TomoJ: tomography software for three-dimensional reconstruction in transmission electron microscopy

<p>Abstract</p> <p>Background</p> <p>Transmission electron tomography is an increasingly common three-dimensional electron microscopy approach that can provide new insights into the structure of subcellular components. Transmission electron tomography fills the gap between high resolution structural methods (X-ray diffraction or nuclear magnetic resonance) and optical microscopy. We developed new software for transmission electron tomography, TomoJ. TomoJ is a plug-in for the now standard image analysis and processing software for optical microscopy, ImageJ.</p> <p>Results</p> <p>TomoJ provides a user-friendly interface for alignment, reconstruction, and combination of multiple tomographic volumes and includes the most recent algorithms for volume reconstructions used in three-dimensional electron microscopy (the algebraic reconstruction technique and simultaneous iterative reconstruction technique) as well as the commonly used approach of weighted back-projection.</p> <p>Conclusion</p> <p>The software presented in this work is specifically designed for electron tomography. It has been written in Java as a plug-in for ImageJ and is distributed as freeware.</p

Error analysis in the determination of the electron microscopical contrast transfer function parameters from experimental power Spectra

<p>Abstract</p> <p>Background</p> <p>The transmission electron microscope is used to acquire structural information of macromolecular complexes. However, as any other imaging device, it introduces optical aberrations that must be corrected if high-resolution structural information is to be obtained. The set of all aberrations are usually modeled in Fourier space by the so-called Contrast Transfer Function (CTF). Before correcting for the CTF, we must first estimate it from the electron micrographs. This is usually done by estimating a number of parameters specifying a theoretical model of the CTF. This estimation is performed by minimizing some error measure between the theoretical Power Spectrum Density (PSD) and the experimentally observed PSD. The high noise present in the micrographs, the possible local minima of the error function for estimating the CTF parameters, and the cross-talking between CTF parameters may cause errors in the estimated CTF parameters.</p> <p>Results</p> <p>In this paper, we explore the effect of these estimation errors on the theoretical CTF. For the CTF model proposed in <abbrgrp><abbr bid="B1">1</abbr></abbrgrp> we show which are the most sensitive CTF parameters as well as the most sensitive background parameters. Moreover, we provide a methodology to reveal the internal structure of the CTF model (which parameters influence in which parameters) and to estimate the accuracy of each model parameter. Finally, we explore the effect of the variability in the detection of the CTF for CTF phase and amplitude correction.</p> <p>Conclusion</p> <p>We show that the estimation errors for the CTF detection methodology proposed in <abbrgrp><abbr bid="B1">1</abbr></abbrgrp> does not show a significant deterioration of the CTF correction capabilities of subsequent algorithms. All together, the methodology described in this paper constitutes a powerful tool for the quantitative analysis of CTF models that can be applied to other models different from the one analyzed here.</p

Transfer function restoration in 3D electron microscopy via iterative data refinement

Three-dimensional electron microscopy (3D-EM) is a powerful tool for visualizing complex biological systems. As with any other imaging device, the electron microscope introduces a transfer function (called in this field the contrast transfer function, CTF) into the image acquisition process that modulates the various frequencies of the signal. Thus, the 3D reconstructions performed with these CTF-affected projections are also affected by an implicit 3D transfer function. For high-resolution electron microscopy, the effect of the CTF is quite dramatic and limits severely the achievable resolution. In this work we make use of the iterative data refinement (IDR) technique to ameliorate the effect of the CTF. It is demonstrated that the approach can be successfully applied to noisy data.Partial support is acknowledged to the Comisión Interministerial de Ciencia y Tecnología of Spain through projects BIO98-0761 and BIO2001-1237 and to National Institutes of Health through grant HL70472. The work of Y. Censor was done in part at the Center for Computational Mathematics and Scientific Computation (CCMSC) at the University of Haifa and supported by Research Grant 592/00 from the Israel Science Foundation founded by the Israel Academy of Sciences and Humanities

Consistent and elastic registration of histological sections using vector-spline regularization

The final publication is available at Springer via http://dx.doi.org/10.1007/11889762_8Revised Papers on Second International ECCV Workshop, CVAMIA 2006 Graz, Austria, May 12, 2006Here we present a new image registration algorithm for the alignment of histological sections that combines the ideas of B-spline based elastic registration and consistent image registration, to allow simultaneous registration of images in two directions (direct and inverse). In principle, deformations based on B-splines are not invertible. The consistency term overcomes this limitation and allows registration of two images in a completely symmetric way. This extension of the elastic registration method simplifies the search for the optimum deformation and allows registering with no information about landmarks or deformation regularization. This approach can also be used as the first step to solve the problem of group-wise registration.Ignacio Arganda-Carreras is being supported by a predoctoral FPI-CAM fellow- ship since October 2003. Carlos Ortiz-de-Solorzano is supported by a Ramon y Cajal (Spanish Ministry of Education and Science ryc-2004-002353) and a Marie Curie International Reintegration Grant (FP6-518688). Jan Kybic was sponsored by the Czech Ministery of Education under project number MSM210000012. Par- tial support is acknowledged to Comunidad de Madrid through grant GR/SAL/0234, to Instituto de Salud Carlos III-Fondo de Investigaciones Sanitarias (FIS) through the IM3 Network and grant 040683 and to the Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica (I+D+I)

On the development of three new tools for organizing and sharing information in three-dimensional electron microscopy

This work was funded by the Spanish Ministerio de Economía y Competividad through grants BFU2009-09331, BIO2010-16566, ACI2009-1022, ACI2010-1088 and AIC-A- 2011-0638, by the Comunidad Autonoma de Madrid through grant S2010/BMD-2305, by NFS grant No. 1114901 and by the Spanish National Institute of Bioinformatics (a project funded by the Instituto de Salud Carlos III). This work was conducted using the Protégé resource, which is supported by grant LM007885 from the United States National Library of Medicine. COSS is a Ramón y Cajal researcher financed by the European Social Fund and the Ministerio de Economía y Competitividad. JV is a Juan de la Cierva Postdoctoral Fellow (JCI-2011-10185). This work was funded by Instruct, which is part of the European Strategy Forum on Research Infrastructures (ESFRI) and is supported by national member subscriptions

Involvement of the Cellular Phosphatase DUSP1 in Vaccinia Virus Infection

Poxviruses encode a large variety of proteins that mimic, block or enhance host cell signaling pathways on their own benefit. It has been reported that mitogen-activated protein kinases (MAPKs) are specifically upregulated during vaccinia virus (VACV) infection. Here, we have evaluated the role of the MAPK negative regulator dual specificity phosphatase 1 (DUSP1) in the infection of VACV. We demonstrated that DUSP1 expression is enhanced upon infection with the replicative WR virus and with the attenuated VACV viruses MVA and NYVAC. This upregulation is dependent on early viral gene expression. In the absence of DUSP1 in cultured cells, there is an increased activation of its molecular targets JNK and ERK and an enhanced WR replication. Moreover, DUSP1 knock-out (KO) mice are more susceptible to WR infection as a result of enhanced virus replication in the lungs. Significantly, MVA, which is known to produce non-permissive infections in most mammalian cell lines, is able to grow in DUSP1 KO immortalized murine embryo fibroblasts (MEFs). By confocal and electron microscopy assays, we showed that in the absence of DUSP1 MVA morphogenesis is similar as in permissive cell lines and demonstrated that DUSP1 is involved at the stage of transition between IVN and MV in VACV morphogenesis. In addition, we have observed that the secretion of pro-inflammatory cytokines at early times post-infection in KO mice infected with MVA and NYVAC is increased and that the adaptive immune response is enhanced in comparison with WT-infected mice. Altogether, these findings reveal that DUSP1 is involved in the replication and host range of VACV and in the regulation of host immune responses through the modulation of MAPKs. Thus, in this study we demonstrate that DUSP1 is actively involved in the antiviral host defense mechanism against a poxvirus infection

FSC-Q: a CryoEM map-to-atomic model quality validation based on the local Fourier shell correlation

In recent years, advances in cryoEM have dramatically increased the resolution of reconstructions and, with it, the number of solved atomic models. It is widely accepted that the quality of cryoEM maps varies locally; therefore, the evaluation of the maps-derived structural models must be done locally as well. In this article, a method for the local analysis of the map-to-model fit is presented. The algorithm uses a comparison of two local resolution maps. The first is the local FSC (Fourier shell correlation) between the full map and the model, while the second is calculated between the half maps normally used in typical single particle analysis workflows. We call the quality measure “FSC-Q”, and it is a quantitative estimation of how much of the model is supported by the signal content of the map. Furthermore, we show that FSC-Q may be helpful to detect overfitting. It can be used to complement other methods, such as the Q-score method that estimates the resolvability of atomsWe thank Prof. David Veesler for providing us the half maps of the spike glycoprotein of SARS-CoV-2. The authors would like to acknowledge financial support from: the Comunidad de Madrid through grant CAM (S2017/BMD-3817), the Spanish National Research Council (PIE/COVID-19 number 202020E079), the Spanish Ministry of Economy and Competitiveness through grants SEV 2017-0712, PID2019-104757RB-I00/AEI/10.13039/501100011033, the Instituto de Salud Carlos III through grant PT17/0009/0010 (ISCIII-GEFI/ERDF-). Instruct-ULTRA (Grant 731005), an EU H2020 project to further develop the services of Instruct-ERIC. UE H2020 grant HighResCells (ERC-2018-SyG, Proposal: 810057). This work was supported by the Intramural Research Program of the National Institute for Arthritis, musculoskeletal, and Skin Diseases, NIH. The authors acknowledge the support and the use of resources of Instruct, a Landmark ESFRI projec