Researchers develop universally accessible application that can simulate complex molecular interactions

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Completed Collaboration & Service Projects

one) Relationship between structural Dynamics and Sequence Evolution; Application to molecular machined such every bit bacterial chaperonons and HSP70 chaperones

PI: I. Bahar,University of Pittsburgh
Collaborators: Amnon Horovitz,Weizmann Institute; R. Altman,Stanford University; Fifty. Gierasch,University of Massachusetts.

We have expanded the telescopic of this project to include Dr. Amnon Horovitz from Weizmann Institute, Israel, an expert in the allostery of chaperonins every bit well as sequence co-evolution. We are exploring the sequence->structure->dynamics->function mapping of allosteric proteins. Our overarching goal is to establish the computational methodology for simulating the mechanism of biomolecular systems on the order of Megadaltons. The molecular machines that nosotros selected for developing and implementing the methodology are bacterial chaperonins (GroEL), molecular chaperones (East. coli Hsp70 DnaK) and their co-chaperones. We study the ATP-regulated mechanisms of intersubunit (GroEL) or interdomain (DnaK) communication, also as the role of co-chaperonins or co-chaperones in modulating the conformational changes occurring along the allosteric wheel of these machines.

In Year 3 we fabricated progress in quantitative analysis of the development of proteins, and in item in assessing the relationship, if whatsoever, between their structural dynamics and sequence evolution. In a recent study performed toward this goal (Mao et al., 2015), we carried out a systematic comparative analysis of current methods for analyzing sequence co-evolution and nosotros proposed a hybrid model that yields high performance. The ii metrics for performance were the elimination of fake positives betwixt non-interacting proteins, and the verification of tertiary contacts between co-evolving pairs. The methods used in this newly published (that acknowledged MMBioS support) are currently being implemented into our ProDy API.

Publications Resulting from This Piece of work

    • General IJ, Liu Y, Blackburn Grand, Mao W, Gierasch LM, Bahar I (2014) ATPase subdomain IA is a mediator of interdomain allostery in Hsp70 molecular chaperonesPLoS Comp Bio
      10: e1003624 PMID: 24831085, PMC4022485
    • Mao W, Kaya C, Dutta A, Horovitz A, Bahar I (2015) Comparative Study of the Effectiveness and Limitations of Electric current Methods for Detecting Sequence CoevolutionBioinformatics
      31: 1929-37 PMID: 25697822; PMC4481699


ii) Membrane Proteins Structure and Dynamics Consortium (MPSDC) Computational Cadre, and AMPA receptors structural dynamics

PI: I. Bahar,University of PittsburghCollaborators: B. Roux and E. Perozo,University of Chicago; Grand. Schulten and E. Tajkhorshid,University of Illinois at Urbana-Champaign; I. Greger,Academy of Cambridge, U.k.; H. Weinstein, Cornell University

To assess whether ANM-predicted AMPAR NTD pivoting motions tin occurin cellulo, the Greger lab mutated K262 to cysteine (K262C), hypothesizing that this mutation would trap the NTD tetramer in a closed conformer via germination of a disulfide bridge. GluA2 wild blazon (WT) and K262C mutant were expressed in HEK293 cells and protein extracts were analyzed by Western absorb. The results showed that our hypothesis was valid, thus lending support to computational predictions. Nosotros further examined the dynamics of AMPAR and NMDAR to observe that the differ along a few slow modes only (Effigy one). Overall, our data provided a commencement glimpse into the dynamic spectrum of AMPAR and NMDARs and delineated conserved mechanisms underlying allosteric advice in iGluRs.

csp2Figure ane. AMPAR to NMDAR conformations differ by rearrangements along a few soft modes.

The dark blue bars show the overlap (correlation cosine) between ANM modes of AMPAR and the structural deviation vector between AMPAR and NMDAR; the red curve displays the cumulative overlap; the dotted green bend shows the expected cumulative overlap if the modes were every bit contributing each. The inset shows the initial structures of AMPAR (left) and NMDAR (right).

Publications Resulting from This Work

    • Krieger J, Bahar I, Greger IH (2015) Structure, Dynamics, and Allosteric Potential of Ionotropic Glutamate Receptor North-Terminal DomainsBiophys J
      109: 1136-48 PMID: 26255587, PMC4576161
    • Dutta A, Krieger J, Lee JY, Garcia-Nafria J, Greger IH, Bahar I (2015) Cooperative Dynamics of Intact AMPA and NMDA Glutamate Receptors: Similarities and Subfamily-Specific DifferencesStructure
      23: 1692-1704 PMID: 26256538, PMC4558295
    • Dutta A, Shrivastava IH, Sukumaran M, Greger IH, Bahar I (2012) Comparative dynamics of NMDA- and AMPA-Glutamate receptor N-concluding domainsStructure
      xx: 1838-49 PMID: 22959625, PMC3496038


3) Integration, prediction, and generation of mixed mode information using graphical models, with applications to protein-poly peptide interactions

PIs: C. J. Langmead,Carnegie Mellon University
Collaborators: C. Bailey-Kellog,Dartmouth University; N. Ramakrishnan,Virginia Tech;  A. Friedman,Purdue Academy

This C&SP was an NSF-funded project titled: “Integration, prediction, and generation of mixed mode information using graphical models, with applications to protein-protein interactions” (NSF IIS-0905193). It was collaboration between Carnegie Mellon Academy, Dartmouth College, Virginia Tech, and Purdue. The projection concluded in August, 2013. The focus of this C&SP was to develop novel probabilistic graphical models for modeling poly peptide-protein interactions. Three broad classes of techniques were adult to integrate attribute-value and relational data, integrate statistical and physical information, and utilize probabilistic models generatively. The research has resulted in new graphical models, new algorithms, and the application of those algorithms to the modeling of poly peptide structures and dynamics, including protein-poly peptide interactions. Specific accomplishments include: the first optimal algorithm for learning regularized undirected graphical models of poly peptide sequences; the first method for performing binding gratuitous energy calculations for protein-protein interactions via graphical models; the first probabilistic graphical models of molecular dynamics; the first algorithm for learning the parameters of molecular mechanics strength fields by minimizing functionals over Boltzmann distributions; the first undirected model of distributions on the hypersphere (for modeling distributions over angles); a game-theoretic method for modeling the emergence of drug resistance-causing mutations in proteins; new algorithms for learning semi- and not-parametric distributions; and a new class of regression models for predicting binding free energies. The project resulted in seventeen publications, three Ph.D. dissertations, and i patent. Primary applications of this research include computer-aided drug design and computer-aided protein engineering. Secondary applications of this work include techniques for identifying functionally important residues mediating binding and allostery.

Publications Resulting from This Work

    • Kamisetty H, Ghosh B, Langmead CJ, Bailey-Kellogg C (2014) Learning Sequence Determinants of Poly peptide: protein Interaction Specificity with Sparse Graphical ModelsRes Comput Mol Biol
      8394:129-143 PMID: 25414914, PMC4235964.
    • Kamisetty H, Ghosh B, Langmead CJ, Bailey-Kellogg C (2015) Learning sequence determinants of protein: poly peptide interaction specificity with sparse graphical modelsJ Comput Biol
      22:474- 86 PMID: 25973864, PMC4449715


5) Modeling immunoreceptor signaling, autophagy, and endocytosis, Hardening Software for Rule-Based Modeling

PI:J. Faeder,University of PittsburghCollaborator:Westward. S. Hlavacek,Los Alamos National Laboratory

The fundamental events in FcεRI signaling initiation were considered during the pioneering work of Goldstein, Faeder, Hlavacek and colleagues to found the utility of rule-based methods for mechanistic modeling of the dynamics of molecular interactions in signaling networks. This led to the creation of BioNetGen, already introduced in the BTRC proposal as a computational framework for modeling biochemical networks. Here, we propose to take advantage of successful initiatives at the BTRC for integration of the BioNetGen rule-based modeling arroyo into the mesh-based architecture that underlies MCell spatial simulations. In particular, we volition focus on the proposed new capabilities for adaptive meshing, which can superimpose well-characterized changes in surface geometry over time during simulations. This unique feature volition let us to develop novel computational models to depict simultaneously dull endocytic vesicle dyanamics and fast signaling dynamics at vesicle surfaces.

Aim 1. To utilize the CellBlender interface to create a framework for prison cell shape changes that occur during mast jail cell signaling, such as microvillar-to-ruffled transformation, advent of signaling patches, and changes associated with enhanced endocytosis.

Aim 2. To evaluate the effects of a irresolute 3D geometry on signaling output, by incorporating existing Rule-Based Models for FcεRI chemical reaction networks into MCell-BioNetGen and refining the model with new parameters for spatial distributions, rate constants and diffusion characteristics collected through funded initiatives.

During the project we adult MCell-R, which at present enables network-gratis simulations to exist performed straight in MCell based on a BioNetGen model specification. We accept validated that the simulator gives correct results for a range of FceRI signaling models and too for the bivalent-ligand bivalent receptor, a simple example of unrestricted assemblage. In principle the simulator can at present be used to simulate spatial signaling dynamics of any model that tin be described in BNGL with any geometry that can be specified in MCell, including a geometry that changes in time. We program to implement several such models in the future.

Publications Resulting from This Piece of work

    • Chylek LA, Harris LA, Tung CS, Faeder JR, Lopez CF, Hlavacek WS (2013) Rule-based modeling: a computational approach for studying biomolecular site dynamics in prison cell signaling systemsWiley Interdiscip Rev Syst Biol Med
      vi: 13-36 PMID: 24123887, PMC3947470
    • Chylek LA, Harris LA, Faeder JR, Hlavacek WS (2015) Modeling for (concrete) biologists: an introduction to the dominion-based arroyoPhys Biol
      12: 045007 PMID: 26178138, PMC4526164
    • Lin YT, Feng S, Hlavacek WS. (2019) Scaling methods for accelerating kinetic Monte Carlo simulations of chemic reaction networks.J Chem Phys. 2019 Jun 28;150(24):244101. doi: 10.1063/1.5 096774. PMID: 31255063


half dozen) Distance-dependent structure and office of neuronal dendrites

PIs: T. Sejnowski and T. Bartol,The Salk Institute
Collaborator: M. Harris,University of Texas at Austin

The purpose of this CS&P is to collaborate with Kristen Harris’ lab to create realistic MCell models of dendritic structure and function.

MCell can simulate the diffusion and interaction of molecules involved in biochemical signaling pathways within the 3D subcellular structure of cells. To practice so, surface meshes used to represent prison cell membranes and subcellular structures must run into very strict geometric standards (e.g. h2o-tight, not-intersecting, manifold). We help create realistic MCell models of CA1 dendritic spines, in particular to learn how more the authentic serial section tomographic reconstructions of core organelles impact simulations of molecular signaling inside dendritic spines.

Recently two papers were published relating to this C&SP. The first paper, published in Frontiers in Synaptic Neuroscience, reported on reconstitution of calcium dynamics in dendritic spines. Nine parameters of the model were optimized within realistic experimental limits by a process that compared results of simulations to published information. Simulations in the optimized model reproduce the timing and amplitude of Ca(2+) transients measured experimentally in intact neurons. Thus, the characteristics of individual isolated proteins determined in vitro could accurately reproduce the dynamics of experimentally measured Ca(2+) transients in spines. The 2nd paper, published in eLife, found that dendritic spines that receive input from the same axon are the same size. This finding allowed estimation of the variability of synaptic plasticity and we found that the amount of information stored at synapses is approximately 4.7 $.25, which is an social club of magnitude larger than previous estimates.

Publications Resulting from This Piece of work

    • Edwards J, Daniel Eastward, Kinney J, Bartol T, Sejnowski T, Johnston D, Harris G, Bajaj C (2014) VolRoverN: enhancing surface and volumetric reconstruction for realistic dynamical simulation of cellular and subcellular roleNeuroinformatics
      12:277-89 PMID: 24100964, PMC4033674
    • Kinney JP, Spacek J, Bartol TM, Bajaj CL, Harris KM, Sejnowski TJ (2013) Extracellular sheets and tunnels modulate glutamate diffusion in hippocampal neuopilJ Comp Neurol
      521: 448-64 PMID: 22740128, PMC3540825
    • Bartol TM, Keller DX, Kinney JP, Bajaj CL, Harris KM, Sejnowski TJ, Kennedy MB (2015) Computational reconstitution of spine calcium transients from individual proteinsFront Synaptic Neurosci
      7: 17 PMID: 26500546, PMC4595661
    • Bartol TM, Bromer C, Kinney J, Chirillo MA, Bourne JN, Harris KM, Sejnowski TJ (2015) Nanoconnectomic upper bound on the variability of synaptic plasticityElife
      4.pii: e10778 PMID: 26618907, PMC4737657


8) Using generative models of prison cell system to investigate tumor cell heterogeneity

PIs:R. Potato and G. Rohde,Carnegie Mellon University
Collaborators: S. Altschuler and Fifty. Wu,Academy of Texas Southwestern Medical Center

Tumor cell populations have long been known to be circuitous mixtures of multiple phenotypes. Given the precision with which one is able to measure properties of cellular construction and role, information technology is often the example that the mensurate of two cells volition differ. Paramount to deriving useful information from a potentially large set of measurements is to exist able to determine which differences are statistically pregnant as well every bit their biological pregnant. The established framework for analyzing the heterogeneity of tumor cell populations involves analyzing these measurements in a high dimensional feature infinite. Features can be simple and easily interpretable (such as cell size), or circuitous and hard to translate (such as texture features). The goal of this subproject is to endeavor to correspond prison cell images using generative models that are more easily interpretable and could potentially offer more than insights into the underlying phenomena that touch the distribution of phenotypes in tumor cells.


9) Large-scale electron microscopy of calcium-imaged neuron populations

PIs: A. Wetzel and G. Hood,Pittsburgh Supercomputing Center
Collaborator: D. Bock,Janelia Farm Research Campus, Howard Hughes Medical Institute

During the concluding year of this project, the biological focus of Bock’south work has shifted from localized mouse brain circuits to the mapping of complete drosophila brains with an emphasis on visual circuits. This alter did not affect the relation of our collaboration which is focused on the technical aspects of maximum scale series-TEM data associates.

Bock’due south grouping has improved the throughput of their automatic 4 camera TEM system. Although the cameras and their raw data rates are unchanged new mechanisms take been tested in the last year for automated sample cassette loading. This has essentially improved the ratio of imaging to sample exchange fourth dimension. The long term goal for this new sample loading method is to enable continuous runs up to two weeks for datasets of xv,000 sections and >50 TBytes. Private sections are imaged in arrays of ~xx,000 five MPixel tiles, typically 112 columns by 196 rows, with a unique stepping pattern to accommodate constraints on the spacing of the optical cameras and lenses. One difficult aspect of these data is the use of the small 5 MPixel tiles which, particularly in the case of hierarchical registration methods, become limited by poor performance of large filesystems when processing small files.

Regular production of datasets at this scale would be difficult to annals using our previous method of raw data transmission to the PSC followed by the return of aligned results to Janelia Farm. Therefore nosotros are preparing for Bock’s team to run our alignment codes on the large 6,144 core Janelia Farm computing cluster. Wetzel and Hood were role of an April 1-11, 2015 Janelia Farm image registration Hackathon organized past Stephan Saafeld’due south grouping. During that trip we were able to examination both the AlignTK and SWiFT codes on the Janelia systems using data from Bock’due south work as well as many others from Janelia and elsewhere (i.due east. Winfried Denk and our C&SP10 collaborator Jeff Lichtman). We were also able to adjust a key component of the SWiFT method for utilise inside i of the Janelia registration codes and found that information technology greatly improved the reliability of registration on difficult regions of Bock’s data.

Publications Resulting from This Work

    • Bock DD, Lee WC, Kerlin AM, Andermann ML, Hood G, Wetzel AW, Yurgenson S, Soucy ER, Kim HS, Reid RC (2011) Network anatomy in vivo physiology of visual cortical neuronsNature
      471: 177-82 PMID: 21390124, PMC3095821


10) Advancing high-throughput thin-section scanning EM to study relationships between neuronal circuit structure and role

PIs: A. Wetzel and 1000. Hood,Pittsburgh Supercomputing Middle
Collaborator: J. Lichtman,Harvard University

Nosotros have continued work on automated alignment of serial SEM paradigm sets produced using tape collecting ultramicrotome sectioning and wafer mounted imaging procedures adult at Harvard. The primary datasets accept been the 16,000 section zebrafish stack, which is the focus of a new CS&P project with the Engert lab and the 114 TB lateral geniculate nucleus (LGN) volume that was previously acquired and is the focus of this C&SP.

The different requirements and resolutions of these datasets have highlighted the demand for a multifariousness of approaches to handle different types of biological content and dissimilar types of prototype characteristics. The large number of tape cracks in the LGN dataset has been particularly difficult since the cracks from whatsoever section to its neighbors interfere with one another during the alignment process. Even though the detailed smashing design is different on each department the fact that most of the cracks have like sizes and orientations ways that they tend to pull the alignment such that we accept seen errors on the social club of 1 per m sections. We are currently working on a strategy to produce a low-resolution map of the cracks, ane/4th calibration, that can then be used to constrain the SWiFT alignment procedure.

We are inbound a new and much higher-throughput stage of EM data capture. Previous datasets, including the zebrafish and LGN, were captured over several months using a Zeiss Merlin microscope and pixel rates up to 20MHz. Lichtman’due south team recently installed the commencement Zeiss 61-beam 1.2 Gpixel/sec scanning microscope. This microscope is now being tested with large single sections in the 400GB/section range, which are captured in a hexagonal mosaic design. These information volition require a substantial number of registration lawmaking adaptations. Nosotros will test speed and accuracy of these modifications once the new equipment is producing serial spans of at least 100 sections, which is where the power of the SWiFT approach is most useful.

Publications Resulting from This Work

    • Morgan JL, Berger DR, Wetzel AW, Lichtman JW (2016) The fuzzy logic of network connectivity in mouse visual thalamusCell
      165: 192-206 PMID: 27015312, PMC4808248
    • Hildebrand DGC, Torres RM, Choi Westward, Tran Minh Quan, Arthur Willis Wetzel, George ScottPlummer, Ruben Portugues, Isaac Henry Bianco, Owen Randlett, Stephan Saalfeld, Alex Baden, Kunal Lillaney, Randal Burns,Joshua Tzvi Vogelstein, Won-Ki Jeong, Jeff William Lichtman, Florian Engert (2016) Whole-brain serial-section electron microscopy in larval zebrafishNature, revised version in preparation.


xi) Morphological and regulatory models of neuronal differentiation

PIs: H Busch and Yard Böerries,University of Freiburg
Collaborators:R Murphy and G Rohde,Carnegie Mellon University

The goal of this project is to develop a spatiotemporal generative model of the changes in cell size, shape and subcellular organization that occur during the differentiation of PC12 cells induced by nerve growth cistron. This model volition exist related to a regulatory model constructed from parallel measurements of RNA expression over the time grade of differentiation. The results are expected to lead to the identification of gene expression changes that lead to specific morphological changes and to exam their importance for the differentiation process.

Publications Resulting from This Piece of work

    • Ruan X,Johnson GR, Bierschenk I, Nitschke R, Boerries M, Busch H, Irish potato RF (2020). Image-derived models of cell organization changes during differentiation and drug treatments.
      Mol Biol Cell
      31(vii):655-666. PMID: 31774723


12) Generative models of plant organelle distribution and differentiation

PI: K Palme,University of Freiburg
Collaborator:R Murphy,Carnegie Mellon University

Plants exhibit the power to undergo dramatic dedifferentiation and redifferentiation such that both protoplasts (cells extracted from mature plants) and microspores (cells that arise during gametogenesis) can give ascent to mature plants nether the right circumstances. The goal of this project is to identify the specific changes in protein expression and localization that are associated with such processes, in function by edifice spatiotemporal generative models straight from images and movies of cells obtained by light microscopy.

Publications Resulting from This Piece of work

    • Johnson GR, Kangas JD, Dovzhenko A, Trojok R, Voigt K, Majarian TD, Palme M, Irish potato RF. (2017) A method for characterizing phenotypic changes in highly variable cell populations and its awarding to high content screening of
      Arabidopsis thaliana
      Cytometry A
      91(four):326-335. PMID: 28245335


13) Microtubule pattern analysis and drug sensitivity

PI: P Giannakakou,Cornell UniversityCollaborator: R White potato,Carnegie Mellon University

The goal of this new project is to decide whether and how the distributions of microtubules differ between tumor cells sensitive and resistant to drugs that act on microtubules. Initial work has focused on preparation systems to recognize these patterns, with a complicating factor being the large differences in prison cell size, shape and unperturbed microtubule blueprint between cells from dissimilar tumors. Preliminary results indicate that three broad simply distinct patterns tin can be distinguished, respective to untreated cells, drug-treated resistant cells, and drug-treated sensitive cells. Future piece of work volition focus on using generative models to remove variation due to cell size and shape. Ultimately, this would exist used to determine whether a item tumor is likely to exist resistant to a particular drug and thereby to choose an appropriate drug for that tumor.


xiv) Actin filament patterns and cell motion

PI:J Theriot,Stanford UniversityCollaborators: Thou Rohde and D Slepčev,Carnegie Mellon University

This is a new project, the goal of which is to quantitatively describe the patterns of actin filaments and related proteins as they relate to cell motility in neutrophils. The project aims to apply recently adult methods for measuring the similarity between such patterns, as well equally transport-based method for finding correlation between intensity patterns, to decode relationships between different proteins as cells motility. Results are expected is to help characterize cell motility equally a role of the subcellular protein patterns.


sixteen) Latent Factor Models for Identification of Novel Neuroprotectives for Huntington’s Affliction

PI:Ivet Bahar and Bing Liu,Academy of Pittsburgh;
 Collaborators: Robert Friedlander, Mark Shurdak, Andrew Stern, D. Lansing Taylor,University of Pittsburgh

This C&SP is supported by NIH honour 5R01NS077748-04 (PI: Friedlander) entitled “Functional part of microRNAs in Huntington’s disease pathogenesis”. Our focus is the identification of novel neuroprotectives using latent gene modeling (LFM). Firstly, nosotros congenital a fix of known neuroprotectives based on the data in the literature by collecting compounds that were identified in a large scale screen for neuroprotectives in Huntington’s disease (Hard disk drive) also as compounds that take been tested in clinical trials for their neuroprotective result. And then, the LFM-based algorithm learns the most successful latent factor model of the drug-target interactions in the Run up dataset after assessing the performance of multiple LFM learning algorithms. The LFM of STITCH allowed us to quantitatively assess the interaction profiles of the known neuroprotectives, which we then used to find new compounds toward (i) the discovery of new neuroprotectives past target-based diversification of known neuroprotectives, (ii) the discovery of mechanism of activity hypotheses by identifying targets of involvement. Specifically, for each target of known neuroprotectives, nosotros identified other chemicals known to exist interacting with high confidence and sorted them for maximal contrast to the known neuroprotectives interacting with that target. This method identifies compounds that share the target of interest, but are otherwise every bit diverse in their interaction contour as possible. Compounds selected volition be active if the target of interest is of import for the HD etiology, therefore they represent new potential treatment candidates. Furthermore if i or more than targets yield many neuroprotectives, that target is implicated as existence central to the disease pathophysiology; therefore this method allows usa to infer about the proteins regulating the disease phenotype by testing compounds lonely.

Publications Resulting from This Work

      • Stern AM, Schurdak ME, Bahar I, Berg JM, Taylor DL (2016) A Perspective on Implementing a Quantitative Systems Pharmacology Platform for Drug Discovery and the Advocacy of Personalized Medicine.
        J Biomol Screen
        21(vi):521-34. PMID: 26962875
      • Taylor DL, Gough A, Schurdak ME, Vernetti L, Chennubhotla CS, Lefever D, Pei F, Faeder JR, Lezon TR, Stern AM, Bahar I. (2019) Harnessing Homo Microphysiology Systems as Key Experimental Models for Quantitative Systems Pharmacology.Handb Exp Pharmacol.260:327-367. doi: x.1007/164_2019_239.PMID: 31201557


18) Effect of RNA-editing of ADAR1 on the activity of ionotropic glutamate receptors

PI: Ivet Bahar, Mary H Cheng,University of Pittsburgh
 Collaborators:QingDe Wang,University of Pittsburgh

This is a C&SP, supported past NIH 5R21CA158650-02 (PI: Wang). The goal of the project is to define the function of RNA editing enzyme adenosine deaminase interim on RNA 1 (ADAR1) in hematopoietic and leukemia stem cells. ADAR1 is an essential protein for embryonic and adult hematopoiesis, while the leukemia cells are more susceptible to the gene deletion that codes ADAR1. The bear upon of ADAR1 on normal and leukemia stem cells (LSC) remain unknown, and there is a need to develop a therapeutic strategy to eliminate LSCs by targeting ADAR1. Although ADAR1 is identified as an RNA editing enzyme and RNA editing has been shown to play critical roles in stem cells and other biological processes, multiple attempts to appointment have been unable to place an editing target that accounts for the decease of normal hematopoietic and leukemia cells of ADAR1 knockouts. The R21 seeks a definitive reply whether the editing activity of ADAR1 is necessary for the proliferation and differentiation of LSCs.

ADAR has been found to modify the AMPA receptor GluR2 and G-protein coupling receptors, i.due east, SR2C and GABA receptor. The knockout animal models (Wang lab) have shown that this molecule is essential for embryonic survival and tissue homeostasis. However the molecular mechanism is largely unknown. The Bahar lab has initiated a new collaboration with the Wang Lab, to help investigate (a) the effects of the mutations induced upon ADAR1 editing on calcium channeling properties of iGluRs, (ii the issue of specific mutations on the functions of serotonin receptors and potassium channels, using molecular modeling, and (c) mathematical modeling and quantitative cess of the effect of suppressing cytosolic RNA pathway by ADAR1 on relevant cellular pathways networks.


21) Functional significance of the dynamics of AMPAR extracellular region

Collaborating Investigators: Ingo H. Greger,MRC Lab of Molecular Biology, Ivet Bahar, QingDe Wang,University of Pittsburgh, Tom Grand. Bartol, Terry J. Sejnowski,Salk Institute

Ionotropic glutamate receptor (iGluRs) are ligand-gated ion channels that permit for the menstruum of cations into the postsynaptic cell in response to glutamate binding, thus regulating neurotransmission upon depolarization of the jail cell membrane. Among iGluR subfamilies, AMPAR and NMDAR play a fundamental office in learning and memory, and in particular the AMPAR is essential to rapid neurotransmission and synaptic plasticity. The Greger and Bahar labs have been productively collaborating in recent years on AMPAR dynamics, beginning using the NTD dimer structures and more recently the intact tetrameric structures. These studies demonstrated that the NTD domains exhibit structural flexibilities comparable to those of AMPAR NTDs. Furthermore, the global modes of motions predicted by ANM (orProDy) revealed the propensity of homotetrameric AMPAR to assume more compact forms similar to NMDARs. The validity of these modes of motions were confirmed by cross-linking experiments between NTD sites predicted by ANM to come into shut proximity. In the new term, we volition first adopt ANM-based analysis to narrate the mode spectrum of the heterotetrameric AMPAR.ProDy analysis already revealed that the O ↔ Northward transition is enabled by a global ANM style. We volition characterize thoroughly the whole spectrum of motions and generate the energy landscape of Glu2/three heterotetramer, using the recently introduced extension of coMD. And so nosotros volition focus on the ECR motions that induce a pore opening (or cooperative twisting) at the TMD and analyze the conformational events that enable the allosteric coupling between the ECR and the TMD with the help of accelerated Md simulations. In the next phase, we plan to examine the significance of GluA2/3 ECR flexibility in adapting to its interactions with auxiliary proteins such as cornichon homologs, TARPs or in forming clusters, which volition exist further tested/validated with structural and unmarried-particle tracking methods in the Greger lab.

Publications Resulting from This Work

    • Krieger J,Bahar I,Greger IH (2015) Structure, Dynamics, and Allosteric Potential of Ionotropic Glutamate Receptor N-Last DomainsBiophys J109: 1136-48 PMID: 26255587, PMC45761612
    • Dutta A, Krieger J, Lee JY, Garcia-Nafria J,Greger IH,Bahar I (2015) Cooperative Dynamics of Intact AMPA and NMDA Glutamate Receptors: Similarities and Subfamily-Specific DifferencesConstruction23: 1692-1704 PMID: 26256538, PMC4558295
    • Dutta A, Shrivastava IH, Sukumaran M,Greger IH,Bahar I (2012) Comparative dynamics of NMDA- and AMPA-Glutamate receptor N-final domainsStructure20: 1838-49 PMID: 22959625, PMC3496038
    • Lee JY, Krieger J, Herguedas B, García-Nafría J, Dutta A, Shaikh SA, Greger IH, Bahar I. (2019) Druggability Simulations and X-ray Crystallography Reveal a Ligand-binding Site in the GluA3 AMPA Receptor Due north-final Domain.Structure
      27: 241-252.
    • Krieger J, Lee JY, Greger IH, Bahar I. (2019) Activation and Desensitization of Ionotropic Glutamate Receptors by Selectively Triggering Pre-existing Motions.Neurosci Lett700: 22-29 PMID: 29481851 PMCID: 6107436
    • Lee JY, Krieger J, Herguedas B, García-Nafría J, Dutta A, Shaikh SA, Greger IH, Bahar I. (2019) Druggability Simulations and Ten-Ray Crystallography Reveal a Ligand-Binding Site in the GluA3 AMPA Receptor N-Terminal Domain.
      Structure. 27(ii):241-252 PMID: 30528594


23) CaMKII Structural Dynamics

Collaborating Investigators: John Kuriyan,Academy of California-Berkeley, Mary Kennedy,Caltech, Ivet Bahar, James R. Faeder,Academy of Pittsburgh, Tom M. Bartol, Terry J. Sejnowski,Salk Establish

This C&SP will focus on studying mechanisms of function of the complex biomolecular assembly of CaMKII and its dynamics. Nosotros recently showed that the human alpha isoform, CaMKIIalpha, literally acts as a device for propagating the activation by calcium signaling. This is achieved through subunit exchanges betwixt activated CaMKII and holoenzyme assemblies that have non been activated. The spread of activation through these exchanges now suggests that activation can be maintained long later the disappearance of the initial signal. This is remarkable because information technology reveals a possible mechanism by which CaMKII assists in the induction of long-term retention in the brain. Information technology is clear that the phosphorylation of the calmodulin-recognition element triggers the exchange. But in addition to localized interactions, there is something unique near the overall compages of CaMKII then that it accommodates different oligomerization states. Starting from early on crystal structures and EM images, we have seen that the holoenzyme could assume both dodecameric and tetradecameric forms. Our new observations now show that they coexist. An intriguing observation was: why does the vertical dimer dissociate, rather than a single subunit, or a horizontal dimer? ANM results unambiguously show that the dodecamer structure indeed favors the vertical dimers to be released. The type of assay performed by ProDy can open the way to a better understanding of CaMKII dynamics. Additionally, coevolution analysis may give us additional information on interfacial interactions that modulate subunit exchange. These studies will aid further understanding the dynamics and interactions of CaMKII.


25) Efficient parallel sampling at multiple scales using the weighted ensemble strategy

Collaborating Investigators: Lillian T. Chong,University of Pittsburgh, Daniel Zuckerman,Oregon Health and Scientific discipline University

There is a “silicon ceiling” that ultimately limits many, if not most, types of dynamical biological simulations. That is, even the world’southward most powerful computers cannot generate sufficiently long simulations, whether for atomistic models of proteins or for realistic models of cell behavior. In many cases, the key events may occur across simulation timescales – such as protein folding, conformational transitions of proteins, assembly of protein complexes, or transitions of cell behavior from healthy to pathological states. The work in this C&SP will continue the development of WESTPA, a tool for controlling other software tools: information technology orchestrates up to thousands of trajectories run natively by other software at any scale (e.1000., Gromacs, Amber, BioNetGen, MCell) using a “weighted ensemble” strategy. Non simply does WESTPA parallelize the apply of dynamics engines – but because of the statistical process past which trajectories are added and removed, WESTPA can obtain estimates of key kinetic every bit well equally equilibrium observables in significantly less computing fourth dimension than would be required in ordinary parallelization. Our aims are to improve the ease of use and interoperability of WESTPA; to improve its functioning and reliability; to demonstrate the effectiveness of WESTPA through a serial of “showcase” examples from molecular to cellular scale using a variety of dynamics engines; and to meliorate instructional materials based on the showcase examples.

Publications Resulting from This Work

    • Zwier MC, Adelman JL, Kaus JW, Pratt AJ, Wong KF, Rego NB, Suárez E, Lettieri S, Wang DW, Grabe M, Zuckerman DM, Chong LT (2015) WESTPA: an interoperable, highly scalable software package for weighted ensemble simulation and analysisJ Chem Theory Comput 11: 800-809. PMID: 26392815; PMC4573570
    • Suárez Eastward, Pratt AJ, Chong LT, Zuckerman DM (2016) Estimating First Passage Time Distributions from Weighted Ensemble Simulations and non-Markovian AnalysesProtein Scientific discipline 25: 67-78. PMID: 26131764; PMC4815309
    • Suárez E, Pratt AJ, Chong LT, Zuckerman DM (2016) Estimating First Passage Fourth dimension Distributions from Weighted Ensemble Simulations and non-Markovian AnalysesPoly peptide Science 25: 67-78. PMID: 26131764; PMC4815309


26) GPCR signaling and interactions. Learning from SILAC/proteomics

Collaborating Investigators: Kunhong (Kevin) Xiao, Ivet Bahar, James R. Faeder,University of Pittsburgh

This focus of this C&SP will focus on the development and awarding of quantitative proteomics and systems biology approaches to study GPCR (G protein couple receptor) structures, their signaling networks, and regulation. The approach will be to use mass spectrometry-based quantitative proteomics, in combination with systems, chemical, and structural biology to study the receptor construction-office relationship, macromolecular interactions, and the expression and posttranslational modifications of signaling molecules downstream of GPCR. The ultimate goal of this research is to provide a better understanding of structure-role relationship of GPCRs and their related proteins and provide tools and resources for structure-based drug design.


28) Dynamic modulation of interferon bounden affinity as a mechanism to regulate interferon receptor signaling

Collaborating Investigators: Gideon Schreiber,Weizmann Found, Ivet Bahar, James R. Faeder,University of Pittsburgh

Type I interferons (IFNs) are multifunctional cytokines that mediate/induce various cellular responses, including both innate and adaptive immune responses, stimulation of antiviral responses, and cancer surveillance, upon forming a ternary complex with 2 surface receptors, IFNAR1 and IFNAR2. The activities of IFN-a subtypes correlate with their affinities to demark to IFNAR1 and IFNAR2. While the Schreiber lab made seminal contributions to understanding the molecular basis of IFNARs, the mechanism of regulation of differential IFN activities through interactions with IFNAR1 and 2, remains unclear. Our integrated computational (TR&D1) and experimental preliminary studies point to the significance of the intrinsic dynamics in modulating binding affinity. We adopted a closely integrated computational/experimental strategy that yielded promising results, which we are currently further pursuing and that illustrate the adaptability of proteins to different jump states or to sequence variations/mutations via their softest modes of move. Further cross-linking, fluorescence quenching and cistron induction experiments will be conducted in the Schreiber lab, in shut coordination with TR&D1 computational studies at the Bahar lab.

Publications Resulting from This Work

  • Li H, Sharma Due north, General IJ, Schreiber G, Bahar I. (2017) Dynamic Modulation of Binding Analogousness as a Machinery for Regulating Interferon Signaling.J Mol Biol429: 2571-2589 PMID: 28648616 PMCID: 5545807


32) Integrating compartmental dominion-based modeling into VCell

Collaborating Investigators: James C. Shcaff, Mikhail L. Blinov, Ion I. Moraru, Lew Loew,University of Connecticut, James R. Faeder,Academy of Pittsburgh

This C&SP is a collaboration with The National Resource for Cell Analysis and Modeling (NRCAM), which develops new technologies for modeling cell biological processes. The technologies are integrated through Virtual Cell (VCell), a trouble solving environment built on a cardinal database and disseminated as a customer-server application. It will develop methods to simulate multi-molecular interactions at a sub-cellular scale for issues where molecular shape and cellular geometry influence the system behavior. MMBioS investigators volition interact to integrate compartmental rule-based modeling into the VCell platform.


C&SP Summary

Collaboration & Service Projects Summary

C&SP # and title

Collaborator PI (due south)
and co-Is





MMBioS Leader


External Funding

(source/grant code, title, and start-terminate dates)

v. Modeling immunoreceptor signaling, autophagy, and endocytosis

William Hlavacek

U of New Mexico, Los Alamos Natl lab



NCI, 5R01-GM111510-02, Hardening Software for Rule-Based Modeling (Hlavacek)


7. Interfacing image-derived generative models with cell simulation engines

Michael Hucka,

Leslie One thousand. Loew,

James C. Schaff

Caltech, U Conn

2, 3, 4



Bartol, Hood


National Resources for Jail cell Analysis and Modeling (Loew)


8. Using generative models of cell organization to investigate drug mechanisms and tumor cell heterogeneity

Steven J.


Lani Wu



White potato

NCI: R01-CA133253-06:

Lung Cancer Heterogeneity and its Bear upon on Drug Resistance


15. Reconstructing zebrafish neural circuits controlling visually induced behaviors

Florian Engert,

David L. Hildebrand,




NINDS: 5U01-NS090449

Neural Circuits in Zebrafish Form, Part and Plasticity (Engert) 9/30/fourteen-seven/31/17

16. Latent factor models for identification of novel neuroprotectives for Huntington’s Disease

Robert M.


ME. Schurdak,

Andrew M. Stern, D. Lansing Taylor

U of Pittsburgh,





Functional Role of Micro RNAS in Huntington’s Disease Pathogenesis (Friedlander) 2/15/12-1/31/17

17. NMJ Calcium signaling

Stephen D. Meriney

Peter Wipf

U of Pittsburgh

1, 2

Bartol, Bahar,

Hood, Cheng,

Laghaei, Czech

NINDS, 5R01-NS090644-02,

CRCNS: Transmitter Release Site Organization in Plasticity and Disease at the NMJ 8/1/14-5/31/xix

18.Effect of RNA-editing of ADAR1 on the activeness of ionotropic glutamate receptors

QingDe Wang




NIH 5R21CA158650-02

Role of a novel molecule ADAR1 in endothelial cells for angiogenesis iv/15/2016 – 3/31/2018

19. Mapping the supramolecular organization of dendritic spines to model the regulation of synaptic signal transfer

Mark H. Ellisman

Andreas Herz


Ludwig-Maximilians U, Munich, Deutschland

2, iv



NIDA: R01-DA038896-02:

CRCNS: Deciphering the dynamical multiscale structure-function of dendritic spines 7/1/xiv-6/30/nineteen

20. Structural plasticity of chaperonins adamant by cryo-electron microscopy: Modeling the mechanism of GroEL and TRiC

Wah Chiu

Baylor Higher of Medicine

1, 4


P01NS092525 “From structure to therapy: the TRiC chaperonin network in Huntington’due south disease” (Chiu, Co-PI) 4/1/2016-31/three/2021

5P41-GM103832-31: 3D Electron Microscopy of

Macromolecules 12/1/96-12/31/xix

22. Modeling T cell fate decisions

Penelope Morel,
Robin Eastward. C. Lee

U of Pittsburgh



Juvenile Diabetes Research

Foundation 1-INO-2016-215-A-N

(Morel) 06/01/2016-05/31/2017

23. CaMKII structural dynamics

John Kuriyan


ane, two, 3


Faeder Sejnowski,


24. Spatio-temporal cell biology

Alan Horwitz,

Gregory Johnson

Allen Institute



The Allen Foundation has provided $100 million to create the Allen Institute for Cell Science to support the collection of prison cell images

26. GPCR signaling and interactions. Learning from

SILAC/ proteomics

Kunhong (Kevin) Xiao

U Pitt

1, iii

Bahar, Faeder

NHLBI: 2P01-HL07544311, Novel

Mechanisms and Therapies in Centre Failure (PI: Rockman, Duke U) Proteomics and Structure Core (PI: Xiao)
viii/ane/2015 7/31/2020

27. Circuit reconstruction of association cortex




NIDCD: 5R03-DC013622-03: Network

Anatomy of Olfactory Processing


28. Dynamic modulation of interferon binding affinity as a machinery to regulate interferon receptor signaling

Gideon Schreiber

Weizmann Found, Israel

1, 3

Bahar, Faeder

ISF (State of israel Science Foundation) Eye of Inquiry Excellence for

Integrated Structural Prison cell Biological science,


29. Spatiotemporal models of autophagy

Joern Dengjel

Academy of Frieburg



Swiss National Fund 31003A_ 166482/1 Stimulus and time dependent poly peptide dynamics in autophagy analyzed by quantitative MS

30. Modeling of Tetrahymena basal body dynamics

Chad K. Pearson

University of Colorado,




5R01GM099820-04, Mechanisms of Centriole Assembly and Stability ix/24/12-7/31/17

31. Receptor Clustering and the Kinetics of T Cell-Mediated Killing

Yuri Sykulev

Thomas Jefferson University



5R21AI113819-02 Exploiting an Bogus APC to Induce Dissimilar T Subsets ane/one/15-12/31/sixteen

32. Integrating compartmental rule-based modeling into VCell


Les Loew

U Connecticut Wellness Center




5P41-GM103313-18: National Resource for Prison cell Analysis

and Modeling (Loew) 9/xxx/98-4/30/17

33. Integration of rule-based modeling capabilities with pySB modeling platform

Carlos F. Lopez




34. Development of a high-level, rule-based whole-jail cell modeling language

Jonathan Karr

Mt. Sinai



NSF 1548123L, ERASynBio:

MiniCell – A Model-driven approach to minimal cell engineering science (Karr) 12/17/2015 – 7/31/2018

C&SP34: Evolution of a loftier-level, rule-based whole-cell modeling language


Collaborating Investigators:

Jonathan Karr

James R. Faederii




Mt. Sinai and
twoUniversity of Pittsburgh


Funding Condition of Project:

NSF grant
1548123 “

ERASynBio: MiniCell – A Model-driven Approach to Minimal Cell Engineering”

(Karr) 12/17/2015–7/31/2018


Biomedical Research Problem:

Despite decades of enquiry and the growing wealth of data, we do not have a unified understanding of cell biological science. Whole-cell (WC) models that represent every factor role and that predict the cell cycle dynamics of every molecular species have the potential to unify our understanding of cell biology.


Recently, Karr and others adult the beginning WC model.


This was achieved by combining multiple mathematically distinct submodels of private cellular pathways, and by developing custom software to simulate the model. However, this
ad hoc
approach is not easily reusable, expandable, or reproducible.


New WC modeling tools are needed to build better models that tin can guide bioengineering and medicine.

Fig VIII.9

Transcription factor binding in

Rules ascertain specific sequences of DNA and protein-DNA interactions in terms of Deoxyribonucleic acid-sequence motifs and specific proteins. Evaluation of WC-Lang rules generates bounden sites on specific sequences.

Sequences and proteins are instantiated as BioNetGen species and rules.

This system can be efficiently simulated using NFsim.

Due east.

Methods and Procedures:

This project will develop a high-level rule-based WC modeling language and a rule-based multi-algorithm simulator. The language will enable researchers to compactly describe biological processes, such as the translation of each amino acid, using high-level rule patterns and sequence, genomic, and biochemical data. WC-Lang will provide loftier-level data-based rules to succinctly describe large reactions networks in terms of reaction patterns and sequence, genomic, and other data, that will be encoded in BioNetGen.


This loftier-level language will make information technology easier for researchers to develop and communicate WC models, and the BioNetGen implementation will get in possible to apace develop a reusable and efficient WC model simulator that will have advantage of BioNetGen’south network generation as well as the network-free simulation capabilities of NFsim.


WC-Lang will be implemented as a Python library which encodes the loftier-level rules into lower-level BioNetGen rules. Utilise of rules to encode molecular interactions will open many possibilities for the visualization of WC models using contact maps and regulatory graphs (meet
Aim 2

). In the kickoff phase of this project we will focus on implementing modules from a reference WC model the Karr lab is building that will exist described completely independently from whatever simulation code. This is a major first step in the evolution of WC-lang. We will develop lawmaking to translate reference model modules into BioNetGen language code (
Fig. Eight.9
) and test simulation results confronting those obtained with reference simulators. In the second stage, we will aggrandize BioNetGen and NFSim to support multi-algorithm modeling, which will be enabled past development of the libasal bodyNG API in
Aim 3.1

. Submodels will be implemented as singled-out processes, which is already possible for existing interfaces to BioNetGen and NFsim, and these volition in turn be controlled by the multi-algorithm simulator that the Karr lab is developing in the in a higher place-referenced NSF-funded project. Together, the new language and expanded simulator will enable researchers to build much larger and more accurate WC models that tin guide bioengineering and precision medicine.