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

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.

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

Contact ◽

10.1177/2515256418768996 ◽

2018 ◽

Vol 1 ◽

pp. 251525641876899 ◽

Cited By ~ 5

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.

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Targeted disruption of the aromatase P450 gene (Cyp19) in mice and their ovarian and uterine responses to 17beta-oestradiol

Journal of Endocrinology ◽

10.1677/joe.0.1700099 ◽

2001 ◽

Vol 170 (1) ◽

pp. 99-111 ◽

Cited By ~ 87

Author(s):

K Toda ◽

K Takeda ◽

T Okada ◽

S Akira ◽

T Saibara ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Corpus Luteum ◽

Lipid Droplets ◽

Smooth Endoplasmic Reticulum ◽

Ovarian Follicles ◽

Interstitial Cells ◽

Aromatase Activity ◽

Tubular Structures ◽

Wild Type ◽

Targeted Disruption

Aromatase P450 (CYP19) is an enzyme catalysing the conversion of androgens into oestrogens. We generated mice lacking aromatase activity (ArKO) by targeted disruption of Cyp19 and report the characteristic features of the ArKO ovaries and uteri as revealed by histological and biochemical analyses. ArKO females were totally infertile but there were as many developing follicles in their ovaries at 8 weeks of age as in wild-type ovaries. Nevertheless, no typical corpus luteum was observed in the ArKO ovaries. Electron microscopy revealed the presence of well-developed smooth endoplasmic reticulum, few lipid droplets and mitochondria with less organized tubular structures in the ArKO luteinized interstitial cells. These ultrastructural features were different from those of the wild-type interstitial cells, where there are many lipid droplets and mitochondria with well-developed tubular structures, characteristic of steroid-producing cells. When ArKO mice were supplemented with 17beta-oestradiol (E(2); 15 microg/mouse) every fourth day from 4 weeks of age for 1 month, increased numbers of follicles were observed in the ovaries as compared with those of untreated ArKO mice, although no typical corpus luteum was detectable. Ultrastructural analysis revealed the disappearance of the accumulated smooth endoplasmic reticulum in the luteinized interstitial cells after E(2 )supplementation. Transcripts of pro-apoptotic genes such as p53 and Bax genes were markedly elevated in the ArKO ovaries as compared with those of wild-type mice. Although E(2) supplementation did not cause suppression of the elevated expression of p53 and Bax mRNAs, it caused marked enhancement of expression levels of lactoferrin and progesterone receptor mRNAs in the uteri as well as increases in uterine wet weight. At 8 months of age, ArKO mice developed haemorrhages in the ovaries, in which follicles were nearly depleted, while age-matched wild-type females still had many ovarian follicles. Furthermore, macrophage-like cells were occasionally observed in the ArKO ovarian follicles. These results suggested that targeted disruption of Cyp19 caused anovulation and precocious depletion of ovarian follicles. Additionally, analysis of mice supplemented with E(2) demonstrated that E(2) apparently supports development of ovarian follicles, although it did not restore the defect in ovulation.

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

The Journal of Cell Biology ◽

10.1083/jcb.201502070 ◽

2015 ◽

Vol 211 (4) ◽

pp. 829-844 ◽

Cited By ~ 106

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.

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

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.650186 ◽

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.

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

Molecular Biology of the Cell ◽

10.1091/mbc.e20-09-0590 ◽

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]

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Neutral lipids regulate amphipathic helix affinity for model lipid droplets

The Journal of Cell Biology ◽

10.1083/jcb.201907099 ◽

2020 ◽

Vol 219 (4) ◽

Cited By ~ 8

Author(s):

Aymeric Chorlay ◽

Abdou Rachid Thiam

Keyword(s):

Neutral Lipid ◽

Lipid Membranes ◽

Lipid Droplets ◽

Neutral Lipids ◽

Aqueous Environment ◽

Major Step ◽

Amphipathic Helices ◽

Binding Preference ◽

Hydrophobic Nature ◽

Binding Level

Cellular lipid droplets (LDs) have a neutral lipid core shielded from the aqueous environment by a phospholipid monolayer containing proteins. These proteins define the biological functions of LDs, and most of them bear amphipathic helices (AH), which can selectively target to LDs, or to LD subsets. How such binding preference happens remains poorly understood. Here, we found that artificial LDs made of different neutral lipids but presenting equal phospholipid packing densities differentially recruit AHs. Varying the phospholipid density shifts the binding levels, but the differential recruitment is unchanged. We found that the binding level of AHs is defined by their interaction preference with neutral lipids and ability to decrease surface tension. The phospholipid packing level regulates mainly the amount of neutral lipid accessible. Therefore, it is the hydrophobic nature of the phospholipid packing voids that controls the binding level of AHs. Our data bring us a major step closer to understanding the binding selectivity of AHs to lipid membranes.

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Regulation of the yeast triacylglycerol lipases Tgl4p and Tgl5p by the presence/absence of nonpolar lipids

Molecular Biology of the Cell ◽

10.1091/mbc.e15-09-0633 ◽

2016 ◽

Vol 27 (13) ◽

pp. 2014-2024 ◽

Cited By ~ 7

Author(s):

Isabella Klein ◽

Lisa Klug ◽

Claudia Schmidt ◽

Martina Zandl ◽

Martina Korber ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Lipid Droplets ◽

Lipolytic Activity ◽

Wild Type ◽

Yeast Saccharomyces Cerevisiae ◽

Steryl Esters ◽

Lysophospholipid Acyltransferases ◽

Nonpolar Lipids ◽

The Stability ◽

In Cells

Tgl3p, Tgl4p, and Tgl5p are the major triacylglycerol lipases of the yeast Saccharomyces cerevisiae. Recently we demonstrated that properties of Tgl3p are regulated by the formation of nonpolar lipids. The present study extends these investigations to the two other yeast triacylglycerol lipases, Tgl4p and Tgl5p. We show that Tgl4p and Tgl5p, which are localized to lipid droplets in wild type, are partially retained in the endoplasmic reticulum in cells lacking triacylglycerols and localize exclusively to the endoplasmic reticulum in a mutant devoid of lipid droplets. In cells lacking steryl esters, the subcellular distribution of Tgl4p and Tgl5p is unaffected, but Tgl5p becomes unstable, whereas the stability of Tgl4p increases. In cells lacking nonpolar lipids, Tgl4p and Tgl5p lose their lipolytic activity but retain their side activity as lysophospholipid acyltransferases. To investigate the regulatory network of yeast triacylglycerol lipases in more detail, we also examined properties of Tgl3p, Tgl4p, and Tgl5p, respectively, in the absence of the other lipases. Surprisingly, lack of two lipases did not affect expression, localization, and stability of the remaining Tgl protein. These results suggest that Tgl3p, Tgl4p, and Tgl5p, although they exhibit similar functions, act as independent entities.

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Motility Plays an Important Role in the Lifetime of Mammalian Lipid Droplets

International Journal of Molecular Sciences ◽

10.3390/ijms22083802 ◽

2021 ◽

Vol 22 (8) ◽

pp. 3802

Author(s):

Yi Jin ◽

Zhuqing Ren ◽

Yanjie Tan ◽

Pengxiang Zhao ◽

Jian Wu

Keyword(s):

Signal Transduction ◽

Endoplasmic Reticulum ◽

Lipid Droplet ◽

Molecular Basis ◽

Lipid Droplets ◽

Neutral Lipids ◽

Future Research ◽

Biological Processes ◽

Cellular Processes ◽

Resistance To Stress

The lipid droplet is a kind of organelle that stores neutral lipids in cells. Recent studies have found that in addition to energy storage, lipid droplets also play an important role in biological processes such as resistance to stress, immunity, cell proliferation, apoptosis, and signal transduction. Lipid droplets are formed at the endoplasmic reticulum, and mature lipid droplets participate in various cellular processes. Lipid droplets are decomposed by lipase and lysosomes. In the life of a lipid droplet, the most important thing is to interact with other organelles, including the endoplasmic reticulum, mitochondria, peroxisomes, and autophagic lysosomes. The interaction between lipid droplets and other organelles requires them to be close to each other, which inevitably involves the motility of lipid droplets. In fact, through many microscopic observation techniques, researchers have discovered that lipid droplets are highly dynamic organelles that move quickly. This paper reviews the process of lipid droplet motility, focusing on explaining the molecular basis of lipid droplet motility, the factors that regulate lipid droplet motility, and the influence of motility on the formation and decomposition of lipid droplets. In addition, this paper also proposes several unresolved problems for lipid droplet motility. Finally, this paper makes predictions about the future research of lipid droplet motility.

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Seipin accumulates and traps diacylglycerols and triglycerides in its ring-like structure

10.1101/2020.10.27.357079 ◽

2020 ◽

Author(s):

Valeria Zoni ◽

Wataru Shinoda ◽

Stefano Vanni

Keyword(s):

Molecular Dynamics ◽

Endoplasmic Reticulum ◽

Molecular Dynamics Simulations ◽

Lipid Droplets ◽

Neutral Lipids ◽

Health Concern ◽

Intracellular Organelles ◽

Dynamics Simulations ◽

Precise Role

AbstractLipid droplets (LD) are intracellular organelles responsible for lipid storage, and they emerge from the endoplasmic reticulum (ER) upon the accumulation of neutral lipids, mostly triglycerides (TG), between the two leaflets of the ER membrane. LD biogenesis takes place at ER sites that are marked by the protein seipin, which subsequently recruits additional proteins to catalyse LD formation. Deletion of seipin, however, does not abolish LD biogenesis, and its precise role in controlling LD assembly remains unclear. Here we use molecular dynamics simulations to investigate the molecular mechanism through which seipin promotes LD formation. We find that seipin clusters TG molecules inside its unconventional ring-like oligomeric structure, and that both its luminal and transmembrane regions contribute to this process. Diacylglycerol, the precursor of TG, also clusters inside the seipin oligomer, in turn promoting TG accumulation. Our results suggest that seipin remodels the membrane of specific ER sites to prime them for LD biogenesis.Significance statementMetabolic disorders related to aberrant fat accumulation, including lipodystrophy and obesity, are a particularly serious health concern. In cells, fat accumulates in intracellular organelles, named lipid droplets (LDs). LDs form in the endoplasmic reticulum, where triglycerides, the most abundant form of fat, is produced. The Bernardinelli-Seip congenital lipodystrophy type 2 protein, seipin, has been identified as a key regulator of LD formation, but its mechanism of action remains debated and its molecular details mostly obscure. Here, we use molecular dynamics simulations to investigate the mechanism of seipin. We find that seipin can cluster and trap both triglycerides and its precursor, diacylglycerol. Our results suggest that seipin organizes the lipid composition of specific ER sites to prime them for LD biogenesis.

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Pre-existing bilayer stresses modulate triglyceride accumulation in the ER versus lipid droplets

eLife ◽

10.7554/elife.62886 ◽

2021 ◽

Vol 10 ◽

Author(s):

Valeria Zoni ◽

Rasha Khaddaj ◽

Pablo Campomanes ◽

Abdou Rachid Thiam ◽

Roger Schneiter ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Fluorescence Microscopy ◽

Conceptual Understanding ◽

Lipid Droplets ◽

Neutral Lipids ◽

Yeast Genetics ◽

Triglyceride Accumulation ◽

Acyl Chains ◽

And Function ◽

Er Homeostasis

Cells store energy in the form of neutral lipids (NLs) packaged into micrometer-sized organelles named lipid droplets (LDs). These structures emerge from the endoplasmic reticulum (ER) at sites marked by the protein seipin, but the mechanisms regulating their biogenesis remain poorly understood. Using a combination of molecular simulations, yeast genetics, and fluorescence microscopy, we show that interactions between lipids’ acyl-chains modulate the propensity of NLs to be stored in LDs, in turn preventing or promoting their accumulation in the ER membrane. Our data suggest that diacylglycerol, which is enriched at sites of LD formation, promotes the packaging of NLs into LDs, together with ER-abundant lipids, such as phosphatidylethanolamine. On the opposite end, short and saturated acyl-chains antagonize fat storage in LDs and promote accumulation of NLs in the ER. Our results provide a new conceptual understanding of LD biogenesis in the context of ER homeostasis and function.

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