Monitoring the autophagy pathway in cancer.

abstract：:Microscopy is a simple, direct technique for examining the morphology of cells and their organelles. Embedding cells in agarose and then in paraffin as 'cell blocks' allows for them to be processed in the same manner that tissue specimens are processed for histology. This method is advantageous because numerous sectio...

journal_title：Methods in enzymology

authors： Koh CM

更新日期：2013-01-01 00:00:00

abstract：:Autophagy is a catabolic process that is crucial for cellular homeostasis and adaptive response to changing environments. Importantly, autophagy has been shown to be induced in many longevity-associated scenarios and to be required to maintain lifespan extension. Notably, autophagy is a highly conserved cellular proce...

journal_title：Methods in enzymology

authors： Kainz K,Tadic J,Zimmermann A,Pendl T,Carmona-Gutierrez D,Ruckenstuhl C,Eisenberg T,Madeo F

更新日期：2017-01-01 00:00:00

abstract：:An optimal tool to unravel the role of a specific player within a cellular network or process requires its spatiotemporally resolved perturbation. Chemically induced dimerization (CID) by the rapamycin system has proven useful to induce protein dimerization or translocation with high spatiotemporal precision. Recently...

journal_title：Methods in enzymology

authors： Schifferer M,Feng S,Stein F,Schultz C

更新日期：2017-01-01 00:00:00

abstract：:Epitope tagged alpha 2-AR subtypes have been used to address a variety of cell biological questions, and the strategies used are readily applicable to all GPCR as well as other cell surface proteins. We have provided detailed protocols for successful utilization of the epitope-tagged receptor in the studies of protein...

journal_title：Methods in enzymology

authors： Wozniak M,Saunders C,Schramm N,Keefer JR,Limbird LE

更新日期：2002-01-01 00:00:00

abstract：:The aim of this chapter is to discuss the basic principles and reasoning behind elementary flux mode analysis (EFM analysis)--an important tool for the analysis of metabolic networks. We begin with a short introduction into metabolic pathway analysis and subsequently outline in detail fundamentals of EFM analysis by w...

journal_title：Methods in enzymology

authors： Schäuble S,Schuster S,Kaleta C

更新日期：2011-01-01 00:00:00

abstract：:Transfer RNA is extensively modified by the actions of a variety of enzymes. The radical S-adenosyl-l-methionine enzyme TYW1 modifies tRNAPhe forming the characteristic tricyclic ring via the condensation of carbons 2 and 3 of pyruvate. This chapter details methods that are required for studies of TYW1. ...

journal_title：Methods in enzymology

authors： Young AP,Bandarian V

更新日期：2018-01-01 00:00:00

abstract：:Desmoplakin (DP) is an obligate component of desmosomes, where it links the desmosomal cadherin/plakoglobin/plakophilin assembly to intermediate filaments. DP contains a large amino-terminal domain (DPNT) that binds to the cadherin/plakoglobin/plakophilin complex, a central coiled-coil domain that dimerizes the molecu...

journal_title：Methods in enzymology

authors： Choi HJ,Weis WI

更新日期：2016-01-01 00:00:00

abstract：:A synthetic ligand regulable system for gene transfer and expression has been developed in our laboratory based on mechanistic studies of steriod hormone receptor and transcriptional regulation. This gene switch system possesses most of the important features that are required for application of the system in biologic...

journal_title：Methods in enzymology

authors： Ye X,Schillinger K,Burcin MM,Tsai SY,O'Malley BW

更新日期：2002-01-01 00:00:00

abstract：:Retinoic acid (RA), the biologically active metabolite of vitamin A, regulates a vast spectrum of biological processes, such as cell differentiation, proliferation, apoptosis, and morphogenesis. microRNAs (miRNAs) play a crucial role in regulating gene expression by binding to messenger RNA (mRNA) which leads to mRNA ...

journal_title：Methods in enzymology

authors： Wang L,Rohatgi AP,Wan YY

更新日期：2020-01-01 00:00:00

abstract：:Growing appreciation for the biogeochemical significance of uncultured microorganisms is changing the focus of environmental microbiology. Techniques designed to investigate microbial metabolism in situ are increasingly popular, from mRNA-targeted fluorescence in situ hybridization (FISH) to the "-omics" revolution, i...

journal_title：Methods in enzymology

authors： Dekas AE,Orphan VJ

更新日期：2011-01-01 00:00:00

abstract：:This chapter presents methodologies for RNA extraction from soils coupled with competitive reverse transcription-polymerase chain reaction and capillary electrophoresis techniques. Combined, these approaches provide new capabilities to quantify gene expression in different environments and can aid our understanding of...

journal_title：Methods in enzymology

authors： Semrau JD,Han JI

更新日期：2005-01-01 00:00:00

abstract：:The anaphase-promoting complex (APC) is a central regulator of the eukaryotic cell cycle and functions as an E3 ubiquitin protein ligase to catalyze the ubiquitination of a number of cell cycle regulatory proteins. The APC contains at least 13 subunits in addition to two activator subunits, Cdc20 and Cdh1, that associ...

journal_title：Methods in enzymology

authors： Passmore LA,Barford D,Harper JW

更新日期：2005-01-01 00:00:00

abstract：:Fluorescence live-cell RNA imaging to monitor the intracellular localization and dynamics of the target RNA is a challenging subject. One of the difficulties to achieve this is to establish a precise method to enable a fluorescent labeling to the target RNA in living cells. Technologies to reduce the background fluore...

journal_title：Methods in enzymology

authors： Yoshimura H,Ozawa T

更新日期：2016-01-01 00:00:00

abstract：:Mitochondrial uncoupling protein-2 (UCP2) regulates glucose-stimulated insulin secretion (GSIS) by pancreatic beta cells-the physiological role of the beta cell UCP2 remains a subject of debate. Experimental studies informing this debate benefit from reliable measurements of UCP2 protein level and activity. In this ch...

journal_title：Methods in enzymology

authors： Barlow J,Hirschberg V,Brand MD,Affourtit C

更新日期：2013-01-01 00:00:00

abstract：:Protein matrix cavities and extended tunnels can effectively channel substrates and products to and from active sites in enzymes. Substrate and product channeling can enhance catalytic efficiency by reducing the intramolecular diffusion times to and from reaction centers. Moreover, protected transfer between sites may...

journal_title：Methods in enzymology

authors： Pesce A,Milani M,Nardini M,Bolognesi M

更新日期：2008-01-01 00:00:00

abstract：:It is widely acknowledged that mitochondria are highly active structures that rapidly respond to cellular and environmental perturbations by changing their shape, number, and distribution. Mitochondrial remodeling is a key component of diverse biological processes, ranging from cell cycle progression to autophagy. In ...

journal_title：Methods in enzymology

authors： Marchi S,Bonora M,Patergnani S,Giorgi C,Pinton P

更新日期：2017-01-01 00:00:00

abstract：:Although many players of the DNA damage response and DNA repair have been identified in several systems their biochemical role is still poorly understood. The use of the Xenopus laevis egg extract cell-free system allowed biochemical dissection of DNA replication and cell cycle events in a complex biological context. ...

journal_title：Methods in enzymology

authors： Sannino V,Pezzimenti F,Bertora S,Costanzo V

更新日期：2017-01-01 00:00:00

abstract：:Histone acetylation is a dynamic epigenetic modification that functions in the regulation of DNA-templated reactions, such as transcription. This lysine modification is reversibly controlled by histone (lysine) acetyltransferases and deacetylases. Here, we present methods employing isotopic labeling and mass spectrome...

journal_title：Methods in enzymology

authors： Fan J,Baeza J,Denu JM

更新日期：2016-01-01 00:00:00

abstract：:In human, rat, and mice, a UGT1 complex locus provides for developmental-, inducer-, and cell-specific synthesis of a family of chemical-detoxifying isozymes, UDP-glucuronosyltransferases, which prevent toxicities, mutagenesis, and/or carcinogenesis. Between 10 and 14 first exons with individual promoter elements are ...

journal_title：Methods in enzymology

authors： Owens IS,Basu NK,Banerjee R

更新日期：2005-01-01 00:00:00

abstract：:Nucleic acid aptamers find widespread use as targeting and sensing agents in nature and biotechnology. Their ability to bind an extensive range of molecular targets, including small molecules, proteins, and ions, with high affinity and specificity enables their use in diverse diagnostic, therapeutic, imaging, and gene...

journal_title：Methods in enzymology

authors： Chang AL,McKeague M,Smolke CD

更新日期：2014-01-01 00:00:00

abstract：:Acidic organelles and vesicles, such as endosomes, lysosomes, autophagosomes, trans-Golgi network, and synaptic vesicles, are known to play important roles in a broad range of cellular events. To facilitate studying these multifunctional systems, we describe here an acid-brightening fluorescent protein (abFP), which f...

journal_title：Methods in enzymology

authors： Wang N,Wang L

更新日期：2020-01-01 00:00:00

abstract：:In eukaryotes, determination of the subcellular location of a novel protein encoded in genomic or transcriptomic data provides useful clues as to its possible function. However, experimental localization studies are expensive and time-consuming. As a result, accurate in silico prediction of subcellular localization fr...

journal_title：Methods in enzymology

authors： Gaston D,Tsaousis AD,Roger AJ

更新日期：2009-01-01 00:00:00

abstract：:Advanced glycation is one of the major pathophysiological posttranslational modifications. Under hyperglycemic conditions or oxidative stress, proteins and DNA are nonenzymatically modified by oxidative glycation and converted to advanced glycation endproducts (AGEs), which induce the loss of protein functions or apop...

journal_title：Methods in enzymology

authors： Inagi R

更新日期：2011-01-01 00:00:00

abstract：:In this chapter we have detailed a method that can be generalized to link virtually any DNA substrate to a chromatography matrix at its ends via an avidin-biotin linkage. We have used this technique to construct a left-handed Z-DNA column for the purpose of identification and purification of Z-DNA-binding proteins. Th...

journal_title：Methods in enzymology

authors： Fishel R,Anziano P,Rich A

更新日期：1990-01-01 00:00:00

abstract：:The field of therapeutic application of the A₃ adenosine receptor (A₃AR) antagonists represents a rapidly growing and intense area of research in the adenosine field. Even if there are currently no A₃AR antagonists in clinical phases, in light of the plethora of biological effects attributed to A₃ARs, substantial effo...

journal_title：Methods in enzymology

authors： Paoletta S,Federico S,Spalluto G,Moro S

更新日期：2010-01-01 00:00:00

abstract：:The combination of PCR amplification and chemiluminescent detection of PCR products provides a highly sensitive system for the quantitation of DNA and RNA. The broad dynamic range of the chemiluminescent detection assay simplifies the selection of cycling and concentration parameters critical to harnessing the quantit...

journal_title：Methods in enzymology

authors： Martin CS,Voyta JC,Bronstein I

更新日期：2000-01-01 00:00:00

abstract：:Proteins are fragile molecules that often require great care during purification to ensure that they remain intact and fully active. Nowadays, many proteins are also purified in small amounts under denaturing conditions by various gel electrophoretic techniques, such that inactive proteins are obtained. But even here,...

journal_title：Methods in enzymology

authors： Deutscher MP

更新日期：2009-01-01 00:00:00

abstract：:The identification of novel forms of O-linked glycosylation on epidermal growth factor and thrombospondin type 1 repeats, and their emerging functional significance, require the development of new methods for their analysis. This chapter describes detailed methods to analyze both the structure and the site of modifica...

journal_title：Methods in enzymology

authors： Nita-Lazar A,Haltiwanger RS

更新日期：2006-01-01 00:00:00

abstract：:Here we describe an optimized recombineering protocol for the creation of conditional alleles. Recombineering is a DNA engineering methodology based on homologous recombination in Escherichia coli. It presents a number of advantages over conventional engineering methods, including the ability to place target sites for...

journal_title：Methods in enzymology

authors： Fu J,Teucher M,Anastassiadis K,Skarnes W,Stewart AF

更新日期：2010-01-01 00:00:00

abstract：:Guanine nucleotide exchange factors (GEFs) are essential regulators of the spatiotemporal conditions of small GTP-binding protein (SMG) activation. Their cellular activities combine the biochemical stimulation of GDP/GTP exchange, which leads to the active conformation of the SMG, to the detection of upstream signals ...

journal_title：Methods in enzymology

authors： Zeeh JC,Antonny B,Cherfils J,Zeghouf M

更新日期：2008-01-01 00:00:00