Showing papers by "Rockefeller University published in 2003"
TL;DR: It is suggested that several clinical situations, including autoimmunity and certain infectious diseases, can be influenced by the antigen-specific tolerogenic role of DCs.
Abstract: Dendritic cells (DCs) have several functions in innate and adaptive immunity. In addition, there is increasing evidence that DCs in situ induce antigen-specific unresponsiveness or tolerance in central lymphoid organs and in the periphery. In the thymus DCs generate tolerance by deleting self-reactive T cells. In peripheral lymphoid organs DCs also induce tolerance to antigens captured by receptors that mediate efficient uptake of proteins and dying cells. Uptake by these receptors leads to the constitutive presentation of antigens on major histocompatibility complex (MHC) class I and II products. In the steady state the targeting of DC antigen capture receptors with low doses of antigens leads to deletion of the corresponding T cells and unresponsiveness to antigenic rechallenge with strong adjuvants. In contrast, if a stimulus for DC maturation is coadministered with the antigen, the mice develop immunity, including interferon-gamma-secreting effector T cells and memory T cells. There is also new evidence that DCs can contribute to the expansion and differentiation of T cells that regulate or suppress other immune T cells. One possibility is that distinct developmental stages and subsets of DCs and T cells can account for the different pathways to peripheral tolerance, such as deletion or suppression. We suggest that several clinical situations, including autoimmunity and certain infectious diseases, can be influenced by the antigen-specific tolerogenic role of DCs.
3,082 citations
TL;DR: The results reaffirm the thesis that miRNAs have an important role in establishing the complex spatial and temporal patterns of gene activity necessary for the orderly progression of development and suggest additional roles in the function of the mature organism.
Abstract: Background: The recent discoveries of microRNA (miRNA) genes and characterization of the first few target genes regulated by miRNAs in Caenorhabditis elegans and Drosophila melanogaster have set the stage for elucidation of a novel network of regulatory control. We present a computational method for wholegenome prediction of miRNA target genes. The method is validated using known examples. For each miRNA, target genes are selected on the basis of three properties: sequence complementarity using a position-weighted local alignment algorithm, free energies of RNA-RNA duplexes, and conservation of target sites in related genomes. Application to the D. melanogaster, Drosophila pseudoobscura and Anopheles gambiae genomes identifies several hundred target genes potentially regulated by one or more known miRNAs.
2,997 citations
TL;DR: The concept of allostasis is discussed, maintaining stability through change, as a fundamental process through which organisms actively adjust to both predictable and unpredictable events, using the balance between energy input and expenditure as the basis for applying the concept.
2,782 citations
TL;DR: Two approaches presented are endocytic uptake of QDs and selective labeling of cell surface proteins with QDs conjugated to antibodies, which should permit the simultaneous study of multiple cells over long periods of time as they proceed through growth and development.
Abstract: Luminescent quantum dots (QDs)—semiconductor nanocrystals—are a promising alternative to organic dyes for fluorescence-based applications. We have developed procedures for using QDs to label live cells and have demonstrated their use for long-term multicolor imaging of live cells. The two approaches presented are (i) endocytic uptake of QDs and (ii) selective labeling of cell surface proteins with QDs conjugated to antibodies. Live cells labeled using these approaches were used for long-term multicolor imaging. The cells remained stably labeled for over a week as they grew and developed. These approaches should permit the simultaneous study of multiple cells over long periods of time as they proceed through growth and development.
2,085 citations
TL;DR: The prevalence of self-reactive antibody formation and its regulation in human B cells is determined and a majority (55 to 75%) of all antibodies expressed by early immature B cells displayedSelf-reactivity, including polyreactive and anti-nuclear specificities.
Abstract: During B lymphocyte development, antibodies are assembled by random gene segment reassortment to produce a vast number of specificities. A potential disadvantage of this process is that some of the antibodies produced are self-reactive. We determined the prevalence of self-reactive antibody formation and its regulation in human B cells. A majority (55 to 75%) of all antibodies expressed by early immature B cells displayed self-reactivity, including polyreactive and anti-nuclear specificities. Most of these autoantibodies were removed from the population at two discrete checkpoints during B cell development. Inefficient checkpoint regulation would lead to substantial increases in circulating autoantibodies.
1,863 citations
TL;DR: The structure of KvAP, a voltage-dependent K+ channel from Aeropyrum pernix, is presented and a crystal structure of the full-length channel at a resolution of 3.2 Å is determined, which suggests that the voltage-sensor paddles move in response to membrane voltage changes, carrying their positive charge across the membrane.
Abstract: Voltage-dependent K+ channels are members of the family of voltage-dependent cation (K+, Na+ and Ca2+) channels that open and allow ion conduction in response to changes in cell membrane voltage. This form of gating underlies the generation of nerve and muscle action potentials, among other processes. Here we present the structure of KvAP, a voltage-dependent K+ channel from Aeropyrum pernix. We have determined a crystal structure of the full-length channel at a resolution of 3.2 A, and of the isolated voltage-sensor domain at 1.9 A, both in complex with monoclonal Fab fragments. The channel contains a central ion-conduction pore surrounded by voltage sensors, which form what we call 'voltage-sensor paddles'-hydrophobic, cationic, helix-turn-helix structures on the channel's outer perimeter. Flexible hinges suggest that the voltage-sensor paddles move in response to membrane voltage changes, carrying their positive charge across the membrane.
1,758 citations
Massachusetts Institute of Technology1, University of California, Riverside2, University of Oregon3, University of Edinburgh4, Celera Corporation5, Texas A&M University6, University of New Mexico7, University of Colorado Denver8, University of Kansas9, University of Florida10, Hebrew University of Jerusalem11, University of Düsseldorf12, Rockefeller University13, Technische Universität München14, University of Kentucky15, University of Texas at Austin16, J. Craig Venter Institute17, University of California, Berkeley18, University of California, Los Angeles19, Sapienza University of Rome20, Flinders University21, Ohio State University22, University of Arizona23, University of Missouri–Kansas City24, Dartmouth College25, University of Leeds26, California State Polytechnic University, Pomona27, Oregon Health & Science University28
TL;DR: A high-quality draft sequence of the N. crassa genome is reported, suggesting that RIP has had a profound impact on genome evolution, greatly slowing the creation of new genes through genomic duplication and resulting in a genome with an unusually low proportion of closely related genes.
Abstract: Neurospora crassa is a central organism in the history of twentieth-century genetics, biochemistry and molecular biology. Here, we
report a high-quality draft sequence of the N. crassa genome. The approximately 40-megabase genome encodes about 10,000
protein-coding genes—more than twice as many as in the fission yeast Schizosaccharomyces pombe and only about 25% fewer
than in the fruitfly Drosophila melanogaster. Analysis of the gene set yields insights into unexpected aspects of Neurospora biology
including the identification of genes potentially associated with red light photobiology, genes implicated in secondary metabolism,
and important differences in Ca21 signalling as compared with plants and animals. Neurospora possesses the widest array of
genome defence mechanisms known for any eukaryotic organism, including a process unique to fungi called repeat-induced
point mutation (RIP). Genome analysis suggests that RIP has had a profound impact on genome evolution, greatly slowing the
creation of new genes through genomic duplication and resulting in a genome with an unusually low proportion of closely related
genes.
1,659 citations
TL;DR: Using electrophysiological recordings, ghrelin stimulated the activity of arcuate NPY neurons and mimicked the effect of NPY in the paraventricular nucleus of the hypothalamus (PVH), thus representing a novel regulatory circuit controlling energy homeostasis.
1,578 citations
TL;DR: The cellular response to telomere dysfunction is governed by proteins that also control the DNA damage response, and induction of TIFs through TRF2 inhibition provides an opportunity to study theDNA damage response within the context of well-defined, physically marked lesions.
1,370 citations
TL;DR: The idea that the in vivo action of the different factors in E-cadherin repression can be modulated by their relative concentrations as well as by specific cellular or tumour contexts is supported.
Abstract: Transcriptional repression mechanisms have emerged as one of the crucial processes for the downregulation of E-cadherin expression during development and tumour progression. Recently, several E-cadherin transcriptional repressors have been characterized (Snail, E12/E47, ZEB-1 and SIP-1) and shown to act through an interaction with proximal E-boxes of the E-cadherin promoter. We have analyzed the participation of another member of the Snail family, Slug, and observed that it also behaves as a repressor of E-cadherin expression. Stable expression of Slug in MDCK cells leads to the full repression of E-cadherin at transcriptional level and triggers a complete epithelial to mesenchymal transition. Slug-induced repression of E-cadherin is mediated by its binding to proximal E-boxes, particularly to the E-pal element of the mouse promoter. Detailed analysis of the binding affinity of different repressors to the E-pal element indicates that Slug binds with lower affinity than Snail and E47 proteins. These results, together with the known expression patterns of these factors in embryonic development and carcinoma cell lines, support the idea that the in vivo action of the different factors in E-cadherin repression can be modulated by their relative concentrations as well as by specific cellular or tumour contexts.
1,138 citations
TL;DR: CLIP reveals that Nova coordinately regulates a biologically coherent set of RNAs encoding multiple components of the inhibitory synapse, an observation that may relate to the cause of abnormal motor inhibition in POMA.
Abstract: Nova proteins are neuron-specific antigens targeted in paraneoplastic opsoclonus myoclonus ataxia (POMA), an autoimmune neurologic disease characterized by abnormal motor inhibition Nova proteins regulate neuronal pre-messenger RNA splicing by directly binding to RNA To identify Nova RNA targets, we developed a method to purify protein-RNA complexes from mouse brain with the use of ultraviolet cross-linking and immunoprecipitation (CLIP)Thirty-four transcripts were identified multiple times by Nova CLIPThree-quarters of these encode proteins that function at the neuronal synapse, and one-third are involved in neuronal inhibitionSplicing targets confirmed in Nova-/- mice include c-Jun N-terminal kinase 2, neogenin, and gephyrin; the latter encodes a protein that clusters inhibitory gamma-aminobutyric acid and glycine receptors, two previously identified Nova splicing targetsThus, CLIP reveals that Nova coordinately regulates a biologically coherent set of RNAs encoding multiple components of the inhibitory synapse, an observation that may relate to the cause of abnormal motor inhibition in POMA
TL;DR: The amygdala becomes hyperactive in posttraumatic stress disorder (PTSD) and depressive illness, and hypertrophy of amygdala nerve cells is reported after repeated stress in an animal model.
TL;DR: It is shown that the chromodomain proteins Polycomb (Pc) and HP1 (heterochromatin protein 1) are highly discriminatory for binding to these sites in vivo and in vitro, and a role for their Chromodomains in both target site binding and discrimination is indicated.
Abstract: On the histone H3 tail, Lys 9 and Lys 27 are both methylation sites associated with epigenetic repression, and reside within a highly related sequence motif ARKS. Here we show that the chromodomain proteins Polycomb (Pc) and HP1 (heterochromatin protein 1) are highly discriminatory for binding to these sites in vivo and in vitro. In Drosophila S2 cells, and on polytene chromosomes, methyl-Lys 27 and Pc are both excluded from areas that are enriched in methyl-Lys 9 and HP1. Swapping of the chromodomain regions of Pc and HP1 is sufficient for switching the nuclear localization patterns of these factors, indicating a role for their chromodomains in both target site binding and discrimination. To better understand the molecular basis for the selection of methyl-lysine binding sites, we solved the 1.8 A structure of the Pc chromodomain in complex with a H3 peptide bearing trimethyl-Lys 27, and compared it with our previously determined structure of the HP1 chromodomain in complex with a H3 peptide bearing trimethyl-Lys 9. The Pc chromodomain distinguishes its methylation target on the H3 tail via an extended recognition groove that binds five additional residues preceding the ARKS motif.
TL;DR: The small RNA profile of Drosophila melanogaster is described as a function of development and 178 repeat-associated small interfering RNAs (rasiRNAs) are isolated, suggesting that small RNAs participate in defining chromatin structure.
TL;DR: It is found that CD25+ CD4+ T cells can proliferate in the absence of added cytokines in culture and in vivo when stimulated by antigen-loaded dendritic cells (DCs), especially mature DCs.
Abstract: An important pathway for immune tolerance is provided by thymic-derived CD25+ CD4+ T cells that suppress other CD25− autoimmune disease–inducing T cells. The antigen-presenting cell (APC) requirements for the control of CD25+ CD4+ suppressor T cells remain to be identified, hampering their study in experimental and clinical situations. CD25+ CD4+ T cells are classically anergic, unable to proliferate in response to mitogenic antibodies to the T cell receptor complex. We now find that CD25+ CD4+ T cells can proliferate in the absence of added cytokines in culture and in vivo when stimulated by antigen-loaded dendritic cells (DCs), especially mature DCs. With high doses of DCs in culture, CD25+ CD4+ and CD25− CD4+ populations initially proliferate to a comparable extent. With current methods, one third of the antigen-reactive T cell receptor transgenic T cells enter into cycle for an average of three divisions in 3 d. The expansion of CD25+ CD4+ T cells stops by day 5, in the absence or presence of exogenous interleukin (IL)-2, whereas CD25− CD4+ T cells continue to grow. CD25+ CD4+ T cell growth requires DC–T cell contact and is partially dependent upon the production of small amounts of IL-2 by the T cells and B7 costimulation by the DCs. After antigen-specific expansion, the CD25+ CD4+ T cells retain their known surface features and actively suppress CD25− CD4+ T cell proliferation to splenic APCs. DCs also can expand CD25+ CD4+ T cells in the absence of specific antigen but in the presence of exogenous IL-2. In vivo, both steady state and mature antigen-processing DCs induce proliferation of adoptively transferred CD25+ CD4+ T cells. The capacity to expand CD25+ CD4+ T cells provides DCs with an additional mechanism to regulate autoimmunity and other immune responses.
TL;DR: Dengue virus is a single-stranded, enveloped RNA virus that productively infects human dendritic cells primarily at the immature stage of their differentiation and DC-SIGN, a C-type lectin, may be considered as a new target for designing therapies that block dengue infection.
Abstract: Dengue virus is a single-stranded, enveloped RNA virus that productively infects human dendritic cells (DCs) primarily at the immature stage of their differentiation. We now find that all four serotypes of dengue use DC-SIGN (CD209), a C-type lectin, to infect dendritic cells. THP-1 cells become susceptible to dengue infection after transfection of DC-specific ICAM-3 grabbing nonintegrin (DC-SIGN), or its homologue L-SIGN, whereas the infection of dendritic cells is blocked by anti–DC-SIGN antibodies and not by antibodies to other molecules on these cells. Viruses produced by dendritic cells are infectious for DC-SIGN– and L-SIGN–bearing THP-1 cells and other permissive cell lines. Therefore, DC-SIGN may be considered as a new target for designing therapies that block dengue infection.
TL;DR: By 2050, the human population will probably be larger by 2 to 4 billion people, more slowly growing (declining in the more developed regions), more urban, especially in less developed regions, and older than in the 20th century.
Abstract: By 2050, the human population will probably be larger by 2 to 4 billion people, more slowly growing (declining in the more developed regions), more urban, especially in less developed regions, and older than in the 20th century. Two major demographic uncertainties in the next 50 years concern international migration and the structure of families. Economies, nonhuman environments, and cultures (including values, religions, and politics) strongly influence demographic changes. Hence, human choices, individual and collective, will have demographic effects, intentional or otherwise.
TL;DR: Incomplete control of HCV replication by memory CD8+ Tcells in the absence of adequate CD4+ Tcell help was associated with emergence of viral escape mutations in class I major histocompatibility complex–restricted epitopes and failure to resolve HCV infection.
Abstract: Spontaneous resolution of hepatitis C virus (HCV) infection in humans usually affords long-term immunity to persistent viremia and associated liver diseases. Here, we report that memory CD4+ Tcells are essential for this protection. Antibody-mediated depletion of CD4+ Tcells before reinfection of two immune chimpanzees resulted in persistent, low-level viremia despite functional intra-hepatic memory CD8+ Tcell responses. Incomplete control of HCV replication by memory CD8+ Tcells in the absence of adequate CD4+ Tcell help was associated with emergence of viral escape mutations in class I major histocompatibility complex-restricted epitopes and failure to resolve HCV infection.
TL;DR: The results show that newly born cells recruited into the olfactory bulb become neurons, and a unique sequence of events leads to their functional integration.
Abstract: New neurons are continually recruited throughout adulthood in certain regions of the adult mammalian brain. How these cells mature and integrate into preexisting functional circuits remains unknown. Here we describe the physiological properties of newborn olfactory bulb interneurons at five different stages of their maturation in adult mice. Patch-clamp recordings were obtained from tangentially and radially migrating young neurons and from neurons in three subsequent maturation stages. Tangentially migrating neurons expressed extrasynaptic GABAA receptors and then AMPA receptors, before NMDA receptors appeared in radially migrating neurons. Spontaneous synaptic activity emerged soon after migration was complete, and spiking activity was the last characteristic to be acquired. This delayed excitability is unique to cells born in the adult and may protect circuits from uncontrolled neurotransmitter release and neural network disruption. Our results show that newly born cells recruited into the olfactory bulb become neurons, and a unique sequence of events leads to their functional integration.
TL;DR: A sensitive imaging system in the Drosophila brain that couples two-photon microscopy with the specific expression of the calcium-sensitive fluorescent protein, G-CaMP is developed, demonstrating that the response pattern of a given glomerulus is a function of the specificity of a single odorant receptor.
TL;DR: It is concluded that the voltage-sensor paddles operate somewhat like hydrophobic cations attached to levers, enabling the membrane electric field to open and close the pore.
Abstract: The steep dependence of channel opening on membrane voltage allows voltage-dependent K+ channels to turn on almost like a switch. Opening is driven by the movement of gating charges that originate from arginine residues on helical S4 segments of the protein. Each S4 segment forms half of a ‘voltage-sensor paddle’ on the channel's outer perimeter. Here we show that the voltage-sensor paddles are positioned inside the membrane, near the intracellular surface, when the channel is closed, and that the paddles move a large distance across the membrane from inside to outside when the channel opens. KvAP channels were reconstituted into planar lipid membranes and studied using monoclonal Fab fragments, a voltage-sensor toxin, and avidin binding to tethered biotin. Our findings lead us to conclude that the voltage-sensor paddles operate somewhat like hydrophobic cations attached to levers, enabling the membrane electric field to open and close the pore.
TL;DR: 6-bromoindirubins provide a new scaffold for the development of selective and potent pharmacological inhibitors of GSK-3, and closely mimicked Wnt signaling in Xenopus embryos.
TL;DR: Results indicate that abnormal information storage in corticostriatal synapses is linked with the development of L-DOPA–induced dyskinesia.
Abstract: Long-term treatment with the dopamine precursor levodopa (L-DOPA) induces dyskinesia in Parkinson's disease (PD) patients We divided hemiparkinsonian rats treated chronically with L-DOPA into two groups: one showed motor improvement without dyskinesia, and the other developed debilitating dyskinesias in response to the treatment We then compared the plasticity of corticostriatal synapses between the two groups High-frequency stimulation of cortical afferents induced long-term potentiation (LTP) of corticostriatal synapses in both groups of animals Control and non-dyskinetic rats showed synaptic depotentiation in response to subsequent low-frequency synaptic stimulation, but dyskinetic rats did not The depotentiation seen in both L-DOPA-treated non-dyskinetic rats and intact controls was prevented by activation of the D1 subclass of dopamine receptors or inhibition of protein phosphatases The striata of dyskinetic rats contained abnormally high levels of phospho[Thr34]-DARPP-32, an inhibitor of protein phosphatase 1 These results indicate that abnormal information storage in corticostriatal synapses is linked with the development of L-DOPA-induced dyskinesia
TL;DR: The development of an ERβ-selective antibody that cross-reacts with mouse, rat, and human ERβ protein and its use to determine the distribution of ERβ in the murine brain is reported.
Abstract: Estrogen receptor α (ERα) and ERβ are members of the steroid nuclear receptor family that modulate gene transcription in an estrogen-dependent manner. ER mRNA and protein have been detected both peripherally and in the central nervous system, with most data having come from the rat. Here we report the development of an ERβ-selective antibody that cross-reacts with mouse, rat, and human ERβ protein and its use to determine the distribution of ERβ in the murine brain. Further, a previously characterized polyclonal antibody to ERα was used to compare the distribution of the two receptors in the first comprehensive description of ER distribution specifically in the mouse brain. ERβ immunoreactivity (ir) was primarily localized to cell nuclei within select regions of the brain, including the olfactory bulb, cerebral cortex, septum, preoptic area, bed nucleus of the stria terminalis, amygdala, paraventricular hypothalamic nucleus, thalamus, ventral tegmental area, substantia nigra, dorsal raphe, locus coeruleus...
TL;DR: The theoretical shift for adsorption of a single protein is found to be extremely sensitive to the target region, with adsor adaptation in the most sensitive region varying as 1/R(5/2).
Abstract: Biosensors based on the shift of whispering-gallery modes in microspheres accompanying protein adsorption are described by use of a perturbation theory. For random spatial adsorption, theory predicts that the shift should be inversely proportional to micorsphere radius R and proportional to protein surface density and excess polarizability. Measurements are found to be consistent with the theory, and the correspondence enables the average surface area occupied by a single protein to be estimated. These results are consistent with crystallographic data for bovine serum albumin. The theoretical shift for adsorption of a single protein is found to be extremely sensitive to the target region, with adsorption in the most sensitive region varying as 1R 52 . Specific parameters for single protein or virus particle detection are predicted. © 2003 Optical
TL;DR: An overview of the molecular biology of the flaviviruses is presented, which are enveloped positive-strand RNA viruses capable of causing a number of important human diseases.
Abstract: Publisher Summary This chapter summarizes the current understanding of the molecular biology of flaviviruses and points out promising avenues for future work. The molecular biology of flaviviruses is best understood in the context of the viral life cycle, which provides a framework for the organization of this chapter. Flavivirus particles bind to cells via interactions between the viral surface glycoprotein and cellular receptors. Several cell surface proteins have been described as putative receptors. In addition, opsonization with immunoglobulins enhances virus particle binding and infection of cells expressing immunoglobulin Fc receptors. Virions are internalized into clathrin-coated pits via receptor-mediated endocytosis. It is thought that virions are brought into a prelysosomal endocytic compartment where low pH induces fusion among the viruses and host cell membranes to release the virus nucleocapsid. The viral genome is released into the host cytoplasm by the process of nucleocapsid uncoating, which is not yet fully understood. Translation of the viral genome produces proteins that lead to replication of the viral genome and assembly of new virus particles. Flavivirus infection induces rearrangement of cytoplasmic membranes in the perinuclear region. Virus particles are thought to assemble by budding into the endoplasmic reticulum. A few studies have shown evidence for budding at the plasma membrane. Based on trans-complementation studies, it appears that genome packaging is coupled to RNA replication. Nascent virus particles pass through the host secretory pathway, where virion maturation occurs, and are released.
TL;DR: A single dose of αGalCer rapidly stimulates the full maturation of dendritic cells in situ, and this accounts for the induction of combined Th1 CD4+ and CD8+ T cell immunity to a coadministered protein.
Abstract: The maturation of dendritic cells (DCs) allows these antigen-presenting cells to initiate immunity. We pursued this concept in situ by studying the adjuvant action of α-galactosylceramide (αGalCer) in mice. A single i.v. injection of glycolipid induced the full maturation of splenic DCs, beginning within 4 h. Maturation was manifest by marked increases in costimulator and major histocompatibility complex class II expression, interferon (IFN)-γ production, and stimulation of the mixed leukocyte reaction. These changes were not induced directly by αGalCer but required natural killer T (NKT) cells acting independently of the MyD88 adaptor protein. To establish that DC maturation was responsible for the adjuvant role of αGalCer, mice were given αGalCer together with soluble or cell-associated ovalbumin antigen. Th1 type CD4+ and CD8+ T cell responses developed, and the mice became resistant to challenge with ovalbumin-expressing tumor. DCs from mice given ovalbumin plus adjuvant, but not the non-DCs, stimulated ovalbumin-specific proliferative responses and importantly, induced antigen-specific, IFN-γ producing, CD4+ and CD8+ T cells upon transfer into naive animals. In the latter instance, immune priming did not require further exposure to ovalbumin, αGalCer, NKT, or NK cells. Therefore a single dose of αGalCer i.v. rapidly stimulates the full maturation of DCs in situ, and this accounts for the induction of combined Th1 CD4+ and CD8+ T cell immunity to a coadministered protein.
TL;DR: This work shows that the TRF1 complex interacts with a single-stranded telomeric DNA-binding protein—protection of telomeres 1 (POT1)—and that human POT1 controls telomerase-mediated telomere elongation.
Abstract: Human telomere maintenance is essential for the protection of chromosome ends, and changes in telomere length have been implicated in ageing and cancer. Human telomere length is regulated by the TTAGGG-repeat-binding protein TRF1 and its interacting partners tankyrase 1, TIN2 and PINX1 (refs 5-9). As the TRF1 complex binds to the duplex DNA of the telomere, it is unclear how it can affect telomerase, which acts on the single-stranded 3' telomeric overhang. Here we show that the TRF1 complex interacts with a single-stranded telomeric DNA-binding protein--protection of telomeres 1 (POT1)--and that human POT1 controls telomerase-mediated telomere elongation. The presence of POT1 on telomeres was diminished when the amount of single-stranded DNA was reduced. Furthermore, POT1 binding was regulated by the TRF1 complex in response to telomere length. A mutant form of POT1 lacking the DNA-binding domain abrogated TRF1-mediated control of telomere length, and induced rapid and extensive telomere elongation. We propose that the interaction between the TRF1 complex and POT1 affects the loading of POT1 on the single-stranded telomeric DNA, thus transmitting information about telomere length to the telomere terminus, where telomerase is regulated.
TL;DR: The results of this study of patients registered in the IFAR over a 20-year period provide information that will enable better prediction of outcome and aid clinicians with decisions regarding major therapeutic modalities.
TL;DR: In this review, the development of this technology is discussed, its broad application to biological systems, and its possible role in the area of proteomics are discussed.
Abstract: ▪ Abstract Expressed protein ligation (EPL) is a protein engineering approach that allows recombinant and synthetic polypeptides to be chemoselectively and regioselectively joined together. The approach makes the primary structure of most proteins accessible to the tools of synthetic organic chemistry, enabling the covalent structure of proteins to be modified in an unprecedented fashion. The ability to incorporate noncoded amino acids, biophysical probes, and stable isotopes into specific locations within proteins provides research tools to peer into the inner workings of these molecules. In this review I discuss the development of this technology, its broad application to biological systems, and its possible role in the area of proteomics.