Christian Renken

Structural and functional features and significance of the physical linkage between ER and mitochondria

The role of mitochondria in cell metabolism and survival is controlled by calcium signals that are commonly transmitted at the close associations between mitochondria and endoplasmic reticulum (ER). However, the physical linkage of the ER–mitochondria interface and its relevance for cell function remains elusive. We show by electron tomography that ER and mitochondria are adjoined by tethers that are ∼10 nm at the smooth ER and ∼25 nm at the rough ER. Limited proteolysis separates ER from mitochondria, whereas expression of a short “synthetic linker” (<5 nm) leads to tightening of the associations. Although normal connections are necessary and sufficient for proper propagation of ER-derived calcium signals to the mitochondria, tightened connections, synthetic or naturally observed under apoptosis-inducing conditions, make mitochondria prone to Ca2+ overloading and ensuing permeability transition. These results reveal an unexpected dependence of cell function and survival on the maintenance of proper spacing between the ER and mitochondria.

The bactericidal effect of a complement-independent antibody is osmolytic and specific to Borrelia

A complement-independent bactericidal IgG1 against the OspB of Borrelia burgdorferi increased the permeability of the outer membrane through the creation of openings of 2.8 – 4.4 nm, resulting in its osmotic lysis. Cryo-electron microscopy and tomography demonstrated that exposure to the antibody causes the formation of outer membrane projections and large breaks which may precede the increase in permeability of the outer membrane. The bactericidal effect of this antibody is not transferable to Escherichia coli expressing rOspB on its outer membrane. Additionally, the porin P66, the only protein that coprecipitated with OspB, is dispensable for the bactericidal mechanism.

Chapter 6 Principles and Practice in Electron Tomography

Publisher Summary In electron tomography, a three-dimensional (3D) image is reconstructed from 2D projection images, usually by back-projection methods. The same back-projection approach is used in medical imaging methods such as computerized axial tomography (CAT) scanning, but electron tomography entails special considerations arising from the nature of the specimens examined and their interactions with the electron beam. Electron tomography is an invaluable tool for exploring cellular architecture with sufficient resolution to characterize extended structures and identify large macromolecular assemblies within a cell. Problems created by incomplete angular coverage of the input data can be minimized by using a fine angular sampling interval and by collecting a dual-axis tilt series over the greatest possible tilt range. Cryo-electron tomography has made particularly impressive gains during the last five years, and its application to frozen-hydrated specimens is expected to grow in the near future, despite the low tolerance of frozen-hydrated specimens to electron exposure. The efficacy of electron tomography can be enhanced by prior knowledge of structural components located in the 3D reconstruction. In some cases, a priori knowledge is formally incorporated into motif searches or model-based segmentation, but more frequently it manifests as the observers ability to discern familiar landmarks within the reconstruction.