Cell News 01/2017
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In situ cryo-electron tomography reveals the molecular
sociology at the HeLa cell nuclear periphery
Julia Mahamid
ABSTRACTS CHAIRS AND SPEAKERS
Department of Molecular Structural Biology,
Max-Planck Institute of Biochemistry, Martinsried
Most structural biology focuses on the structure and function
of individual proteins or complexes. However, cellular functions
arise from an intricate network of macromolecular interac-
tions. Hence it is of fundamental importance to be able to
decipher the ‘molecular sociology’ of cells, ideally by direct vi-
sualization – the goal of visual proteomics. Traditional electron
microscopy has established cellular ultrastructure as we know
it, but falls short of revealing the finer molecular detail. Here,
cryo-electron tomography provides the unique opportunity for
imaging cellular organization at near-native conditions and at
molecular resolution. Identification and assignment of molec-
ular structures directly from three-dimensional stills was so
far hampered by the low-contrast and the low signal-to-noise
ratio of the tomograms. To image the dense, crowded interior
of mammalian cells was further complicated by the ‘immense’
sample thickness (immense as seen from a nanoscopic per-
spective). Thus intracellular structures, especially those in close
proximity to the eukaryotic nucleus remained largely unex-
plored.
The combination of recent developments in the field of
cryo-electron microscopy, namely cryo-focused ion beam
micromachining, direct electron detectors and the volta phase
plate made it possible to unambiguously identify molecular
components in situ based simply and solely on their visible fea-
tures and structural signatures. Averaging and classification of
80S ribosomes depicted the native structure and the supramo-
lecular organization of the cytoplasmic translation machinery
in unperturbed human cells, while analysis of large dynamic
structures such as the nuclear pore complex revealed structural
variations detectable on the level of individual complexes. This
technical advance rendered previously elusive structures, such
as nucleosome chains and the nuclear lamina, directly visible.
Depiction of the lamina structure provides new insight into its
contribution to metazoan nuclear stiffness.
Signalling myoblast fusion in Drosophila
Andreas Löwer, Stafanie Lübke, Carina Braukmann, Anja Rudolf and Susanne Filiz Önel
Philipps-Universität Marburg, FB17, Biologie,
Karl-von-Frisch Str. 8, 35043 Marburg
The formation of multinucleated skeletal muscle fibres depends
on the fusion of myoblasts, a process that requires multiple
morphological changes of the cells. As a first step, myoblasts
must migrate towards each other. Next, they recognize and ad-
here, so that close membrane proximity is achieved. This step
further involves the activation of a protein-machinery leading
to membrane instability and fusion pore formation.
In Drosophila, multinucleated muscles form via the fusion of
two different myoblast-types: founder cells (FC) that determine
the size, position and epidermal attachment of the muscle;
and naïve fusion-competent myoblasts (FCM). Cell adhesion
molecules of the Immunoglobulin superfamiliy (IgSF) mediate
the recognition and adhesion of both myoblast-types. They or-
chestrate a polarisation process in both myoblast-types, which
involves the translocation of intracellular proteins and vesicles
to the site of cell-cell contact, cytoskeleton rearrangements
and the dissolution of electron-dense plaques. This polarisation
process is very similar to what is observed in the immunologi-
cal synapse that forms between cells of the immune system.
To understand the signalling mechanism that controls myoblast
fusion during development, we have analyzed genetic loss-of-
function mutants on ultrastructural level and performed classic
epistasis experiments. Our data indicate that myoblast fusion
is a tightly controlled mechanism involving multiple layers of
coordinated Arp2/3-dependent F-actin polymerization to bring
plasma membranes in close apposition and to induce fusion
pore formation.
