HSCB, a co-chaperone in mitochondrial iron-sulfur cluster biogenesis, is a novel candidate gene for congenital sideroblastic anemia

Congenital sideroblastic anemias (CSA) are inherited diseases resulting from defects in heme biosynthesis, mitochondrial iron-sulfur cluster (ISC) assembly, or mitochondrial translation. CSAs are characterized by pathological iron deposits in the mitochondria of bone marrow erythroblasts. Recently the Fleming Lab at Boston Children’s Hospital has reported mutations in HSPA9, a chaperone involved in ISC assembly, as a cause of nonsyndromic CSA. Here we identified a CSA patient harboring two variants in HSCB, encoding a binding partner of HSPA9: a paternally inherited promoter variant (c-134C>A) and a maternally inherited frameshift variant (T87fs) predicted to result in a truncated protein. To better understand the pathophysiology of these variants, we investigated HSCB protein expression and function in patient-derived skin fibroblasts. Patient fibroblasts show evidence of decreased HSCB protein levels. shRNA targeting HSCB was employed to specifically suppress HSCB expression in the K562 erythroid-like cell line model. shRNA-infected K562 cells presented with perturbed iron homeostasis, a shift to glycolytic energy production, and diminished hemoglobinization. Targeted deletion of murine Hscb is embryonic lethal prior day E7.0. Tissue-specific lox-Cre transgenic lines, including Vav-, EpoR- and Mx-Cre demonstrate that Hscb is essential for hematopoiesis and erythropoiesis. Mutant mice present with hematopoietic defects similar to the index patient. Vav-Cre animals die prior to post-natal day 9 with decreased red cell counts, white cell counts, and decreased hemoglobin compared to wild-type animals. Floxed-null EpoR-Cre animals die before embryonic day 13. To excise Hscb specifically in the hematopoietic compartment of adult animals, conditional Mx-Cre animals were generated through bone marrow transplantation and temporally induced with polyinosinic-polycytidylic acid treatment. The animals died 22 days post-injection with decreased red blood cells, white blood cells, hemoglobin, and an overall decline in hematopoiesis of the bone marrow. These data demonstrate that HSCB is required for erythropoiesis and hematopoiesis and that the patient mutations are a pathogenic cause of CSA

Solution Structure of the Iron−Sulfur Cluster Cochaperone HscB and Its Binding Surface for the Iron−Sulfur Assembly Scaffold Protein IscU†‡

ABSTRACT: The interaction between IscU and HscB is critical for successful assembly of iron-sulfur clusters. NMR experiments were performed on HscB to investigate which of its residues might be part of the IscU binding surface. Residual dipolar couplings ( 1 DHN and 1 DCRHR) indicated that the crystal structure of HscB [Cupp-Vickery, J. R., and Vickery, L. E. (2000) Crystal structure of Hsc20, a J-type cochaperone from Escherichia coli, J. Mol. Biol. 304, 835-845] faithfully represents its solution state. NMR relaxation rates ( 15 N R1, R2) and 1 H- 15 N heteronuclear NOE values indicated that HscB is rigid along its entire backbone except for three short regions which exhibit flexibility on a fast time scale. Changes in the NMR spectrum of HscB upon addition of IscU mapped to the J-domain/C-domain interface, the interdomain linker, and the C-domain. Sequence conservation is low in the interface and in the linker, and NMR changes observed for these residues likely result from indirect effects of IscU binding. NMR changes observed in the conserved patch of residues in the C-domain (L92, M93, L96, E97, E100, E104, and F153) were suggestive of a direct interaction with IscU. To test this, we replaced several of these residues with alanine and assayed for the ability of HscB to interact with IscU and to stimulate HscA ATPase activity. HscB(L92A,M93A,F153A) and HscB(E97A,E100A,E104A) both showed decreased binding affinity for IscU; the (L92A,M93A,F153A) substitution also strongly perturbed the allosteric interactio

Non-Abelian Quantum Hall States and their Quasiparticles: from the Pattern of Zeros to Vertex Algebra

In the pattern-of-zeros approach to quantum Hall states, a set of data {n;m;S_a|a=1,...,n; n,m,S_a in N} (called the pattern of zeros) is introduced to characterize a quantum Hall wave function. In this paper we find sufficient conditions on the pattern of zeros so that the data correspond to a valid wave function. Some times, a set of data {n;m;S_a} corresponds to a unique quantum Hall state, while other times, a set of data corresponds to several different quantum Hall states. So in the latter cases, the patterns of zeros alone does not completely characterize the quantum Hall states. In this paper, We find that the following expanded set of data {n;m;S_a;c|a=1,...,n; n,m,S_a in N; c in R} provides a more complete characterization of quantum Hall states. Each expanded set of data completely characterize a unique quantum Hall state, at least for the examples discussed in this paper. The result is obtained by combining the pattern of zeros and Z_n simple-current vertex algebra which describes a large class of Abelian and non-Abelian quantum Hall states \Phi_{Z_n}^sc. The more complete characterization in terms of {n;m;S_a;c} allows us to obtain more topological properties of those states, which include the central charge c of edge states, the scaling dimensions and the statistics of quasiparticle excitations.Comment: 42 pages. RevTeX

Three hydrophobic amino acids in Escherichia coli HscB make the greatest contribution to the stability of the HscB-IscU complex

<p>Abstract</p> <p>Background</p> <p>General iron-sulfur cluster biosynthesis proceeds through assembly of a transient cluster on IscU followed by its transfer to a recipient apo-protein. The efficiency of the second step is increased by the presence of HscA and HscB, but the reason behind this is poorly understood. To shed light on the function of HscB, we began a study on the nature of its interaction with IscU. Our work suggested that the binding site of IscU is in the C-terminal domain of HscB, and two different triple alanine substitutions ([L92A, M93A, F153A] and [E97A, E100A, E104A]) involving predicted binding site residues had detrimental effects on this interaction. However, the individual contribution of each substitution to the observed effect remains to be determined as well as the possible involvement of other residues in the proposed binding site.</p> <p>Results</p> <p>In the work reported here, we used isothermal titration calorimetry to characterize the affinity of single alanine HscB mutants for IscU, and subsequently confirmed our results with nuclear magnetic resonance spectroscopy. Alanine substitutions of L92, L96, and F153 severely impaired the ability of HscB to form a complex with IscU; substitutions of R87, R99, and E100 had more modest effects; and substitutions of T89, M93, E97, D103, E104, R152, K156, and S160 had only minor or no detectable effects.</p> <p>Conclusions</p> <p>Our results show that the residues of HscB most important for strong interaction with IscU include three hydrophobic residues (L92, L96, and F153); in addition, we identified a number of other residues whose side chains contribute to a lesser extent to the interaction. Our results suggest that the triple alanine substitution at HscB positions 92, 96, and 153 will destabilize the HscB-IscU complex by ΔΔ<it>G</it>_b≅ 5.7 kcal/mol, equivalent to a ≅ 15000-fold reduction in the affinity of HscB for IscU. We propose that this triple mutant could provide a more definitive test of the functional importance of the HscB-IscU interaction in vivo than those used previously that yielded inconclusive results.</p

Thoughts about a General Theory of Influence in a DIME/PMESII/ASCOP/IRC2 Model

The leading question of this paper is: “How would influence warfare (“iWar”) work and how can we simulate it?” The paper discusses foundational aspects of a theory and model of influence warfare by discussing a framework built along the DIME/PMESII/ASCOP dimension forming a prism with three axes. The DIME concept groups the many instruments of power a nation state can muster into four categories: Diplomacy, Information, Military and Economy. PMESII describes the operational environment in six domains: Political, Military, Economic, Social, Information and Infrastructure. ASCOPE is used in counter insurgency (COIN) environments to analyze the cultural and human environment (aka the “human terrain”) and encompasses Areas, Structures, Capabilities, Organization, People and Events. In addition, the model reflects about aspects of information collection requirements (ICR) and information capabilities requirements (ICR) - hence DIME/PMESII/ASCOP/ICR2. This model was developed from an influence wargame that was conducted in October 2018. This paper introduces basic methodical questions around model building in general and puts a special focus on building a framework for the problem space of influence/information/hybrid warfare takes its shape in. The article tries to describe mechanisms and principles in the information/influence space using cross discipline terminology (e.g. physics, chemistry and literature). On a more advanced level this article contributes to the Human, Social, Culture, Behavior (HSCB) models and community. One goal is to establish an academic, multinational and whole of government influence wargamer community. This paper introduces the idea of the perception field understood as a molecule of a story or narrative that influences an observer. This molecule can be drawn as a selection of vectors that can be built inside the DIME/PMESII/ASCOP prism. Each vector can be influenced by a shielding or shaping action. These ideas were explored in this influence wargame

Studies on the mechanism of catalysis of iron-sulfur cluster transfer from IscU[2Fe2S] by HscA/HscB chaperones

The HscA/HscB chaperone/cochaperone system accelerates transfer of iron-sulfur clusters from the FeS-scaffold protein IscU (IscU(2)[2Fe2S], holo-IscU) to acceptor proteins in an ATP-dependent manner. We have employed visible region circular dichroism (CD) measurements to monitor chaperone-catalyzed cluster transfer from holo-IscU to apoferredoxin and to investigate chaperone-induced changes in properties of the IscU(2)[2Fe2S] cluster. HscA-mediated acceleration of [2Fe2S] cluster transfer exhibited an absolute requirement for both HscB and ATP. A mutant form of HscA lacking ATPase activity, HscA(T212V), was unable to accelerate cluster transfer, suggesting that ATP hydrolysis and conformational changes accompanying the ATP (T-state) to ADP (R-state) transition in the HscA chaperone are required for catalysis. Addition of HscA and HscB to IscU(2)[2Fe2S] did not affect the properties of the [2Fe2S] cluster, but subsequent addition of ATP was found to cause a transient change of the visible region CD spectrum, indicating distortion of the IscU-bound cluster. The dependence of the rate of decay of the observed CD change on ATP concentration and the lack of an effect of the HscA(T212V) mutant were consistent with conformational changes in the cluster coupled to ATP hydrolysis by HscA. Experiments carried out under conditions with limiting concentrations of HscA, HscB, and ATP further showed that formation of a 1:1:1 HscA-HscB-IscU(2)[2Fe2S] complex and a single ATP hydrolysis step are sufficient to elicit the full effect of the chaperones on the [2Fe2S] cluster. These results suggest that acceleration of iron-sulfur cluster transfer involves a structural change in the IscU(2)[2Fe2S] complex during the T --> R transition of HscA accompanying ATP hydrolysis

Melt blending and characterization of carbon nanoparticles-filled thermoplastic polyurethane elastomers

In this work, thermoplastic polyurethane (TPU) elastomers reinforced with carbon nanosized particles were produced by a special melt blending technique. A TPU was melt blended with high-structured carbon black and carbon nanofibres (1 wt%). A miniature asymmetric batch mixer, which applies high shear levels to the melt, ensured good particles dispersion. The TPU material systems were then thoroughly characterized using thermogravimetric analysis, differential scanning calorimetry, tensile mechanical testing, electrical resistance measurements and flammability tests. The different nanofillers exhibited different influences on the TPU properties, these materials featuring interesting and improved multifunctional behaviours, with high propensity for large deformation sensors applications.This work was supported by FCT – Portuguese Foundation for Science and Technology through projects NANOSens – PTDC/CTM/73465/2006

Setting a Course for the Human Social Culture Behavior (HSCB) Modeling Standards Study Group

Documents include Paper and Presentation.Simulation Interoperability Standards Organization (SISO) SIW Conference PaperResearch and development efforts in modeling and simulation are increasingly focusing on the modeling of human societies, culture, and behavior for improved gaming experiences, analytical and decision tools, training, experimentation, and many other purposes. The rapid growth in this area is creating increasing demands for data, models, tools, and techniques. To investigate the potential need for standards or best practices in this area, the Simulation Interoperability Standards Organization (SISO) has initiated the Human Social Culture Behavior (HSCB) Modeling Standards Study Group. This paper examines the opportunity for development of standards and best practices in this domain and lays out a possible plan of action for the new Study Group.Naval Postgraduate School, Monterey, CA.Approved for public release; distribution is unlimited