Putting the Pieces Together: Integrative Modeling Platform Software for Structure Determination of Macromolecular Assemblies

Russel, Daniel; Lasker, Keren; Webb, Ben; Velázquez-Muriel, Javier; Tjioe, Elina; Stroopinsky, Dina; Peterson, Benjamin; Šali, Andrej

doi:10.1371/journal.pbio.1001244

Cited by 492 publications

(552 citation statements)

References 49 publications

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“…1, 7, and 8). The integrative structure modeling protocol ( i.e ., stages 2, 3, and 4) was scripted using the Python Modeling Interface (PMI) package, version 4d97507, a library for modeling macromolecular complexes based on our open-source Integrative Modeling Platform (IMP) package 98 , version 2.6 (https://integrativemodeling.org). The current procedure is an updated version of previously described protocols 66,72,90,101–104 .…”

Section: Methodsmentioning

confidence: 99%

“…Distributions of the FG repeats and NTRs were computed by Brownian dynamics simulation 145 , using our protocol 146 implemented in IMP 98 , version 2.6. The simulated system included the static NPC ring determined in this study, the pore membrane, disordered and flexible FG repeat domains, as well as freely diffusing NTRs and inert macromolecules, all enclosed within a bounding box of 2,000 × 2,000 × 2,000 Å 3 .…”

Section: Methodsmentioning

confidence: 99%

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Integrative structure and functional anatomy of a nuclear pore complex

Kim

Fernández-Martı́nez

Nudelman

et al. 2018

Nature

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478

894

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Summary Despite the central role of Nuclear Pore Complexes (NPCs) as gatekeepers of RNA and protein transport between the cytoplasm and nucleoplasm, their large size and dynamic nature have impeded a full structural and functional elucidation. Here, we have determined a subnanometer precision structure for the entire 552-protein yeast NPC by satisfying diverse data including stoichiometry, a cryo-electron tomography map, and chemical cross-links. The structure reveals the NPC’s functional elements in unprecedented detail. The NPC is built of sturdy diagonal columns to which are attached connector cables, imbuing both strength and flexibility, while tying together all other elements of the NPC, including membrane-interacting regions and RNA processing platforms. Inwardly-directed anchors create a high density of transport factor-docking Phe-Gly repeats in the central channel, organized in distinct functional units. Taken together, this integrative structure allows us to rationalize the architecture, transport mechanism, and evolutionary origins of the NPC.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Integrative structure and functional anatomy of a nuclear pore complex

Kim

Fernández-Martı́nez

Nudelman

et al. 2018

Nature

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478

894

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“…We use simulated annealing dynamics and conjugate gradient optimizations to generate a population of 3D genome structures X for which all of the chromatin contacts in W are physically realized in the genome structures, indicating that the likelihoods of all contacts in the structure population are maximized to approximately one. We implemented the structure optimization tools within the Integrated Modeling Platform (33,34). We applied our method to human lymphoblastoid cells, using TCC experiments with a fivefold increase in sequencing coverage in comparison with our work reported in ref.…”

Section: Population-based Genome Structure Modeling By Maximum Likelimentioning

confidence: 99%

Population-based 3D genome structure analysis reveals driving forces in spatial genome organization

Tjong

Kalhor

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

196

232

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Conformation capture technologies (e.g., Hi-C) chart physical interactions between chromatin regions on a genome-wide scale. However, the structural variability of the genome between cells poses a great challenge to interpreting ensemble-averaged Hi-C data, particularly for long-range and interchromosomal interactions. Here, we present a probabilistic approach for deconvoluting Hi-C data into a model population of distinct diploid 3D genome structures, which facilitates the detection of chromatin interactions likely to co-occur in individual cells. Our approach incorporates the stochastic nature of chromosome conformations and allows a detailed analysis of alternative chromatin structure states. For example, we predict and experimentally confirm the presence of large centromere clusters with distinct chromosome compositions varying between individual cells. The stability of these clusters varies greatly with their chromosome identities. We show that these chromosome-specific clusters can play a key role in the overall chromosome positioning in the nucleus and stabilizing specific chromatin interactions. By explicitly considering genome structural variability, our population-based method provides an important tool for revealing novel insights into the key factors shaping the spatial genome organization. (14), and single-cell (15) and in situ Hi-C (16)], close chromatin contacts can now be identified at increasing resolution, providing new insight into genome organization. These methods measure the relative frequencies of chromosome interactions averaged over a large population of cells. However, individual 3D genome structures can vary dramatically from cell to cell even within an isogenic sample, especially with respect to long-range interactions (15,17,18). This structural variability poses a great challenge to the interpretation of ensemble-averaged Hi-C data (14,(19)(20)(21)(22)(23) and prevents the direct detection of cooperative interactions co-occurring in the same cell. This problem is particularly evident for long-range (cis) and interchromosomal (trans) interactions, which are generally observed at relatively low frequencies and are therefore present only in a small subset of individual cells at any given time (3,11,15). Despite their low frequencies, long-range and interchromosome interaction patterns are not random noise. In fact, these interactions are more informative than short-range interactions in determining the global genome architectures in cells and are often functionally relevant-interactions between transcriptionally active regions are often interchromosomal in nature (14). Owing to their variable nature, long-range and trans interactions can be part of alternative, structurally different conformations, which makes their interpretation in form of consensus structures impossible. However, inferring which of the long-range interactions co-occur in the same cell from ensemble Hi-C data remains a major challenge.These challenges cannot be easily overcome even by the new single-cell Hi-C techno...

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“…An integrative approach to structure determination combines spatial restraints derived from multiple experimental techniques together with theoretical and statistical knowledge (22)(23)(24)(25). Here, we propose including spatial restraints derived from EM class averages into such an integrative approach.…”

Section: Discussionmentioning

confidence: 99%

Assembly of macromolecular complexes by satisfaction of spatial restraints from electron microscopy images

Velázquez-Muriel

Lasker

Russel

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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To obtain a structural model of a macromolecular assembly by single-particle EM, a large number of particle images need to be collected, aligned, clustered, averaged, and finally assembled via reconstruction into a 3D density map. This process is limited by the number and quality of the particle images, the accuracy of the initial model, and the compositional and conformational heterogeneity. Here, we describe a structure determination method that avoids the reconstruction procedure. The atomic structures of the individual complex components are assembled by optimizing a match against 2D EM class-average images, an excluded volume criterion, geometric complementarity, and optional restraints from proteomics and chemical cross-linking experiments. The optimization relies on a simulated annealing Monte Carlo search and a divide-and-conquer message-passing algorithm. Using simulated and experimentally determined EM class averages for 12 and 4 protein assemblies, respectively, we show that a few class averages can indeed result in accurate models for complexes of as many as five subunits. Thus, integrative structural biology can now benefit from the relative ease with which the EM class averages are determined.integrative modeling | structural determination | computational biology E M is an increasingly useful approach for structural characterization of macromolecular assemblies (1-3). Different flavors of EM include electron crystallography, single-particle EM, and electron tomography (4), although tomography is generally limited to resolutions worse than 20 Å. Single-particle EM can be used with negative-stained or cryogenically frozen (cryo-EM) samples. Cryo-EM particularly presents some attractive advantages: It preserves a near-native conformation of the molecules, it can be applied to the study of large assemblies, and it can theoretically achieve atomic resolution (5). Currently, the standard way to analyze single-particle EM data is to align collected 2D single-particle images, cluster the images, calculate a 2D classaverage image for each cluster, and finally perform 3D reconstruction to obtain a 3D density map. Pseudoatomic models for many assemblies have been generated by fitting X-ray crystallographic structures and/or comparative models of the individual components into a density map of the whole assembly (6).Obtaining a high-resolution density map requires a large number of single-particle images and depends critically on determining an initial low-resolution density map as a template for reconstruction, as well as a high signal-to-noise ratio (SNR) in the particle images (7-10). Several methods exist to obtain the template map: randomconical reconstruction (11), common lines determination (12)(13)(14), and maximization of the posterior probability of observing the set of class averages (15). A challenge for template construction arises when the available class averages do not provide significant coverage of all the orientations of the complex; in such a case, an accurate template map cannot be compute...

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Putting the Pieces Together: Integrative Modeling Platform Software for Structure Determination of Macromolecular Assemblies

Cited by 492 publications

References 49 publications

Integrative structure and functional anatomy of a nuclear pore complex

Integrative structure and functional anatomy of a nuclear pore complex

Population-based 3D genome structure analysis reveals driving forces in spatial genome organization

Assembly of macromolecular complexes by satisfaction of spatial restraints from electron microscopy images

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