chemical biology.     

Chemical biology

A selection of interesting papers that were published in the two months before our press date in major journals most likely to report significant results in chemical biology.     

Chemical biology

A selection of interesting papers that were published in the two months before our press date in major journals most likely to report significant results in chemical biology.     

Chemical biology

A selection of interesting papers that were published in the two months before our press date in major journals most likely to report significant results in chemical biology.     

Evolving strategies for the incorporation of bioinformatics within the undergraduate cell biology curriculum

Recent advances in genomics and structural biology have resulted in an unprecedented increase in biological data available from Internet-accessible databases. In order to help students effectively use this vast repository of information, undergraduate biology students at Drake University were introduced to bioinformatics software and databases in three courses, beginning with an introductory course in cell biology. The exercises and projects that were used to help students develop literacy in bioinformatics are described. In a recently offered course in bioinformatics, students developed their own simple sequence analysis tool using the Perl programming language. These experiences are described from the point of view of the instructor as well as the students. A preliminary assessment has been made of the degree to which students had developed a working knowledge of bioinformatics concepts and methods. Finally, some conclusions have been drawn from these courses that may be helpful to instructors wishing to introduce bioinformatics within the undergraduate biology curriculum.     

Chemical dissection of the cell cycle: probes for cell biology and anti-cancer drug development

Cancer cell proliferation relies on the ability of cancer cells to grow, transition through the cell cycle, and divide. To identify novel chemical probes for dissecting the mechanisms governing cell cycle progression and cell division, and for developing new anti-cancer therapeutics, we developed and performed a novel cancer cell-based high-throughput chemical screen for cell cycle modulators. This approach identified novel G1, S, G2, and M-phase specific inhibitors with drug-like properties and diverse chemotypes likely targeting a broad array of processes. We further characterized the M-phase inhibitors and highlight the most potent M-phase inhibitor MI-181, which targets tubulin, inhibits tubulin polymerization, activates the spindle assembly checkpoint, arrests cells in mitosis, and triggers a fast apoptotic cell death. Importantly, MI-181 has broad anti-cancer activity, especially against BRAF^V600E melanomas.The cell cycle is a set of coordinated events that culminate in the formation of two cells from one mother cell. It''s composed of four major phases; G1 (growth phase 1), S (DNA synthesis phase), G2 (growth phase 2) and M (mitosis), which function to integrate environment sensing signaling pathways with cell growth and proliferation.¹ Cancer cells often deregulate the cell cycle and undergo unscheduled cell divisions, therefore inhibition of the cell cycle represents an opportunity for therapeutic intervention in treating proliferative diseases like cancer.² Most anti-cancer drugs perturb the proliferation cycle of tumor cells by inhibiting/damaging cell cycle events, which activate checkpoints, arrest cells and induce apoptosis.³ For example, inhibitors targeting DNA replication (5-fluorouracil) and cell division (microtubule-stabilizing paclitaxel) have been used successfully for treating a broad array of cancers including breast and colorectal cancers.² Nevertheless, due to toxicity issues, drugs targeting the cell division machinery like mitotic kinases (AurKA/B and Plk1) and kinesins (Kif11 and CENP-E) have been developed.³ However, these drugs have shown limited efficacy in vivo.⁴ Thus, there is a critical need to identify novel drug-like molecules that inhibit cancer cell cycle progression, which can be developed into novel cancer therapies.Genome wide studies aimed at depleting the expression of human genes and characterizing their contribution to cell cycle progression have generated a wealth of information regarding the enzymatic machinery required for proliferation.⁵ These enzymes have become the focus of targeted screening campaigns aimed at finding inhibitors to their activities. For example, an in vitro chemical screen targeting Plk1 identified the small molecule BI2536.⁶ BI2536 was not only used to define novel roles for Plk1 during cell division, it was further developed into an anti-cancer drug whose efficacy is being evaluated in clinical trials.⁷ Therefore, beyond their therapeutic potential, inhibitors can be used as molecular probes for dissecting the function of enzymes critical for cell cycle progression in an acute and temporal manner. However, there are no inhibitors to the majority of the cell cycle machinery and the discovery and characterization of such inhibitors would aid our ability to understand the mechanisms regulating cell division.Although molecularly targeted screens have grown in popularity, they rely on the previous identification and validation of specific cancer targets with druggable activities/interactions.⁸ As an alternative, unbiased high-throughput chemical screens have tried to identify inhibitors to a single cell cycle phase,^{9, 10, 11, 12, 13, 14, 15} which limited their ability to identify novel anti-proliferative agents to other phases of the cell cycle. Nonetheless, G2-phase, M-phase, and cytokinesis screens successfully identified inhibitors to Kif11, Plk1, RhoA, and microtubules.^{9, 10, 11, 12, 13, 14, 15} These inhibitors aided the functional characterization of these proteins and were instrumental for developing drugs with therapeutic potential. However, these screens were conducted with a limited number of compounds (100–38 000) or cell extract fractions, with several screens using the same library of 16 320 compounds, thus limiting compound diversity, chemical coverage, and opportunities for novel discoveries. Most screens also lacked chemical analyses to understand the physiochemical properties of bioactive compounds and their cellular targets. In addition, previous screens have not analyzed the four phases of the cell cycle as a biological system. Thus, there is a critical need to develop new screening strategies to discover novel anti-cancer drugs.This, prompted us to establish an integrated high-throughput screening cell-based strategy for identifying small molecule cell cycle modulators, for use in dissecting the mechanisms of cancer cell division, and for developing novel cancer therapies. We report the development of this novel cell-based screening platform, the discovery of cell cycle phase specific inhibitors, the chemical analyses of these inhibitors, the cell culture characterization of cell division inhibitors, and the detailed examination of MI-181, which has potent anti-cancer activity, especially against melanomas.     

Isotope labeling strategies for NMR studies of RNA

The known biological functions of RNA have expanded in recent years and now include gene regulation, maintenance of sub-cellular structure, and catalysis, in addition to propagation of genetic information. As for proteins, RNA function is tightly correlated with structure. Unlike proteins, structural information for larger, biologically functional RNAs is relatively limited. NMR signal degeneracy, relaxation problems, and a paucity of long-range ¹H–¹H dipolar contacts have limited the utility of traditional NMR approaches. Selective isotope labeling, including nucleotide-specific and segmental labeling strategies, may provide the best opportunities for obtaining structural information by NMR. Here we review methods that have been developed for preparing and purifying isotopically labeled RNAs, as well as NMR strategies that have been employed for signal assignment and structure determination.     

Chemical strategies for functional proteomics

With complete genome sequences now available for several prokaryotic and eukaryotic organisms, biological researchers are charged with the task of assigning molecular and cellular functions to thousands of predicted gene products. To address this problem, the field of proteomics seeks to develop and apply methods for the global analysis of protein expression and protein function. Here we review a promising new class of proteomic strategies that utilizes synthetic chemistry to create tools and assays for the characterization of protein samples of high complexity. These approaches include the development of chemical affinity tags to measure the relative expression level and post-translational modification state of proteins in cell and tissue proteomes. Additionally, we discuss the emerging field of activity-based protein profiling, which aims to synthesize and apply small molecule probes that monitor dynamics in protein function in complex proteomes.     

Synthetic biology: new strategies for directing design

The advancement of synthetic biology is thanks, in large part, to continuing improvements in DNA synthesis. The expansion of synthetic biology into the realm of metabolic engineering has shifted the focus from simply making novel synthetic biological parts to answering the question of how we employ these biological parts to construct genomes that ultimately give rise to useful phenotypes. Much like protein engineering, the answer to this will be arrived at following the combination of rational design and evolutionary approaches. This review will highlight some of the new DNA synthesis-enabled search methods and discuss the application of such methods to the creation of synthetic gene networks and genomes.     

Chemical strategies for immobilization of oligonucleotides

The development of oligonucleotide-based microarrays (biochips) is a major thrust area in the rapidly growing biotechnology industry, which encompasses a diverse range of research areas including genomics, proteomics, computational biology, and pharmaceuticals, among other activities. Microarray experiments have proved to be unique in offering cost-effective and efficient analysis at the genomic level. In the last few years, biochips have gained increasing acceptance in the study of genetic and cellular processes. As the increase in experimental throughput has posed many challenges to the research community, considerable progress has been made in the advancement of microarray technology. In this review, chemical strategies for immobilization of oligonucleotides have been highlighted with special emphasis on post-synthetic immobilization of oligonucleotides on glass surface. The major objective of this article is to make the researchers acquainted with some most recent advances in this area.