Analysis of reaction schemes using maximum rates of constituent steps
Ali Hussain Motagamwala and James A. Dumesic
The design of active and selective catalysts is essential for a wide range of industrial applications and societal issues. A fundamental approach is to identify the key elementary steps and to elucidate the predicted reaction kinetics for potential reaction mechanisms. We present a methodology to analyze analytically the performance of catalytic reaction schemes by calculation of the maximum rates of the constituent steps. This proposed methodology can be used to identify the important transition states and adsorbed species, such that more detailed calculations can be carried out for these species, whereas more approximate methods can be used for the remaining species, thereby substantially reducing the computational time required to elucidate how catalyst performance is controlled by the fundamental surface chemistry. (See pp. E2879–E2888.)
Achieving diverse and monoallelic olfactory receptor selection through dual-objective optimization design
Xiao-Jun Tian, Hang Zhang, Jens Sannerud, and Jianhua Xing
For sensitive smell detection, each mammalian olfactory sensory neurons need to express stochastically only one allele of 1 out of possibly more than 1,000 types of olfactory receptors. The mechanism for this monoallelic expression remains as one of the biggest unresolved questions for decades. Using mathematical modeling and computer simulations, we identified a three-layer regulation mechanism the olfactory system adopts to achieve single allelic expression and several other biological requirements such as maximizing the overall diversity of expressed olfactory receptors. The revealed mechanism provides insight for formulating biological processes as multiple-objective optimization problems. (See pp. E2889–E2898.)
Interactions between RNA polymerase and the core recognition element are a determinant of transcription start site selection
Irina O. Vvedenskaya, Hanif Vahedian-Movahed, Yuanchao Zhang, Deanne M. Taylor, Richard H. Ebright, and Bryce E. Nickels
For all cellular RNA polymerases, the position of the transcription start site (TSS) relative to core promoter elements is variable. Furthermore, environmental conditions and regulatory factors that affect TSS selection have profound effects on levels of gene expression. Thus, identifying determinants of TSS selection is important for understanding gene expression control. Here we identify a previously undocumented determinant for TSS selection by Escherichia coli RNA polymerase. We show that sequence-specific protein–DNA interactions between RNA polymerase core enzyme and a sequence element in unwound promoter DNA, the core recognition element, modulate TSS selection. (See pp. E2899–E2905.)
Myosin MyTH4-FERM structures highlight important principles of convergent evolution
Vicente José Planelles-Herrero, Florian Blanc, Serena Sirigu, Helena Sirkia, Jeffrey Clause, Yannick Sourigues, Daniel O. Johnsrud, Beatrice Amigues, Marco Cecchini, Susan P. Gilbert, Anne Houdusse, and Margaret A. Titus
Myosins containing MyTH4-FERM (myosin tail homology 4-band 4.1, ezrin, radixin, moesin, or MF) domains in their tails are found in wide range of phylogenetically divergent organisms. Interestingly, evolutionarily distant MF myosins have similar roles in the extension of actin-filled membrane protrusions, such as filopodia, and microtubule binding, suggesting that their core functions have been highly conserved over evolution. A structural analysis of mammalian and Dd myosin MF domains in combination with comparison of diverse MF myosin sequences illustrate how tuning of existing features can give rise to new structures while preserving the general properties of myosin tails. Thus, tinkering with the MF domain enables it to serve as a multifunctional platform for cooperative recruitment of various partners, allowing common properties to arise through convergent evolution. (See pp. E2906–E2915.)
F1-ATPase conformational cycle from simultaneous single-molecule FRET and rotation measurements
Mitsuhiro Sugawa, Kei-ichi Okazaki, Masaru Kobayashi, Takashi Matsui, Gerhard Hummer, Tomoko Masaike, and Takayuki Nishizaka
The major source of ATP in life is ATP synthase, and its catalytic part is known to be the F1 rotary motor. F1’s structure and function have been characterized in spectacular detail by crystallography and single-molecule techniques, respectively. However, despite more than two decades of intense research, the correspondence of the observed functional states and crystal structures is uncertain. To match structures and states, we perform single-molecule fluorescence-based distance measurements and simultaneous rotary angle measurements on F1-ATPase, and then exploit the wealth of structural data in their analysis. The resulting comprehensive view of the F1’s ATPase cycle reveals the functional principles in the coupling of chemical reactions, stator conformations, and rotary angles for efficient ATP synthesis. (See pp. E2916–E2924.)
Loss of RPGR glutamylation underlies the pathogenic mechanism of retinal dystrophy caused by TTLL5 mutations
Xun Sun, James H. Park, Jessica Gumerson, Zhijian Wu, Anand Swaroop, Haohua Qian, Antonina Roll-Mecak, and Tiansen Li
Mutations affecting two unrelated genes, retinitis pigmentosa GTPase regulator (RPGR) and tubulin tyrosine ligase like 5 (TTLL5), lead to photoreceptor degeneration and blindness in humans. We find that RPGR function in photoreceptor cilia requires glutamylation by TTLL5. Glutamylation is a poorly understood posttranslational modification that consists of the addition of glutamates to target proteins. Moreover, we find that mice lacking RPGR or TTLL5 exhibit similar phenotypes characterized by photoreceptor degeneration and opsin mislocalization. Our work identifies a novel essential regulator of RPGR and demonstrates that disease-causing mutations in these two genes share a common pathogenic pathway in humans. (See pp. E2925–E2934.)
Blockage of neddylation modification stimulates tumor sphere formation in vitro and stem cell differentiation and wound healing in vivo
Xiaochen Zhou, Mingjia Tan, Mukesh K. Nyati, Yongchao Zhao, Gongxian Wang, and Yi Sun
MLN4924, a potent small-molecule inhibitor of NEDD8-activating enzyme, blocks cullin-RING ligase activity through inhibiting cullin neddylation. MLN4924 is widely used in both preclinical and clinical settings for an anticancer application. We report here an unexpected finding: MLN4924 at nanomolar concentration stimulates stem cell proliferation, self-renewal, and differentiation in both tumor and normal stem cell models and promotes skin wound healing in a mouse model and cell migration in vitro. Mechanistic studies revealed that MLN4924 causes c-MYC accumulation and promotes EGFR (epidermal growth factor receptor) dimerization to activate the EGFR signaling pathway. Our study raises a concern in anticancer application of MLN4924, but at the same time provides an opportunity for future development of MLN4924 as an agent for stem cell therapy and tissue regeneration. (See pp. E2935–E2944.)
USP6 oncogene promotes Wnt signaling by deubiquitylating Frizzleds
Babita Madan, Matthew P. Walker, Robert Young, Laura Quick, Kelly A. Orgel, Meagan Ryan, Priti Gupta, Ian C. Henrich, Marc Ferrer, Shane Marine, Brian S. Roberts, William T. Arthur, Jason D. Berndt, Andre M. Oliveira, Randall T. Moon, David M. Virshup, Margaret M. Chou, and Michael B. Major
Ubiquitin-specific protease 6 (USP6) is a deubiquitylase that is overexpressed by chromosome translocation in two human neoplasms, aneurysmal bone cyst and nodular fasciitis. The relevant substrates of this ubiquitin-specific protease are not clear. Here, we identify the Wnt receptor Frizzled (Fzd) as a key target of the USP6 oncogene. Increased expression of USP6 increases the membrane abundance of Fzd, and hence increases cellular sensitivity to Wnts. USP6 opposes the activity of the ubiquitin ligase and tumor suppressor ring finger protein 43 (RNF43). This study identifies a new mechanism for pathological Wnt pathway activation in human disease and suggests a new approach to regulate Wnt activity therapeutically. (See pp. E2945–E2954.)
PDK1–Akt pathway regulates radial neuronal migration and microtubules in the developing mouse neocortex
Yasuhiro Itoh, Maiko Higuchi, Koji Oishi, Yusuke Kishi, Tomohiko Okazaki, Hiroshi Sakai, Takaki Miyata, Kazunori Nakajima, and Yukiko Gotoh
In the developing mammalian neocortex, neurons migrate a long distance from their birthplace to the positions where they form appropriate layers and networks, and dysregulation of this process has been implicated in brain malformation and neurological diseases. Given the fine correlation between temporal order of various sequentially generated neuronal cell types and their spatial distribution, migration speed needs to be tightly controlled to achieve correct neocortical layering, although the underlying mechanisms remain unclear. Here we show that the serine/threonine kinase Akt and its activator phosphoinositide-dependent protein kinase 1 (PDK1) regulate the speed of locomotion of mouse neocortical neurons through the cortical plate. Our data suggest that the PDK1–Akt axis regulates microtubule organization, in part by regulating the cytoplasmic dynein/dynactin complex, in migrating neurons. (See pp. E2955–E2964.)
mir-276a strengthens Drosophila circadian rhythms by regulating timeless expression
Xiao Chen (陈霄) and Michael Rosbash
The circadian clock regulates biochemical, physiological, endocrine, and behavioral features of higher organisms. Although the timekeeping mechanism and many of the functions it governs rely on transcriptional regulation, posttranscriptional regulation also plays an important role. We show here that levels of the abundant Drosophila microRNA mir-276a oscillate under 24-h light–dark cycles and are regulated by the transcription factor Chorion factor 2. mir-276a is important for robust rhythmicity and binds to a single site in the 3' UTR of the important clock gene timeless (tim). We used clustered, regularly interspaced, short palindromic repeats to generate fly strains missing this mir-276a–binding site. Because these strains have elevated TIM levels and enhanced arrhythmicity, the data indicate that mir-276a helps maintain proper TIM levels, which are important for robust rhythmicity. (See pp. E2965–E2972.)
Impaired NK-mediated regulation of T-cell activity in multiple sclerosis is reconstituted by IL-2 receptor modulation
Catharina C. Gross, Andreas Schulte-Mecklenbeck, Anna Rünzi, Tanja Kuhlmann, Anita Posevitz-Fejfár, Nicholas Schwab, Tilman Schneider-Hohendorf, Sebastian Herich, Kathrin Held, Matea Konjević, Marvin Hartwig, Klaus Dornmair, Reinhard Hohlfeld, Tjalf Ziemssen, Luisa Klotz, Sven G. Meuth, and Heinz Wiendl
The importance of natural killer (NK) cells in the control of autoimmunity has recently attracted considerable attention. The current study revealed NK cells as additional players in controlling T-cell activity in CNS autoimmunity. NK-mediated control of T-cell activity in multiple sclerosis (MS) is dysregulated and caused by impaired DNAX accessory molecule-1/CD155 interaction between NK cells and CD4+ T cells. Therapeutic immune modulation of the IL-2 receptor with daclizumab, which has just successfully passed a phase III study in relapsing-remitting MS, not only enhances the cytolytic activity of NK cells but also restores defective NK-mediated immune regulation by increasing the proportion of CD155-expressing CD4+ T cells, thus rendering CD4+ T cells most likely more sensitive to NK-mediated lysis. (See pp. E2973–E2982.)
Restricting nonclassical MHC genes coevolve with TRAV genes used by innate-like T cells in mammals
Pierre Boudinot, Stanislas Mondot, Luc Jouneau, Luc Teyton, Marie-Paule Lefranc, and Olivier Lantz
The conservation and cross-reactivity between species of the T-cell receptor (TR)-V regions and restricting major histocompatibility (MH) molecules characterizing innate-like T cells, natural killer T (NKT) and mucosal-associated invariant T (MAIT), indicate important functions for these cells. Yet, we show that the two MAIT-specific genes, TRAV1 and MR1, have been lost at least three times during the evolution of mammals. In the rabbit, which has few NKT cells and no MR1, we found a candidate invariant TR-α (iTRA) chain and another mammalian MH1Like molecule that seem to coevolve in mammals. Thus, at least three iTRA/MH-like systems were selected during mammalian evolution. The new MH1Like molecule may present a distinct set of antigens to a new innate-like T-cell subset. This study emphasizes the coevolution of TR and MH molecules. (See pp. E2983–E2992.)
Histone deacetylase inhibition enhances antimicrobial peptide but not inflammatory cytokine expression upon bacterial challenge
Natalie Fischer, Emmanuel Sechet, Robin Friedman, Aurélien Amiot, Iradj Sobhani, Giulia Nigro, Philippe J. Sansonetti, and Brice Sperandio
Antimicrobial peptides exert antimicrobial, antifungal, antiviral, and antiprotozoan activity. They are expressed at high concentrations at the intestinal mucosal surface, where they play a crucial role in intestinal homeostasis. Therefore, approaches aiming to boost expression of antimicrobial peptides represent a future therapeutic strategy to treat infections and dysbiosis-driven diseases in humans at a time of increasing incidence of antibiotic resistance. (See pp. E2993–E3001.)
Epigenomic profiling reveals an association between persistence of DNA methylation and metabolic memory in the DCCT/EDIC type 1 diabetes cohort
Zhuo Chen, Feng Miao, Andrew D. Paterson, John M. Lachin, Lingxiao Zhang, Dustin E. Schones, Xiwei Wu, Jinhui Wang, Joshua D. Tompkins, Saul Genuth, Barbara H. Braffett, Arthur D. Riggs, DCCT/EDIC Research Group, and Rama Natarajan
Vascular complications are the main cause of morbidity and mortality in the diabetic population. Clinical trials of diabetic complications show a persistence of benefit from early application of intensive therapy for glycemic control in diabetic patients, a phenomenon referred to as metabolic memory. The mechanisms underlying metabolic memory are not fully understood. In this study, using two groups of type 1 diabetic patients with and without complications development and two sets of genomic DNAs collected 16–17 y apart from the same patients, we showed a persistency of DNA methylation over time at key genomic loci associated with diabetic complications. These data provide direct evidence of a relationship between epigenetics (DNA methylation variations) and human metabolic memory, supporting an epigenetic mechanism. (See pp. E3002–E3011.)
Cos-Seq for high-throughput identification of drug target and resistance mechanisms in the protozoan parasite Leishmania
Élodie Gazanion, Christopher Fernández-Prada, Barbara Papadopoulou, Philippe Leprohon, and Marc Ouellette
Gain-of-function screens using overexpression genomic libraries are powerful tools for discovering drug target/resistance genes, but several limitations make this technique less amenable to high-throughput screening. Using cosmid-based functional screening coupled to next-generation sequencing, an approach that we term Cosmid Sequencing (or “Cos-Seq”), we followed the dynamics of cosmid enrichment during drug pressure in Leishmania, the parasite responsible for leishmaniasis, a neglected tropical disease. This improved and sensitive method has led to the identification and functional characterization of an unprecedented number of drug target/resistance genes against all drugs currently used to treat leishmaniasis. (See pp. E3012–E3021.)
PML plays both inimical and beneficial roles in HSV-1 replication
Pei Xu, Stephen Mallon, and Bernard Roizman
Promyelocytic leukemia protein (PML) is a component of nuclear domain 10 (ND10) bodies and an antiviral effector of IFN-β. A herpes simplex virus (HSV) protein, ICP0, interacts with PML, merges with ND10 bodies, degrades PML, and ultimately takes over the domain of ND10 bodies. Here we show that viral gene expression and growth are reduced in PML−/− cells infected at low ratios of virus to cells. In essence, the results indicate that HSV-1 evolved the means to take advantage of an inimical cellular defense mechanism to degrade it and at the same time use it to gain access to a nuclear domain essential for efficient replication. (See pp. E3022–E3028.)
Fasting induces a form of autonomic synaptic plasticity that prevents hypoglycemia
Manqi Wang, Qian Wang, and Matthew D. Whim
To prevent a fall in blood glucose during fasting, the counter-regulatory response is activated. An important component of this pathway involves the autonomic nervous system and release of epinephrine from the adrenal gland. This autonomic response is often referred to as a reflex, implying the output is hardwired and inflexible. Here we show the strength of the terminal synapse that controls epinephrine release is actually highly plastic. Fasting leads to a long-lasting increase in synaptic strength by a process that requires neuropeptide Y and Y5 receptors. In the absence of neuropeptide Y, synaptic strengthening is absent, epinephrine release is reduced, and the mice become hypoglycemic. These findings indicate that the response to fasting involves significant autonomic synaptic plasticity. (See pp. E3029–E3038.)
Myosin light chain phosphorylation enhances contraction of heart muscle via structural changes in both thick and thin filaments
Thomas Kampourakis, Yin-Biao Sun, and Malcolm Irving
Contraction of heart muscle is triggered by calcium binding to the actin-containing thin filaments but modulated by structural changes in the myosin-containing thick filaments. We showed that phosphorylation of the myosin regulatory light chain generates a structural signal that is transmitted between myosin molecules in the thick filament and from the thick to the thin filaments, altering their calcium sensitivity. A closely related dual-filament signaling pathway underlies the enhanced contractility of heart muscle when it is stretched. These coordinated and cooperative changes in thick and thin filament structure are an essential component of contractile regulation in the healthy heart, and their impairment is likely to underlie the functional effects of mutations in thick filament proteins in heart disease. (See pp. E3039–E3047.)