scholarly journals Multiple C2 domain-containing transmembrane proteins promote lipid droplet biogenesis and growth at specialized ER subdomains

2021 ◽  
pp. mbc.E20-09-0590
Author(s):  
Amit S. Joshi ◽  
Joey V. Ragusa ◽  
William A. Prinz ◽  
Sarah Cohen
Keyword(s):  
Endoplasmic Reticulum ◽  
Lipid Droplets ◽  
Transmembrane Domain ◽  
Neutral Lipids ◽  
Transmembrane Proteins ◽  
C2 Domain ◽  
General Role ◽  
C2 Domains ◽  
Contact Sites ◽  
Er Membrane

Lipid droplets (LDs) are neutral lipid-containing organelles enclosed in a single monolayer of phospholipids. LD formation begins with the accumulation of neutral lipids within the bilayer of the endoplasmic reticulum (ER) membrane. It is not known how the sites of formation of nascent LDs in the ER membrane are determined. Here we show that multiple C2 domain-containing transmembrane proteins, MCTP1 and MCTP2, are at sites of LD formation in specialized ER subdomains. We show that the transmembrane domain (TMD) of these proteins is similar to a reticulon homology domain. Like reticulons, these proteins tubulate the ER membrane and favor highly curved regions of the ER. Our data indicate that the MCTP TMDs promote LD biogenesis, increasing LD number. MCTPs co-localize with seipin, a protein involved in LD biogenesis, but form more stable microdomains in the ER. The MCTP C2 domains bind charged lipids and regulate LD size, likely by mediating ER-LD contact sites. Together, our data indicate that MCTPs form microdomains within ER tubules that regulate LD biogenesis, size, and ER-LD contacts. Interestingly, MCTP punctae colocalized with other organelles as well, suggesting that these proteins may play a more general role in linking tubular ER to organelle contact sites. [Media: see text] [Media: see text]

scholarly journals A Unique Junctional Interface at Contact Sites Between the Endoplasmic Reticulum and Lipid Droplets

2021 ◽  
Vol 9 ◽  
Author(s):  
Vineet Choudhary ◽  
Roger Schneiter
Keyword(s):  
Endoplasmic Reticulum ◽  
Lipid Droplets ◽  
Neutral Lipids ◽  
Close Contact ◽  
Lipid Storage ◽  
Metabolic Energy ◽  
Lipid Monolayer ◽  
Storage Diseases ◽  
Contact Sites ◽  
Ordered Assembly

Lipid droplets (LDs) constitute compartments dedicated to the storage of metabolic energy in the form of neutral lipids. LDs originate from the endoplasmic reticulum (ER) with which they maintain close contact throughout their life cycle. These ER–LD junctions facilitate the exchange of both proteins and lipids between these two compartments. In recent years, proteins that are important for the proper formation of LDs and localize to ER–LD junctions have been identified. This junction is unique as it is generally believed to invoke a transition from the ER bilayer membrane to a lipid monolayer that delineates LDs. Proper formation of this junction requires the ordered assembly of proteins and lipids at specialized ER subdomains. Without such a well-ordered assembly of LD biogenesis factors, neutral lipids are synthesized throughout the ER membrane, resulting in the formation of aberrant LDs. Such ectopically formed LDs impact ER and lipid homeostasis, resulting in different types of lipid storage diseases. In response to starvation, the ER–LD junction recruits factors that tether the vacuole to these junctions to facilitate LD degradation. In addition, LDs maintain close contacts with peroxisomes and mitochondria for metabolic channeling of the released fatty acids toward beta-oxidation. In this review, we discuss the function of different components that ensure proper functioning of LD contact sites, their role in lipogenesis and lipolysis, and their relation to lipid storage diseases.

scholarly journals Lipid droplets form a network interconnected by the endoplasmic reticulum through which they equilibrate their proteins

2021 ◽  
Author(s):  
Stéphanie Cottier ◽  
Roger Schneiter
Keyword(s):  
Endoplasmic Reticulum ◽  
Lipid Droplets ◽  
Neutral Lipids ◽  
Yeast Cells ◽  
Zygote Formation ◽  
Protein Transfer ◽  
Mating Partner ◽  
Functional Connection ◽  
Yeast Mating ◽  
Er Membrane

Lipid droplets (LDs) are globular intracellular structures dedicated to the storage of neutral lipids. They are closely associated with the endoplasmic reticulum (ER) and are delineated by a monolayer of phospholipids that is continuous with the cytoplasmic leaflet of the ER membrane. LDs contain a specific set of proteins, but how these proteins are targeted to the LD surface is not fully understood. Here we devised a yeast mating-based microscopic readout to monitor the transfer of LD proteins upon zygote formation. The results of this analysis indicate that ER fusion between mating partners is required for transfer of LD proteins and that this transfer is continuous, bidirectional and affects most LDs simultaneously. These observations suggest that LDs do not fuse upon mating of yeast cells, but that they form a network that is interconnected through the ER membrane. Consistent with this, ER-localized LD proteins rapidly move onto LDs of a mating partner and this protein transfer is affected by seipin, a protein important for proper LD biogenesis and the functional connection of LDs with the ER membrane.

scholarly journals ORP5 AND ORP8 ORCHESTRATE LIPID DROPLET BIOGENESIS AND MAINTENANCE AT ER-MITOCHONDRIA CONTACT SITES

2021 ◽  
Author(s):  
Valentin Guyard ◽  
Vera F Monteiro-Cardoso ◽  
Mohyeddine Omrane ◽  
Cecile Sauvanet ◽  
Audrey Houcine ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Phosphatidic Acid ◽  
Lipid Droplets ◽  
Neutral Lipids ◽  
Nucleation And Growth ◽  
Lipid Transfer ◽  
Lipid Transfer Proteins ◽  
Membrane Contact Sites ◽  
Contact Sites ◽  
Membrane Contact

Lipid droplets (LDs) are the primary organelles of lipid storage, buffering energy fluctuations of the cell. They store neutral lipids in their core that is surrounded by a protein-decorated phospholipid monolayer. LDs arise from the Endoplasmic Reticulum (ER). The ER-protein seipin, localizing at ER-LD junctions, controls LD nucleation and growth. However, how LD biogenesis is spatially and temporally coordinated remains elusive. Here, we show that the lipid transfer proteins ORP5 and ORP8 control LD biogenesis at Mitochondria-Associated ER Membrane (MAM) subdomains, enriched in phosphatidic acid. We found that ORP5/8 regulate seipin recruitment to these MAM-LD contacts, and their loss impairs LD biogenesis. Importantly, the integrity of ER-mitochondria contact sites is crucial for the ORP5/8 function in regulating seipin-mediated LD biogenesis. Our study uncovers an unprecedented ORP5/8 role in orchestrating LD biogenesis at MAMs and brings novel insights into the metabolic crosstalk between mitochondria, ER, and LDs at membrane contact sites.

scholarly journals A conserved family of proteins facilitates nascent lipid droplet budding from the ER

2015 ◽  
Vol 211 (2) ◽  
pp. 261-271 ◽  
Author(s):  
Vineet Choudhary ◽  
Namrata Ojha ◽  
Andy Golden ◽  
William A. Prinz
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Lipid Metabolism ◽  
Caenorhabditis Elegans ◽  
Lipid Droplet ◽  
Lipid Droplets ◽  
De Novo ◽  
Fat Storage ◽  
Higher Eukaryotes ◽  
Er Membrane

Lipid droplets (LDs) are found in all cells and play critical roles in lipid metabolism. De novo LD biogenesis occurs in the endoplasmic reticulum (ER) but is not well understood. We imaged early stages of LD biogenesis using electron microscopy and found that nascent LDs form lens-like structures that are in the ER membrane, raising the question of how these nascent LDs bud from the ER as they grow. We found that a conserved family of proteins, fat storage-inducing transmembrane (FIT) proteins, is required for proper budding of LDs from the ER. Elimination or reduction of FIT proteins in yeast and higher eukaryotes causes LDs to remain in the ER membrane. Deletion of the single FIT protein in Caenorhabditis elegans is lethal, suggesting that LD budding is an essential process in this organism. Our findings indicated that FIT proteins are necessary to promote budding of nascent LDs from the ER.

scholarly journals In Close Proximity: The Lipid Droplet Proteome and Crosstalk With the Endoplasmic Reticulum

Contact ◽  
2018 ◽  
Vol 1 ◽  
pp. 251525641876899 ◽  
Author(s):  
Kirill Bersuker ◽  
James A. Olzmann
Keyword(s):  
Mass Spectrometry ◽  
Endoplasmic Reticulum ◽  
Lipid Droplets ◽  
Neutral Lipids ◽  
26S Proteasome ◽  
High Confidence ◽  
Close Proximity ◽  
Bona Fide ◽  
A Site ◽  
High Degree

Lipid droplets (LDs) are conserved, endoplasmic reticulum (ER)-derived organelles that act as a dynamic cellular repository for neutral lipids. Numerous studies have examined the composition of LD proteomes by using mass spectrometry to identify proteins present in biochemically isolated buoyant fractions that are enriched in LDs. Although many bona fide LD proteins were identified, high levels of non-LD proteins that contaminate buoyant fractions complicate the detection of true LD proteins. To overcome this problem, we recently developed a proximity-labeling proteomic method to define high-confidence LD proteomes. Moreover, employing this approach, we discovered that ER-associated degradation impacts the composition of LD proteomes by targeting select LD proteins for clearance by the 26S proteasome as they transit between the ER and LDs. These findings implicate the ER as a site of LD protein degradation and underscore the high degree of crosstalk between ER and LDs.

scholarly journals Cotranslational Intersection between the SRP and GET Targeting Pathways to the Endoplasmic Reticulum of Saccharomyces cerevisiae

2016 ◽  
Vol 36 (18) ◽  
pp. 2374-2383 ◽  
Author(s):  
Ying Zhang ◽  
Thea Schäffer ◽  
Tina Wölfle ◽  
Edith Fitzke ◽  
Gerhard Thiel ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Endoplasmic Reticulum ◽  
Transmembrane Domain ◽  
Signal Recognition Particle ◽  
Transmembrane Proteins ◽  
Signal Recognition ◽  
Ribosome Binding ◽  
Channel Proteins ◽  
Membrane Spanning ◽  
Combined Data

Targeting of transmembrane proteins to the endoplasmic reticulum (ER) proceeds via either the signal recognition particle (SRP) or the guided entry of tail-anchored proteins (GET) pathway, consisting of Get1 to -5 and Sgt2. While SRP cotranslationally targets membrane proteins containing one or multiple transmembrane domains, the GET pathway posttranslationally targets proteins containing a single C-terminal transmembrane domain termed the tail anchor. Here, we dissect the roles of the SRP and GET pathways in the sorting of homologous, two-membrane-spanning K+channel proteins termed Kcv, Kesv, and Kesv-VV. We show that Kcv is targeted to the ER cotranslationally via its N-terminal transmembrane domain, while Kesv-VV is targeted posttranslationally via its C-terminal transmembrane domain, which recruits Get4-5/Sgt2 and Get3. Unexpectedly, nascent Kcv recruited not only SRP but also the Get4-5 module of the GET pathway to ribosomes. Ribosome binding of Get4-5 was independent of Sgt2 and was strongly outcompeted by SRP. The combined data indicate a previously unrecognized cotranslational interplay between the SRP and GET pathways.

scholarly journals The yeast lipin orthologue Pah1p is important for biogenesis of lipid droplets

2011 ◽  
Vol 192 (6) ◽  
pp. 1043-1055 ◽  
Author(s):  
Oludotun Adeyo ◽  
Patrick J. Horn ◽  
SungKyung Lee ◽  
Derk D. Binns ◽  
Anita Chandrahas ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Neutral Lipid ◽  
Strong Evidence ◽  
Lipid Droplets ◽  
Neutral Lipids ◽  
Glucose Medium ◽  
Wild Type ◽  
Lipid Levels ◽  
Total Neutral Lipid ◽  
A Site

Lipins are phosphatidate phosphatases that generate diacylglycerol (DAG). In this study, we report that yeast lipin, Pah1p, controls the formation of cytosolic lipid droplets. Disruption of PAH1 resulted in a 63% decrease in droplet number, although total neutral lipid levels did not change. This was accompanied by an accumulation of neutral lipids in the endoplasmic reticulum (ER). The droplet biogenesis defect was not a result of alterations in neutral lipid ratios. No droplets were visible in the absence of both PAH1 and steryl acyltransferases when grown in glucose medium, even though the strain produces as much triacylglycerol as wild type. The requirement of PAH1 for normal droplet formation can be bypassed by a knockout of DGK1. Nem1p, the activator of Pah1p, localizes to a single punctum per cell on the ER that is usually next to a droplet, suggesting that it is a site of droplet assembly. Overall, this study provides strong evidence that DAG generated by Pah1p is important for droplet biogenesis.

scholarly journals The seipin complex Fld1/Ldb16 stabilizes ER–lipid droplet contact sites

2015 ◽  
Vol 211 (4) ◽  
pp. 829-844 ◽  
Author(s):  
Alexandra Grippa ◽  
Laura Buxó ◽  
Gabriel Mora ◽  
Charlotta Funaya ◽  
Fatima-Zahra Idrissi ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Protein ◽  
Neutral Lipid ◽  
Diffusion Barrier ◽  
Lipid Droplets ◽  
Lipid Binding ◽  
Contact Sites ◽  
Packing Defects ◽  
Specific Proteins ◽  
Lipid Binding Proteins

Lipid droplets (LDs) are storage organelles consisting of a neutral lipid core surrounded by a phospholipid monolayer and a set of LD-specific proteins. Most LD components are synthesized in the endoplasmic reticulum (ER), an organelle that is often physically connected with LDs. How LD identity is established while maintaining biochemical and physical connections with the ER is not known. Here, we show that the yeast seipin Fld1, in complex with the ER membrane protein Ldb16, prevents equilibration of ER and LD surface components by stabilizing the contact sites between the two organelles. In the absence of the Fld1/Ldb16 complex, assembly of LDs results in phospholipid packing defects leading to aberrant distribution of lipid-binding proteins and abnormal LDs. We propose that the Fld1/Ldb16 complex facilitates the establishment of LD identity by acting as a diffusion barrier at the ER–LD contact sites.

scholarly journals Endoplasmic Reticulum-Vacuole Contact Sites “Bloom” With Stress-Induced Lipid Droplets

Contact ◽  
2018 ◽  
Vol 1 ◽  
pp. 251525641875611 ◽  
Author(s):  
W. Mike Henne ◽  
Hanaa Hariri
Keyword(s):  
Endoplasmic Reticulum ◽  
Recent Work ◽  
Nutrient Availability ◽  
Lipid Droplets ◽  
Nutrient Sources ◽  
Contact Sites ◽  
Long Term Storage ◽  
Specific Proteins ◽  
Term Storage

Lipid droplets (LDs) serve as specialized cytoplasmic organelles that harbor energy-rich lipids for long-term storage and may be mobilized as nutrient sources during extended starvation. How cells coordinate LD biogenesis and utilization in response to fluctuations in nutrient availability remains poorly understood. Here, we discuss our recent work revealing how yeast spatially organize LD budding at organelle contacts formed between the endoplasmic reticulum and yeast vacuole/lysosome (sites known as nucleus-vacuole junctions [NVJs]). During times of imminent nutrient exhaustion, we observe blooms of stress-induced LDs surrounding the NVJ and find that this LD clustering is regulated by NVJ-resident protein Mdm1. We also discuss several emerging studies revealing specific proteins that demarcate a subpopulation of NVJ-associated LDs. Collectively, these studies reveal a previously unappreciated role for the spatial compartmentalization of LDs at organelle contacts and highlight an important role for interorganellar cross talk in LD dynamics under times of nutritional stress.

scholarly journals Interaction mapping of endoplasmic reticulum ubiquitin ligases identifies modulators of innate immune signalling

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Emma J Fenech ◽  
Federica Lari ◽  
Philip D Charles ◽  
Roman Fischer ◽  
Marie Laétitia-Thézénas ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Interactions ◽  
Protein Quality ◽  
Transmembrane Domain ◽  
Ubiquitin Ligases ◽  
Bioinformatic Analysis ◽  
Innate Immune ◽  
Calcium Flux ◽  
Sting Pathway ◽  
Er Membrane

Ubiquitin ligases (E3s) embedded in the endoplasmic reticulum (ER) membrane regulate essential cellular activities including protein quality control, calcium flux, and sterol homeostasis. At least 25 different, transmembrane domain (TMD)-containing E3s are predicted to be ER-localised, but for most their organisation and cellular roles remain poorly defined. Using a comparative proteomic workflow, we mapped over 450 protein-protein interactions for 21 stably expressed, full-length E3s. Bioinformatic analysis linked ER-E3s and their interactors to multiple homeostatic, regulatory, and metabolic pathways. Among these were four membrane-embedded interactors of RNF26, a polytopic E3 whose abundance is auto-regulated by ubiquitin-proteasome dependent degradation. RNF26 co-assembles with TMEM43, ENDOD1, TMEM33 and TMED1 to form a complex capable of modulating innate immune signalling through the cGAS-STING pathway. This RNF26 complex represents a new modulatory axis of STING and innate immune signalling at the ER membrane. Collectively, these data reveal the broad scope of regulation and differential functionalities mediated by ER-E3s for both membrane-tethered and cytoplasmic processes.

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