Surface tension-driven sorting of human perilipins on lipid droplets.

Perilipins (PLINs), the most abundant proteins on lipid droplets (LDs), display similar domain organization including amphipathic helices (AH). However, the five human PLINs bind different LDs, suggesting different modes of interaction. We established a minimal system whereby artificial LDs covered with defined polar lipids were transiently deformed to promote surface tension.

Established and emerging players in phospholipid scrambling: A structural perspective.

The maintenance of a diverse and non-homogeneous lipid composition in cell membranes is crucial for a multitude of cellular processes. One important example is transbilayer lipid asymmetry, which refers to a difference in lipid composition between the two leaflets of a cellular membrane. Transbilayer asymmetry is especially pronounced at the plasma membrane, where at resting state, negatively-charged phospholipids such as phosphatidylserine (PS) are almost exclusively restricted to the cytosolic leaflet, whereas sphingolipids are mostly found in the exoplasmic leaflet.

Molecular mechanisms of perilipin protein function in lipid droplet metabolism.

Perilipins are abundant lipid droplet (LD) proteins present in all metazoans and also in Amoebozoa and fungi. Humans express five perilipins, which share a similar domain organization: an amino-terminal PAT domain and an 11-mer repeat region, which can fold into amphipathic helices that interact with LDs, followed by a structured carboxy-terminal domain. Variations of this organization that arose during vertebrate evolution allow for functional specialization between perilipins in relation to the metabolic needs of different tissues.

Transport Pathways That Contribute to the Cellular Distribution of Phosphatidylserine.

Phosphatidylserine (PS) is a negatively charged phospholipid that displays a highly uneven distribution within cellular membranes, essential for establishment of cell polarity and other processes. In this review, we discuss how combined action of PS biosynthesis enzymes in the endoplasmic reticulum (ER), lipid transfer proteins (LTPs) acting within membrane contact sites (MCS) between the ER and other compartments, and lipid flippases and scramblases that mediate PS flip-flop between membrane leaflets controls the cellular distribution of PS. Enrichment of PS in specific compartments, in particular in the cytosolic leaflet of the plasma membrane (PM), requires input of energy, which can be supplied in the form of ATP or by phosphoinositides.

Exceptional stability of a perilipin on lipid droplets depends on its polar residues, suggesting multimeric assembly.

Numerous proteins target lipid droplets (LDs) through amphipathic helices (AHs). It is generally assumed that AHs insert bulky hydrophobic residues in packing defects at the LD surface. However, this model does not explain the targeting of perilipins, the most abundant and specific amphipathic proteins of LDs, which are weakly hydrophobic.

A comprehensive library of fluorescent constructs of SARS-CoV-2 proteins and their initial characterisation in different cell types.

Background Information: Comprehensive libraries of plasmids for SARS-CoV-2 proteins with various tags (e.g., Strep, HA, Turbo) are now available.

Osh6 requires Ist2 for localization to ER-PM contacts and efficient phosphatidylserine transport in budding yeast.

Osh6 and Osh7 are lipid transfer proteins (LTPs) that move phosphatidylserine (PS) from the endoplasmic reticulum (ER) to the plasma membrane (PM). High PS levels at the PM are key for many cellular functions. Intriguingly, Osh6 and Osh7 localize to ER-PM contact sites, although they lack membrane-targeting motifs, in contrast to multidomain LTPs that both bridge membranes and convey lipids.

Following Anterograde Transport of Phosphatidylserine in Yeast in Real Time.

In order to understand how lipids are sorted between cellular compartments, kinetic assays are required to selectively follow the transport of lipid species in cells. We present here a microfluidics-based protocol to follow the transport of phosphatidylserine (PS) in yeast cells from the site of its synthesis, the endoplasmic reticulum (ER), to downstream compartments, primarily the plasma membrane under our conditions. This assay takes advantage of yeast cells lacking Cho1, the enzyme responsible for PS synthesis.

The Many Faces of Amphipathic Helices.

Amphipathic helices (AHs), a secondary feature found in many proteins, are defined by their structure and by the segregation of hydrophobic and polar residues between two faces of the helix. This segregation allows AHs to adsorb at polar⁻apolar interfaces such as the lipid surfaces of cellular organelles. Using various examples, we discuss here how variations within this general scheme impart membrane-interacting AHs with different interfacial properties.

A Combinatorial Lipid Code Shapes the Electrostatic Landscape of Plant Endomembranes.

Membrane surface charge is critical for the transient, yet specific recruitment of proteins with polybasic regions to certain organelles. In eukaryotes, the plasma membrane (PM) is the most electronegative compartment of the cell, which specifies its identity. As such, membrane electrostatics is a central parameter in signaling, intracellular trafficking, and polarity.

A giant amphipathic helix from a perilipin that is adapted for coating lipid droplets.

How proteins are targeted to lipid droplets (LDs) and distinguish the LD surface from the surfaces of other organelles is poorly understood, but many contain predicted amphipathic helices (AHs) that are involved in targeting. We have focused on human perilipin 4 (Plin4), which contains an AH that is exceptional in terms of length and repetitiveness. Using model cellular systems, we show that AH length, hydrophobicity, and charge are important for AH targeting to LDs and that these properties can compensate for one another, albeit at a loss of targeting specificity.

Crowd-Sourcing of Membrane Fission: How crowding of non-specialized membrane-bound proteins contributes to cellular membrane fission.

Fission of cellular membranes is ubiquitous and essential for life. Complex protein machineries, such as the dynamin and ESCRT spirals, have evolved to mediate membrane fission during diverse cellular processes, for example, vesicle budding. A new study suggests that non-specialized membrane-bound proteins can induce membrane fission through mass action due to protein crowding.

Advances on the Transfer of Lipids by Lipid Transfer Proteins.

Transfer of lipid across the cytoplasm is an essential process for intracellular lipid traffic. Lipid transfer proteins (LTPs) are defined by highly controlled in vitro experiments. The functional relevance of these is supported by evidence for the same reactions inside cells.

New molecular mechanisms of inter-organelle lipid transport.

Lipids are precisely distributed in cell membranes, along with associated proteins defining organelle identity. Because the major cellular lipid factory is the endoplasmic reticulum (ER), a key issue is to understand how various lipids are subsequently delivered to other compartments by vesicular and non-vesicular transport pathways. Efforts are currently made to decipher how lipid transfer proteins (LTPs) work either across long distances or confined to membrane contact sites (MCSs) where two organelles are at close proximity.

Membrane bending by protein crowding is affected by protein lateral confinement.

Crowding of asymmetrically-distributed membrane proteins has been recently recognized as an important factor in remodeling of biological membranes, for example during transport vesicle formation. In this paper, we theoretically analyze the effect of protein crowding on membrane bending and examine its dependence on protein size, shape, transmembrane asymmetry and lateral confinement. We consider three scenarios of protein lateral organization, which are highly relevant for cellular membranes in general: freely diffusing membrane proteins without lateral confinement, the presence of a diffusion barrier and interactions with a vesicular coat.

INTRACELLULAR TRANSPORT. Phosphatidylserine transport by ORP/Osh proteins is driven by phosphatidylinositol 4-phosphate.

In eukaryotic cells, phosphatidylserine (PS) is synthesized in the endoplasmic reticulum (ER) but is highly enriched in the plasma membrane (PM), where it contributes negative charge and to specific recruitment of signaling proteins. This distribution relies on transport mechanisms whose nature remains elusive. Here, we found that the PS transporter Osh6p extracted phosphatidylinositol 4-phosphate (PI4P) and exchanged PS for PI4P between two membranes.

Traffic of p24 Proteins and COPII Coat Composition Mutually Influence Membrane Scaffolding.

Eukaryotic protein secretion requires efficient and accurate delivery of diverse secretory and membrane proteins. This process initiates in the ER, where vesicles are sculpted by the essential COPII coat. The Sec13p subunit of the COPII coat contributes to membrane scaffolding, which enforces curvature on the nascent vesicle.

Fatty acid metabolism meets organelle dynamics.

Upon nutrient deprivation, cells metabolize fatty acids (FAs) in mitochondria to supply energy, but how FAs, stored as triacylglycerols in lipid droplets, reach mitochondria has been mysterious. Rambold et al. (2015) now show that FA mobilization depends on triacylglycerol lipolysis, whereas autophagy feeds the lipid droplet pool for continued fueling of mitochondria.

Membrane bending: the power of protein imbalance.

Many cellular processes require membrane deformation, which is driven by specialized protein machinery and can often be recapitulated using pure lipid bilayers. However, biological membranes contain a large amount of embedded proteins. Recent research suggests that membrane-bound proteins with asymmetric distribution of mass across the bilayer can influence membrane bending in a nonspecific manner due to molecular crowding.

ER cargo properties specify a requirement for COPII coat rigidity mediated by Sec13p.

Eukaryotic secretory proteins exit the endoplasmic reticulum (ER) via transport vesicles generated by the essential coat protein complex II (COPII) proteins. The outer coat complex, Sec13-Sec31, forms a scaffold that is thought to enforce curvature. By exploiting yeast bypass-of-sec-thirteen (bst) mutants, where Sec13p is dispensable, we probed the relationship between a compromised COPII coat and the cellular context in which it could still function.

Ent3p and Ent5p exhibit cargo-specific functions in trafficking proteins between the trans-Golgi network and the endosomes in yeast.

The phosphoinositide-binding proteins Ent3p and Ent5p are required for protein transport from the trans-Golgi network (TGN) to the vacuole in Saccharomyces cerevisiae. Both proteins interact with the monomeric clathrin adaptor Gga2p, but Ent5p also interacts with the clathrin adaptor protein 1 (AP-1) complex, which facilitates retention of proteins such as Chs3p at the TGN. When both ENT3 and ENT5 are mutated, Chs3p is diverted from an intracellular reservoir to the cell surface.