Designer amphiphilic proteins as building blocks for the intracellular formation of organelle-like compartments.

Nanoscale biological materials formed by the assembly of defined block-domain proteins control the formation of cellular compartments such as organelles. Here, we introduce an approach to intentionally 'program' the de novo synthesis and self-assembly of genetically encoded amphiphilic proteins to form cellular compartments, or organelles, in Escherichia coli. These proteins serve as building blocks for the formation of artificial compartments in vivo in a similar way to lipid-based organelles. We investigated the formation of these organelles using epifluorescence microscopy, total internal reflection fluorescence microscopy and transmission electron microscopy. The in vivo modification of these protein-based de novo organelles, by means of site-specific incorporation of unnatural amino acids, allows the introduction of artificial chemical functionalities. Co-localization of membrane proteins results in the formation of functionalized artificial organelles combining artificial and natural cellular function. Adding these protein structures to the cellular machinery may have consequences in nanobiotechnology, synthetic biology and materials science, including the constitution of artificial cells and bio-based metamaterials.

Coherent total internal reflection dark-field microscopy: label-free imaging beyond the diffraction limit.

Coherent imaging is barely applicable in life-science microscopy due to multiple interference artifacts. Here, we show how these interferences can be used to improve image resolution and contrast. We present a dark-field microscopy technique with evanescent illumination via total internal reflection that delivers high-contrast images of coherently scattering samples. By incoherent averaging of multiple coherent images illuminated from different directions we can resolve image structures that remain unresolved by conventional (incoherent) fluorescence microscopy. We provide images of 190 nm beads revealing resolution beyond the diffraction limit and slightly increased object distances. An analytical model is introduced that accounts for the observed effects and which is confirmed by numerical simulations. Our approach may be a route to fast, label-free, super-resolution imaging in live-cell microscopy.

Gait Analysis of Patients After Allogeneic Hematopoietic Cell Transplantation Reveals Impairments of Functional Performance.

Background: After allogeneic hematopoietic cell transplantation (alloHCT), patients often report functional impairments like reduced gait speed and muscle weakness. These impairments can increase the risk of adverse health events similar to elderly populations. However, they have not been quantified in patients after alloHCT (PATs). Methods: We compared fear of falling (Falls Efficacy Scale-International) and temporal gait parameters recorded on a 10-m walkway at preferred and maximum gait speed and under dual-task walking of 16 PATs (aged 31-73 years) with 15 age-matched control participants (CONs) and 17 seniors (SENs, aged >73 years). Results: Groups' gait parameters especially differed during the maximum speed condition: PATs walked slower and required more steps/10 m than CONs. PATs exhibited greater stride, stance, and swing times than CONs. PATs' swing time was even longer than SENs'. The PATs' ability to accelerate their gait speed from preferred to fast was smaller compared with CONs'. PATs reported a greater fear of falling than CONs and SENs. Conclusion: Gait analysis of alloHCT patients has revealed impairments of functional performance. Patients presented a diminished ability to accelerate gait and extending steps possibly related to a notable strength deficit that impairs power-generation abilities from lower extremities. Furthermore, patients reported a greater fear of falling than control participants and even seniors. Slowing locomotion could be a risk-preventive safety strategy. Since functional disadvantages may put alloHCT patients at a higher risk of frailty, reinforcing appropriate physical exercises already during and after alloHCT could prevent adverse health events and reduce the risk of premature functional aging.

Patchwork organization of the yeast plasma membrane into numerous coexisting domains.

The plasma membrane is made up of lipids and proteins, and serves as an active interface between the cell and its environment. Many plasma-membrane proteins are laterally segregated in the plane of the membrane, but the underlying mechanisms remain controversial. Here we investigate the distribution and dynamics of a representative set of plasma-membrane-associated proteins in yeast cells. These proteins were distributed non-homogeneously in patterns ranging from distinct patches to nearly continuous networks, and these patterns were in turn strongly influenced by the lipid composition of the plasma membrane. Most proteins segregated into distinct domains. However, proteins with similar or identical transmembrane sequences (TMSs) showed a marked tendency to co-localize. Indeed we could predictably relocate proteins by swapping their TMSs. Finally, we found that the domain association of plasma-membrane proteins has an impact on their function. Our results are consistent with self-organization of biological membranes into a patchwork of coexisting domains.