Cyrus A. Wilson

Normal mode analysis of macromolecular motions in a database framework: Developing mode concentration as a useful classifying statistic

We investigated protein motions using normal modes within a database framework, determining on a large sample the degree to which normal modes anticipate the direction of the observed motion and were useful for motions classification. As a starting point for our analysis, we identified a large number of examples of protein flexibility from a comprehensive set of structural alignments of the proteins in the PDB. Each example consisted of a pair of proteins that were considerably different in structure given their sequence similarity. On each pair, we performed geometric comparisons and adiabatic‐mapping interpolations in a high‐throughput pipeline, arriving at a final list of 3,814 putative motions and standardized statistics for each. We then computed the normal modes of each motion in this list, determining the linear combination of modes that best approximated the direction of the observed motion. We integrated our new motions and normal mode calculations in the Macromolecular Motions Database, through a new ranking interface at http://molmovdb.org. Based on the normal mode calculations and the interpolations, we identified a new statistic, mode concentration, related to the mathematical concept of information content, which describes the degree to which the direction of the observed motion can be summarized by a few modes. Using this statistic, we were able to determine the fraction of the 3,814 motions where one could anticipate the direction of the actual motion from only a few modes. We also investigated mode concentration in comparison to related statistics on combinations of normal modes and correlated it with quantities characterizing protein flexibility (e.g., maximum backbone displacement or number of mobile atoms). Finally, we evaluated the ability of mode concentration to automatically classify motions into a variety of simple categories (e.g., whether or not they are “fragment‐like”), in comparison to motion statistics. This involved the application of decision trees and feature selection (particular machine‐learning techniques) to training and testing sets derived from merging the “list” of motions with manually classified ones. Proteins 2002;48:682–695.

Myosin II contributes to cell-scale actin network treadmilling through network disassembly

Crawling locomotion of eukaryotic cells is achieved by a process dependent on the actin cytoskeleton: protrusion of the leading edge requires assembly of a network of actin filaments, which must be disassembled at the cell rear for sustained motility. Although ADF/cofilin proteins have been shown to contribute to actin disassembly, it is not clear how activity of these locally acting proteins could be coordinated over the distance scale of the whole cell. Here we show that non-muscle myosin II has a direct role in actin network disassembly in crawling cells. In fish keratocytes undergoing motility, myosin II is concentrated in regions at the rear with high rates of network disassembly. Activation of myosin II by ATP in detergent-extracted cytoskeletons results in rear-localized disassembly of the actin network. Inhibition of myosin II activity and stabilization of actin filaments synergistically impede cell motility, suggesting the existence of two disassembly pathways, one of which requires myosin II activity. Our results establish the importance of myosin II as an enzyme for actin network disassembly; we propose that gradual formation and reorganization of an actomyosin network provides an intrinsic destruction timer, enabling long-range coordination of actin network treadmilling in motile cells.

Actin–myosin network reorganization breaks symmetry at the cell rear to spontaneously initiate polarized cell motility

We have analyzed the spontaneous symmetry breaking and initiation of actin-based motility in keratocytes (fish epithelial cells). In stationary keratocytes, the actin network flow was inwards and radially symmetric. Immediately before motility initiation, the actin network flow increased at the prospective cell rear and reoriented in the perinuclear region, aligning with the prospective axis of movement. Changes in actin network flow at the cell front were detectable only after cell polarization. Inhibition of myosin II or Rho kinase disrupted actin network organization and flow in the perinuclear region and decreased the motility initiation frequency, whereas increasing myosin II activity with calyculin A increased the motility initiation frequency. Local stimulation of myosin activity in stationary cells by the local application of calyculin A induced directed motility initiation away from the site of stimulation. Together, these results indicate that large-scale actin–myosin network reorganization and contractility at the cell rear initiate spontaneous symmetry breaking and polarized motility of keratocytes.

Emergence of Large-Scale Cell Morphology and Movement from Local Actin Filament Growth Dynamics

Variations in cell migration and morphology are consequences of changes in underlying cytoskeletal organization and dynamics. We investigated how these large-scale cellular events emerge as direct consequences of small-scale cytoskeletal molecular activities. Because the properties of the actin cytoskeleton can be modulated by actin-remodeling proteins, we quantitatively examined how one such family of proteins, enabled/vasodilator-stimulated phosphoprotein (Ena/VASP), affects the migration and morphology of epithelial fish keratocytes. Keratocytes generally migrate persistently while exhibiting a characteristic smooth-edged “canoe” shape, but may also exhibit less regular morphologies and less persistent movement. When we observed that the smooth-edged canoe keratocyte morphology correlated with enrichment of Ena/VASP at the leading edge, we mislocalized and overexpressed Ena/VASP proteins and found that this led to changes in the morphology and movement persistence of cells within a population. Thus, local changes in actin filament dynamics due to Ena/VASP activity directly caused changes in cell morphology, which is coupled to the motile behavior of keratocytes. We also characterized the range of natural cell-to-cell variation within a population by using measurable morphological and behavioral features—cell shape, leading-edge shape, filamentous actin (F-actin) distribution, cell speed, and directional persistence—that we have found to correlate with each other to describe a spectrum of coordinated phenotypes based on Ena/VASP enrichment at the leading edge. This spectrum stretched from smooth-edged, canoe-shaped keratocytes—which had VASP highly enriched at their leading edges and migrated fast with straight trajectories—to more irregular, rounder cells migrating slower with less directional persistence and low levels of VASP at their leading edges. We developed a mathematical model that accounts for these coordinated cell-shape and behavior phenotypes as large-scale consequences of kinetic contributions of VASP to actin filament growth and protection from capping at the leading edge. This work shows that the local effects of actin-remodeling proteins on cytoskeletal dynamics and organization can manifest as global modifications of the shape and behavior of migrating cells and that mathematical modeling can elucidate these large-scale cell behaviors from knowledge of detailed multiscale protein interactions.

Estimating specular roughness and anisotropy from second order spherical gradient illumination

This paper presents a novel method for estimating specular roughness and tangent vectors, per surface point, from polarized second order spherical gradient illumination patterns. We demonstrate that for isotropic BRDFs, only three second order spherical gradients are sufficient to robustly estimate spatially varying specular roughness. For anisotropic BRDFs, an additional two measurements yield specular roughness and tangent vectors per surface point. We verify our approach with different illumination configurations which project both discrete and continuous fields of gradient illumination. Our technique provides a direct estimate of the per‐pixel specular roughness and thus does not require off‐line numerical optimization that is typical for the measure‐and‐fit approach to classical BRDF modeling.

Circularly polarized spherical illumination reflectometry

We present a novel method for surface reflectometry from a few observations of a scene under a single uniform spherical field of circularly polarized illumination. The method is based on a novel analysis of the Stokes reflectance field of circularly polarized spherical illumination and yields per-pixel estimates of diffuse albedo, specular albedo, index of refraction, and specular roughness of isotropic BRDFs. To infer these reflectance parameters, we measure the Stokes parameters of the reflected light at each pixel by taking four photographs of the scene, consisting of three photographs with differently oriented linear polarizers in front of the camera, and one additional photograph with a circular polarizer. The method only assumes knowledge of surface orientation, for which we make a few additional photometric measurements. We verify our method with three different lighting setups, ranging from specialized to off-the-shelf hardware, which project either discrete or continuous fields of spherical illumination. Our technique offers several benefits: it estimates a more detailed model of per-pixel surface reflectance parameters than previous work, it requires a relatively small number of measurements, it is applicable to a wide range of material types, and it is completely viewpoint independent.

Temporal upsampling of performance geometry using photometric alignment

We present a novel technique for acquiring detailed facial geometry of a dynamic performance using extended spherical gradient illumination. Key to our method is a new algorithm for jointly aligning two photographs, under a gradient illumination condition and its complement, to a full-on tracking frame, providing dense temporal correspondences under changing lighting conditions. We employ a two-step algorithm to reconstruct detailed geometry for every captured frame. In the first step, we coalesce information from the gradient illumination frames to the full-on tracking frame, and form a temporally aligned photometric normal map, which is subsequently combined with dense stereo correspondences yielding a detailed geometry. In a second step, we propagate the detailed geometry back to every captured instance guided by the previously computed dense correspondences. We demonstrate reconstructed dynamic facial geometry, captured using moderate to video rates of acquisition, for every captured frame.

Developing INOTS to support interpersonal skills practice

The Immersive Naval Officer Training System (INOTS) is a blended learning environment that merges traditional classroom instruction with a mixed reality training setting. INOTS supports the instruction, practice and assessment of interpersonal communication skills. The goal of INOTS is to provide a consistent training experience to supplement interpersonal skills instruction for Naval officer candidates without sacrificing trainee throughput and instructor control. We developed an instructional design from cognitive task analysis interviews with experts to serve as a framework for system development. We also leveraged commercial student response technology and research technologies including natural language recognition, virtual humans, realistic graphics, intelligent tutoring and automated instructor support tools. In this paper, we describe our methodologies for developing a blended learning environment, and our challenges adding mixed reality and virtual human technologies to a traditional classroom to support interpersonal skills training.1 2

Semantically-aware blendshape rigs from facial performance measurements

We present a framework for automatically generating personalized blendshapes from actor performance measurements, while preserving the semantics of a template facial animation rig. Firstly, we capture various poses from the subject with our photogrammetry apparatus. The 3D reconstruction from each pose is then corresponded by an image-based tracking algorithm. The core of our framework is an optimization algorithm which iteratively refines the initial estimation of the blendshapes such that they can fit the performance measurements better. This framework facilitates creation of an ensemble of realistic digital-double face rigs for each individual with consistent behavior across the character set.

Facial cartography: interactive scan correspondence

We present a semi-automatic technique for computing surface correspondences between 3D facial scans in different expressions, such that scan data can be mapped into a common domain for facial animation. The technique can accurately correspond high-resolution scans of widely differing expressions -- without requiring intermediate pose sequences -- such that they can be used, together with reflectance maps, to create high-quality blendshape-based facial animation. We optimize correspondences through a combination of Image, Shape, and Internal forces, as well as Directable forces to allow a user to interactively guide and refine the solution. Key to our method is a novel representation, called an Active Visage, that balances the advantages of both deformable templates and correspondence computation in a 2D canonical domain. We show that our semi-automatic technique achieves more robust results than automated correspondence alone, and is more precise than is practical with unaided manual input.