Lipid Trafficking sans Vesicles: Where, Why, How?

Eukaryotic cells possess a remarkable diversity of lipids, which distribute among cellular membranes by well-characterized vesicle trafficking pathways. However, transport of lipids by alternate, or “nonvesicular,” routes is also critical for lipid synthesis, metabolism, and proper membrane partitioning. In the past few years, considerable progress has been made in characterizing the mechanisms of nonvesicular lipid transport and how it may go awry in particular diseases, but many fundamental questions remain for this rising field

Ltc1 is an ER-localized sterol transporter and a component of ER-mitochondria and ER-vacuole contacts.

Organelle contact sites perform fundamental functions in cells, including lipid and ion homeostasis, membrane dynamics, and signaling. Using a forward proteomics approach in yeast, we identified new ER-mitochondria and ER-vacuole contacts specified by an uncharacterized protein, Ylr072w. Ylr072w is a conserved protein with GRAM and VASt domains that selectively transports sterols and is thus termed Ltc1, for Lipid transfer at contact site 1. Ltc1 localized to ER-mitochondria and ER-vacuole contacts via the mitochondrial import receptors Tom70/71 and the vacuolar protein Vac8, respectively. At mitochondria, Ltc1 was required for cell viability in the absence of Mdm34, a subunit of the ER-mitochondria encounter structure. At vacuoles, Ltc1 was required for sterol-enriched membrane domain formation in response to stress. Increasing the proportion of Ltc1 at vacuoles was sufficient to induce sterol-enriched vacuolar domains without stress. Thus, our data support a model in which Ltc1 is a sterol-dependent regulator of organelle and cellular homeostasis via its dual localization to ER-mitochondria and ER-vacuole contact sites

Sterol transporters at membrane contact sites regulate TORC1 and TORC2 signaling.

Membrane contact sites (MCSs) function to facilitate the formation of membrane domains composed of specialized lipids, proteins, and nucleic acids. In cells, membrane domains regulate membrane dynamics and biochemical and signaling pathways. We and others identified a highly conserved family of sterol transport proteins (Ltc/Lam) localized at diverse MCSs. In this study, we describe data indicating that the yeast family members Ltc1 and Ltc3/4 function at the vacuole and plasma membrane, respectively, to create membrane domains that partition upstream regulators of the TORC1 and TORC2 signaling pathways to coordinate cellular stress responses with sterol homeostasis

Mutants Affecting the Structure of the Cortical Endoplasmic Reticulum in Saccharomyces cerevisiae

We find that the peripheral ER in Saccharomyces cerevisiae forms a dynamic network of interconnecting membrane tubules throughout the cell cycle, similar to the ER in higher eukaryotes. Maintenance of this network does not require microtubule or actin filaments, but its dynamic behavior is largely dependent on the actin cytoskeleton. We isolated three conditional mutants that disrupt peripheral ER structure. One has a mutation in a component of the COPI coat complex, which is required for vesicle budding. This mutant has a partial defect in ER segregation into daughter cells and disorganized ER in mother cells. A similar phenotype was found in other mutants with defects in vesicular trafficking between ER and Golgi complex, but not in mutants blocked at later steps in the secretory pathway. The other two mutants found in the screen have defects in the signal recognition particle (SRP) receptor. This receptor, along with SRP, targets ribosome–nascent chain complexes to the ER membrane for protein translocation. A conditional mutation in SRP also disrupts ER structure, but other mutants with translocation defects do not. We also demonstrate that, both in wild-type and mutant cells, the ER and mitochondria partially coalign, and that mutations that disrupt ER structure also affect mitochondrial structure. Our data suggest that both trafficking between the ER and Golgi complex and ribosome targeting are important for maintaining ER structure, and that proper ER structure may be required to maintain mitochondrial structure

The dynamin-like GTPase Sey1p mediates homotypic ER fusion in S. cerevisiae

The endoplasmic reticulum (ER) forms a network of tubules and sheets that requires homotypic membrane fusion to be maintained. In metazoans, this process is mediated by dynamin-like guanosine triphosphatases (GTPases) called atlastins (ATLs), which are also required to maintain ER morphology. Previous work suggested that the dynamin-like GTPase Sey1p was needed to maintain ER morphology in Saccharomyces cerevisiae. In this paper, we demonstrate that Sey1p, like ATLs, mediates homotypic ER fusion. The absence of Sey1p resulted in the ER undergoing delayed fusion in vivo and proteoliposomes containing purified Sey1p fused in a GTP-dependent manner in vitro. Sey1p could be partially replaced by ATL1 in vivo. Like ATL1, Sey1p underwent GTP-dependent dimerization. We found that the residual ER–ER fusion that occurred in cells lacking Sey1p required the ER SNARE Ufe1p. Collectively, our results show that Sey1p and its homologues function analogously to ATLs in mediating ER fusion. They also indicate that S. cerevisiae has an alternative fusion mechanism that requires ER SNAREs

Формування законодавчої і нормативної бази архівної справи в республіці Казахстан

Аналізується законодавство Республіки Казахстан, спрямоване на забезпечення збереження документальної спадщини, регулювання діяльності архівної системи в умовах державного суверенітету. Визначаються особливості архівного законодавства кінця 1990-х рр., прослідковуються його зміни та напрями удосконалення в 2000-х рр., процеси формування нормативної бази архівної справи.Анализируется законодательство Республики Казахстан, направленное на обеспечение сохранности документального наследия, регулирование деятельности архивной системы в условиях государственного суверенитета. Определяются особенности архивного законодательства конца 1990-х гг., прослеживаются его изменения и направления усовершенствования в 2000- х гг., процессы формирования нормативной базы архивного дела.The legislation of Republic of Kazakhstan, directed on providing of safety of documentary legacy, adjusting of activity of the archival system in the conditions of the state sovereignty is analysed in the article. The features of the archival legislation the end of 1990-th are determined, its changes and directions of improvement in the 2000-th, the processes of forming of normative base of the archival affairs are traced

Architecture of lipid droplets in endoplasmic reticulum is determined by phospholipid intrinsic curvature

Lipid droplets (LDs) store fats and play critical roles in lipid and energy homeostasis. They form between the leaflets of the endoplasmic reticulum (ER) membrane and consist of a neutral lipid core wrapped in a phospholipid monolayer with proteins. Two types of ER-LD architecture are thought to exist and be essential for LD functioning. Maturing LDs either emerge from the ER into the cytoplasm, remaining attached to the ER by a narrow membrane neck, or stay embedded in the ER and are surrounded by ER membrane. Here, we identify a lipid-based mechanism that controls which of these two architectures is favored. Theoretical modeling indicated that the intrinsic molecular curvatures of ER phospholipids can determine whether LDs remain embedded in or emerge from the ER; lipids with negative intrinsic curvature such as diacylglycerol (DAG) and phosphatidylethanolamine favor LD embedding, while those with positive intrinsic curvature, like lysolipids, support LD emergence. This prediction was verified by altering the lipid composition of the ER in S. cerevisiae using mutants and the addition of exogenous lipids. We found that fat-storage-inducing transmembrane protein 2 (FIT2) homologs become enriched at sites of LD generation when biogenesis is induced. DAG accumulates at sites of LD biogenesis, and FIT2 proteins may promote LD emergence from the ER by reducing DAG levels at these sites. Altogether, our findings suggest that cells regulate LD integration in the ER by modulating ER lipid composition, particularly at sites of LD biogenesis and that FIT2 proteins may play a central role in this process