Liquid-liquid phase separation is ubiquitous in suspensions of nanoparticles, proteins and colloids. It plays an important role in gel formation, protein crystallization and perhaps even as an organizing principle in cellular biology 1,2 .With a few notable exceptions 3,4 , surface-tension-minimizing liquid droplets in bulk suspensions continuously coalesce, increasing in size without bound until achieving macroscale phase separation. In comparison, the phase behavior of colloids, nanoparticles or proteins confined to interfaces, surfaces or membranes is significantly more complex 5-11 . Inclusions distort the local interface structure leading to interactions that are fundamentally different from the well-studied interactions mediated by isotropic solvents 12,13 . Here, we investigate liquid-liquid phase separation in monolayer membranes composed of dissimilar chiral colloidal rods. We demonstrate that colloidal rafts are a ubiquitous feature of binary colloidal membranes. We measure the raft free energy landscape by visualizing its assembly kinetics. Subsequently, we quantify repulsive raft-raft interactions and relate them to directly imaged raft-induced membrane distortions, demonstrating that particle chirality plays a key role in this microphase separation. At high densities, rafts assemble into cluster crystals which constantly exchange monomeric rods with the background reservoir to maintain a self-limited size. Lastly, we 2 demonstrate that rafts can form bonds to assemble into higher-order suprastructures. Our work demonstrates that membrane-mediated liquid-liquid phase separation can be fundamentally different from the well-characterized behavior of bulk liquids. It outlines a robust membrane-based pathway for assembly of monodisperse liquid clusters which is complementary to existing methods which take place in bulk suspensions [14][15][16] . Finally, it reveals that chiral inclusions in membranes acquire long-ranged repulsive interactions, which might play a role in stabilizing assemblages of finite size 11,17 .In the presence of non-adsorbing polymer, mono-disperse rod-like viruses experience effective depletion attractions that drive their lateral association. These interactions can lead to assembly of one-rod-length-thick colloidal monolayer membranes that are held together by the osmotic pressure of the enveloping polymer suspension 18 . We allow membranes to sediment to the bottom of the sample chambers in which case the constituent rods point in the z direction, while all images are taken in the x-y plane.Although they differ on molecular scales, the long-wavelength fluctuations of colloidal monolayers and lipid bilayers are described by the same free energy. In this work, we investigated the behavior of colloidal membranes containing a mixture of two rods: 880 nm long rod-like fd-Y21M virus and 1200 nm long M13KO7 virus 19 . Membranes were prepared by adding a depletant to a dilute isotropic fd-Y21M/M13KO7 mixture. For all parameters investigated, both rods co-assembled into binary membranes. ...

Mycobacterium tuberculosis (M.tb), which requires iron for survival, acquires this element by synthesizing iron-binding molecules known as siderophores and by recruiting a host iron-transport protein, transferrin, to the phagosome. The siderophores extract iron from transferrin and transport it into the bacterium. Here we describe an additional mechanism for iron acquisition, consisting of an M.tb protein that drives transport of human holo-transferrin into M.tb cells. The pathogenic strain M.tb H37Rv expresses several proteins that can bind human holo-transferrin. One of these proteins is the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH, Rv1436), which is present on the surface of M.tb and its relative Mycobacterium smegmatis. Overexpression of GAPDH results in increased transferrin binding to M.tb cells and iron uptake. Human transferrin is internalized across the mycobacterial cell wall in a GAPDH-dependent manner within infected macrophages.

Recycling endosomes (REs) are transient endosomal tubular intermediates of early/sorting endosomes (E/SEs) that function in cargo recycling to the cell surface and deliver the cell type‐specific cargo to lysosome‐related organelles such as melanosomes in melanocytes. However, the mechanism of RE biogenesis is largely unknown. In this study, by using an endosomal Rab‐specific RNAi screen, we identified Rab22A as a critical player during RE biogenesis. Rab22A‐knockdown results in reduced RE dynamics and concurrent cargo accumulation in the E/SEs or lysosomes. Rab22A forms a complex with BLOC‐1, BLOC‐2 and the kinesin‐3 family motor KIF13A on endosomes. Consistently, the RE‐dependent transport defects observed in Rab22A‐depleted cells phenocopy those in BLOC‐1‐/BLOC‐2‐deficient cells. Further, Rab22A depletion reduced the membrane association of BLOC‐1/BLOC‐2. Taken together, these findings suggest that Rab22A promotes the assembly of a BLOC‐1‐BLOC‐2‐KIF13A complex on E/SEs to generate REs that maintain cellular and organelle homeostasis.

In the presence of a nonadsorbing polymer, monodisperse rod-like particles assemble into colloidal membranes, which are one-rodlength-thick liquid-like monolayers of aligned rods. Unlike 3D edgeless bilayer vesicles, colloidal monolayer membranes form open structures with an exposed edge, thus presenting an opportunity to study elasticity of fluid sheets. Membranes assembled from single-component chiral rods form flat disks with uniform edge twist. In comparison, membranes composed of a mixture of rods with opposite chiralities can have the edge twist of either handedness. In this limit, disk-shaped membranes become unstable, instead forming structures with scalloped edges, where two adjacent lobes with opposite handedness are separated by a cuspshaped point defect. Such membranes adopt a 3D configuration, with cusp defects alternatively located above and below the membrane plane. In the achiral regime, the cusp defects have repulsive interactions, but away from this limit we measure effective longranged attractive binding. A phenomenological model shows that the increase in the edge energy of scalloped membranes is compensated by concomitant decrease in the deformation energy due to Gaussian curvature associated with scalloped edges, demonstrating that colloidal membranes have positive Gaussian modulus. A simple excluded volume argument predicts the sign and magnitude of the Gaussian curvature modulus that is in agreement with experimental measurements. Our results provide insight into how the interplay between membrane elasticity, geometrical frustration, and achiral symmetry breaking can be used to fold colloidal membranes into 3D shapes.self-assembly | membranes | liquid crystals | Gaussian curvature | chirality T he possible configurations and shapes of 2D fluid membranes can be described by a continuum energy expression that accounts for the membrane's out-of-plane deformations as well as the line tension associated with the membrane's exposed edge (1, 2). Because an arbitrary deformation of a thin layer can have either mean and/or Gaussian curvature, the full theoretical description of membranes, in principle, requires two parameters, the bending and Gaussian curvature moduli. However, lipid bilayers almost always appear as edgeless 3D vesicles, which further simplify theoretical modeling. In particular, integrating Gaussian curvature over any simply closed surface yields a constant (3). Thus, the shape fluctuations of a closed vesicle only depend on the membrane-bending modulus. Consequently, experiments that interrogated mechanics or shape fluctuations of vesicles provided extensive information about the membrane curvature modulus and how it depends on the structure of the constituent particles (4-6). In comparison, significantly less is known about the Gaussian modulus, despite the significant role it plays in fundamental biological and technological processes such as pore formation as well as vesicle fusion and fission (7)(8)(9)(10)(11).Recent experiments have demonstrated that, in the presence of a d...