RESUMEN
Lipid transport proteins (LTPs) facilitate nonvesicular lipid exchange between cellular compartments and have critical roles in lipid homeostasis1. A new family of bridge-like LTPs (BLTPs) is thought to form lipid-transporting conduits between organelles2. One, BLTP2, is conserved across species but its function is not known. Here, we show that BLTP2 and its homolog directly regulate plasma membrane (PM) fluidity by increasing the phosphatidylethanolamine (PE) level in the PM. BLTP2 localizes to endoplasmic reticulum (ER)-PM contact sites34, 5, suggesting it transports PE from the ER to the PM. We find BLTP2 works in parallel with another pathway that regulates intracellular PE distribution and PM fluidity6, 7. BLTP2 expression correlates with breast cancer aggressiveness8-10. We found BLTP2 facilitates growth of a human cancer cell line and sustains its aggressiveness in an in vivo model of metastasis, suggesting maintenance of PM fluidity by BLTP2 may be critical for tumorigenesis in humans.
RESUMEN
Membrane-bound organelles allow cells to traffic cargo and separate and regulate metabolic pathways. While many organelles are generated by the growth and division of existing organelles, some can also be produced de novo, often in response to metabolic cues. This review will discuss recent advances in our understanding of the early steps in the de novo biogenesis of peroxisomes, lipid droplets, lipoproteins, and autophagosomes. These organelles play critical roles in cellular lipid metabolism and other processes, and their dysfunction causes or is linked to several human diseases. The de novo biogenesis of these organelles occurs in or near the endoplasmic reticulum membrane. This review summarizes recent progress and highlights open questions.
Asunto(s)
Gotas Lipídicas , Peroxisomas , Humanos , Peroxisomas/metabolismo , Gotas Lipídicas/metabolismo , Autofagosomas/metabolismo , Retículo Endoplásmico/metabolismo , Metabolismo de los Lípidos , Lipoproteínas/metabolismoRESUMEN
Glycosylphosphatidylinositol (GPI) is a glycolipid membrane anchor found on surface proteins in all eukaryotes. It is synthesized in the ER membrane. Each GPI anchor requires three molecules of ethanolamine phosphate (P-Etn), which are derived from phosphatidylethanolamine (PE). We found that efficient GPI anchor synthesis in Saccharomyces cerevisiae requires Csf1; cells lacking Csf1 accumulate GPI precursors lacking P-Etn. Structure predictions suggest Csf1 is a tube-forming lipid transport protein like Vps13. Csf1 is found at contact sites between the ER and other organelles. It interacts with the ER protein Mcd4, an enzyme that adds P-Etn to nascent GPI anchors, suggesting Csf1 channels PE to Mcd4 in the ER at contact sites to support GPI anchor biosynthesis. CSF1 has orthologues in Caenorhabditis elegans (lpd-3) and humans (KIAA1109/TWEEK); mutations in KIAA1109 cause the autosomal recessive neurodevelopmental disorder Alkuraya-Kucinskas syndrome. Knockout of lpd-3 and knockdown of KIAA1109 reduced GPI-anchored proteins on the surface of cells, suggesting Csf1 orthologues in human cells support GPI anchor biosynthesis.
Asunto(s)
Retículo Endoplásmico/metabolismo , Glicosilfosfatidilinositoles/metabolismo , Fosfatidiletanolaminas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Autofagia , Mitocondrias/metabolismoRESUMEN
Lipid droplets (LDs) are ubiquitous organelles that fulfill essential roles in response to metabolic cues. The identification of several neutral lipid synthesizing and regulatory protein complexes have propelled significant advance on the mechanisms of LD biogenesis in the endoplasmic reticulum (ER). However, our understanding of signaling networks, especially transcriptional mechanisms, regulating membrane biogenesis is very limited. Here, we show that the nutrient-sensing Target of Rapamycin Complex 1 (TORC1) regulates LD formation at a transcriptional level, by targeting DGA1 expression, in a Sit4-, Mks1-, and Sfp1-dependent manner. We show that cytosolic pH (pHc), co-regulated by the plasma membrane H+-ATPase Pma1 and the vacuolar ATPase (V-ATPase), acts as a second messenger, upstream of protein kinase A (PKA), to adjust the localization and activity of the major transcription factor repressor Opi1, which in turn controls the metabolic switch between phospholipid metabolism and lipid storage. Together, this work delineates hitherto unknown molecular mechanisms that couple nutrient availability and pHc to LD formation through a transcriptional circuit regulated by major signaling transduction pathways.
Asunto(s)
Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Gotas Lipídicas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Factores de Transcripción/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/fisiología , Citosol/metabolismo , Retículo Endoplásmico/metabolismo , Concentración de Iones de Hidrógeno , Gotas Lipídicas/fisiología , Metabolismo de los Lípidos/fisiología , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/fisiología , Proteínas de la Membrana/metabolismo , Proteína Fosfatasa 2/metabolismo , Proteínas Represoras/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/fisiología , Transducción de Señal , Factores de Transcripción/fisiologíaRESUMEN
ESCRT-III proteins, which form filaments that deform, bud, and sever membranes, are found in eukaryotes and some archaea. Three studies in this issue of Cell reveal that PspA and Vipp1 are bacterial and cyanobacterial members of the ESCRT-III superfamily, indicating it is even more ubiquitous and ancient than previously thought.
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Complejos de Clasificación Endosomal Requeridos para el TransporteRESUMEN
Lipid droplets store neutral lipids, primarily triacylglycerol and steryl esters. Seipin plays a role in lipid droplet biogenesis and is thought to determine the site of lipid droplet biogenesis and the size of newly formed lipid droplets. Here we show a seipin-independent pathway of lipid droplet biogenesis. In silico and in vitro experiments reveal that retinyl esters have the intrinsic propensity to sequester and nucleate in lipid bilayers. Production of retinyl esters in mammalian and yeast cells that do not normally produce retinyl esters causes the formation of lipid droplets, even in a yeast strain that produces only retinyl esters and no other neutral lipids. Seipin does not determine the size or biogenesis site of lipid droplets composed of only retinyl esters or steryl esters. These findings indicate that the role of seipin in lipid droplet biogenesis depends on the type of neutral lipid stored in forming droplets.
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Subunidades gamma de la Proteína de Unión al GTP/metabolismo , Gotas Lipídicas/metabolismo , Ésteres de Retinilo/metabolismo , Triglicéridos/metabolismo , Animales , Células Cultivadas , Cricetulus , Subunidades gamma de la Proteína de Unión al GTP/deficiencia , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones TransgénicosRESUMEN
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 colocalize 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 general role in linking tubular ER to organelle contact sites.
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Retículo Endoplásmico/metabolismo , Gotas Lipídicas/metabolismo , Proteínas de la Membrana/metabolismo , Animales , Dominios C2 , Células COS , Chlorocebus aethiops , Células HeLa , HumanosRESUMEN
The VPS13 gene family consists of VPS13A-D in mammals. Although all four genes have been linked to human diseases, their cellular functions are poorly understood, particularly those of VPS13D. We generated and characterized knockouts of each VPS13 gene in HeLa cells. Among the individual knockouts, only VPS13D-KO cells exhibit abnormal mitochondrial morphology. Additionally, VPS13D loss leads to either partial or complete peroxisome loss in several transformed cell lines and in fibroblasts derived from a VPS13D mutation-carrying patient with recessive spinocerebellar ataxia. Our data show that VPS13D regulates peroxisome biogenesis.
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Peroxisomas/genética , Peroxisomas/metabolismo , Proteínas/genética , Proteínas/metabolismo , Células HEK293 , Células HeLa , Humanos , Mitocondrias/genética , Mitocondrias/metabolismo , Mutación/genéticaRESUMEN
We have long known that lipids traffic between cellular membranes via vesicles but have only recently appreciated the role of nonvesicular lipid transport. Nonvesicular transport can be high volume, supporting biogenesis of rapidly expanding membranes, or more targeted and precise, allowing cells to rapidly alter levels of specific lipids in membranes. Most such transport probably occurs at membrane contact sites, where organelles are closely apposed, and requires lipid transport proteins (LTPs), which solubilize lipids to shield them from the aqueous phase during their transport between membranes. Some LTPs are cup like and shuttle lipid monomers between membranes. Others form conduits allowing lipid flow between membranes. This review describes what we know about nonvesicular lipid transfer mechanisms while also identifying many remaining unknowns: How do LTPs facilitate lipid movement from and into membranes, do LTPs require accessory proteins for efficient transfer in vivo, and how is directionality of transport determined?
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Metabolismo de los Lípidos , Vesículas Transportadoras/metabolismo , Animales , Transporte Biológico , Humanos , Modelos Biológicos , Mutación/genéticaRESUMEN
Lipid droplets (LDs) are implicated in conditions of lipid and protein dysregulation. The fat storage-inducing transmembrane (FIT; also known as FITM) family induces LD formation. Here, we establish a model system to study the role of the Saccharomyces cerevisiae FIT homologues (ScFIT), SCS3 and YFT2, in the proteostasis and stress response pathways. While LD biogenesis and basal endoplasmic reticulum (ER) stress-induced unfolded protein response (UPR) remain unaltered in ScFIT mutants, SCS3 was found to be essential for proper stress-induced UPR activation and for viability in the absence of the sole yeast UPR transducer IRE1 Owing to not having a functional UPR, cells with mutated SCS3 exhibited an accumulation of triacylglycerol within the ER along with aberrant LD morphology, suggesting that there is a UPR-dependent compensatory mechanism that acts to mitigate lack of SCS3 Additionally, SCS3 was necessary to maintain phospholipid homeostasis. Strikingly, global protein ubiquitylation and the turnover of both ER and cytoplasmic misfolded proteins is impaired in ScFITΔ cells, while a screen for interacting partners of Scs3 identifies components of the proteostatic machinery as putative targets. Together, our data support a model where ScFITs play an important role in lipid metabolism and proteostasis beyond their defined roles in LD biogenesis.This article has an associated First Person interview with the first author of the paper.
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Lípidos de la Membrana , Saccharomyces cerevisiae , Retículo Endoplásmico/genética , Retículo Endoplásmico/metabolismo , Estrés del Retículo Endoplásmico/genética , Homeostasis , Lípidos de la Membrana/metabolismo , Proteostasis , Saccharomyces cerevisiae/genética , Respuesta de Proteína Desplegada/genéticaRESUMEN
All lipid transport proteins in eukaryotes are thought to shuttle lipids between cellular membranes. In this issue, Li et al. (2020. J. Cell Biol.https://doi.org/10.1083/jcb.202001161) show that Vps13 has a channel-like domain that may allow lipids to flow between closely apposed membranes at contact sites.
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Microscopía por Crioelectrón , Fosfolípidos , Membrana Celular , Membranas , Membranas MitocondrialesRESUMEN
Lipid droplets (LDs) are fat storage organelles that originate from the endoplasmic reticulum (ER). Relatively little is known about how sites of LD formation are selected and which proteins/lipids are necessary for the process. Here, we show that LDs induced by the yeast triacylglycerol (TAG)-synthases Lro1 and Dga1 are formed at discrete ER subdomains defined by seipin (Fld1), and a regulator of diacylglycerol (DAG) production, Nem1. Fld1 and Nem1 colocalize to ER-LD contact sites. We find that Fld1 and Nem1 localize to ER subdomains independently of each other and of LDs, but both are required for the subdomains to recruit the TAG-synthases and additional LD biogenesis factors: Yft2, Pex30, Pet10, and Erg6. These subdomains become enriched in DAG. We conclude that Fld1 and Nem1 are both necessary to recruit proteins to ER subdomains where LD biogenesis occurs.
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Retículo Endoplásmico/metabolismo , Gotas Lipídicas/metabolismo , Metabolismo de los Lípidos/genética , Proteínas de la Membrana/genética , Proteínas Nucleares/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Diacilglicerol O-Acetiltransferasa/genética , Diacilglicerol O-Acetiltransferasa/metabolismo , Diglicéridos/biosíntesis , Retículo Endoplásmico/genética , Regulación Fúngica de la Expresión Génica , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Proteínas de la Membrana/metabolismo , Metiltransferasas/genética , Metiltransferasas/metabolismo , Proteínas Nucleares/metabolismo , Biogénesis de Organelos , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Transducción de Señal , Triglicéridos/biosíntesis , Proteína Fluorescente RojaRESUMEN
Organelles compartmentalize eukaryotic cells, enhancing their ability to respond to environmental and developmental changes. One way in which organelles communicate and integrate their activities is by forming close contacts, often called 'membrane contact sites' (MCSs). Interest in MCSs has grown dramatically in the past decade as it is has become clear that they are ubiquitous and have a much broader range of critical roles in cells than was initially thought. Indeed, functions for MCSs in intracellular signalling (particularly calcium signalling, reactive oxygen species signalling and lipid signalling), autophagy, lipid metabolism, membrane dynamics, cellular stress responses and organelle trafficking and biogenesis have now been reported.
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Membrana Celular/metabolismo , Metabolismo de los Lípidos , Transducción de Señal , Estrés Fisiológico/fisiología , Animales , Autofagosomas/metabolismo , Autofagia , Transporte Biológico , Señalización del Calcio , Membrana Celular/química , Retículo Endoplásmico/metabolismo , Enzimas/metabolismo , Células Eucariotas/metabolismo , Humanos , Membranas Mitocondriales/metabolismo , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Nascent lipid droplet (LD) formation occurs in the endoplasmic reticulum (ER) membrane but it is not known how sites of biogenesis are determined. We previously identified ER domains in S. cerevisiae containing the reticulon homology domain (RHD) protein Pex30 that are regions where preperoxisomal vesicles (PPVs) form. Here, we show that Pex30 domains are also sites where most nascent LDs form. Mature LDs usually remain associated with Pex30 subdomains, and the same Pex30 subdomain can simultaneously associate with a LD and a PPV or peroxisome. We find that in higher eukaryotes multiple C2 domain containing transmembrane protein (MCTP2) is similar to Pex30: it contains an RHD and resides in ER domains where most nascent LD biogenesis occurs and that often associate with peroxisomes. Together, these findings indicate that most LDs and PPVs form and remain associated with conserved ER subdomains, and suggest a link between LD and peroxisome biogenesis.
Asunto(s)
Retículo Endoplásmico/metabolismo , Gotas Lipídicas/metabolismo , Biogénesis de Organelos , Peroxisomas/metabolismo , Saccharomyces cerevisiae/metabolismo , Animales , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Diacilglicerol O-Acetiltransferasa/metabolismo , Subunidades gamma de la Proteína de Unión al GTP/genética , Subunidades gamma de la Proteína de Unión al GTP/metabolismo , Eliminación de Gen , Células HeLa , Humanos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Proteínas de Transporte de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Metiltransferasas/metabolismo , Mutación , Dominios Proteicos , Saccharomyces cerevisiae/citología , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismoRESUMEN
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.
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Proteínas de Transporte de Catión/metabolismo , Glicoproteínas/metabolismo , Gotas Lipídicas/metabolismo , Gotas Lipídicas/fisiología , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Transporte de Catión/fisiología , Simulación por Computador , Diglicéridos/metabolismo , Diglicéridos/fisiología , Retículo Endoplásmico/metabolismo , Retículo Endoplásmico/fisiología , Glicoproteínas/fisiología , Proteínas Asociadas a Gotas Lipídicas/metabolismo , Proteínas Asociadas a Gotas Lipídicas/fisiología , Metabolismo de los Lípidos/fisiología , Proteínas de la Membrana/metabolismo , Fosfatidiletanolaminas/metabolismo , Fosfolípidos/fisiología , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/fisiologíaRESUMEN
Recent studies have reported intrinsic metabolic reprogramming in Pkd1 knock-out cells, implicating dysregulated cellular metabolism in the pathogenesis of polycystic kidney disease. However, the exact nature of the metabolic changes and their underlying cause remains controversial. We show herein that Pkd1 k o /ko renal epithelial cells have impaired fatty acid utilization, abnormal mitochondrial morphology and function, and that mitochondria in kidneys of ADPKD patients have morphological alterations. We further show that a C-terminal cleavage product of polycystin-1 (CTT) translocates to the mitochondria matrix and that expression of CTT in Pkd1 ko/ko cells rescues some of the mitochondrial phenotypes. Using Drosophila to model in vivo effects, we find that transgenic expression of mouse CTT results in decreased viability and exercise endurance but increased CO2 production, consistent with altered mitochondrial function. Our results suggest that PC1 may play a direct role in regulating mitochondrial function and cellular metabolism and provide a framework to understand how impaired mitochondrial function could be linked to the regulation of tubular diameter in both physiological and pathological conditions.
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Riñón , Mitocondrias , Proteínas Mitocondriales/metabolismo , Riñón Poliquístico Autosómico Dominante/metabolismo , Proteolisis , Canales Catiónicos TRPP/metabolismo , Anciano , Animales , Animales Modificados Genéticamente , Perros , Drosophila melanogaster , Embrión de Mamíferos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Ácidos Grasos/metabolismo , Técnicas de Silenciamiento del Gen , Humanos , Riñón/metabolismo , Riñón/patología , Células de Riñón Canino Madin Darby , Masculino , Ratones , Persona de Mediana Edad , Mitocondrias/metabolismo , Mitocondrias/patología , Proteínas Mitocondriales/genética , Canales Catiónicos TRPP/genéticaRESUMEN
Spatial organization of phospholipid synthesis in eukaryotes is critical for cellular homeostasis. The synthesis of phosphatidylcholine (PC), the most abundant cellular phospholipid, occurs redundantly via the ER-localized Kennedy pathway and a pathway that traverses the ER and mitochondria via membrane contact sites. The basis of the ER-mitochondrial PC synthesis pathway is the exclusive mitochondrial localization of a key pathway enzyme, phosphatidylserine decarboxylase Psd1, which generates phosphatidylethanolamine (PE). We find that Psd1 is localized to both mitochondria and the ER. Our data indicate that Psd1-dependent PE made at mitochondria and the ER has separable cellular functions. In addition, the relative organellar localization of Psd1 is dynamically modulated based on metabolic needs. These data reveal a critical role for ER-localized Psd1 in cellular phospholipid homeostasis, question the significance of an ER-mitochondrial PC synthesis pathway to cellular phospholipid homeostasis, and establish the importance of fine spatial regulation of lipid biosynthesis for cellular functions.
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Carboxiliasas/metabolismo , Retículo Endoplásmico/enzimología , Mitocondrias/enzimología , Proteínas Mitocondriales/metabolismo , Fosfatidiletanolaminas/metabolismo , Carboxiliasas/química , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Homeostasis , Mitocondrias/metabolismo , Proteínas Mitocondriales/química , Señales de Clasificación de ProteínaRESUMEN
Squalene monooxygenase (SM), which synthesizes a cholesterol precursor, is degraded when cholesterol levels in the endoplasmic reticulum (ER) membrane are high, but the signal for degradation was not known. In this issue of JBC, Brown and co-workers identify an N-terminal domain in SM that interconverts in a cholesterol-sensitive manner between a membrane-binding amphipathic helix and a soluble degradation-prone segment, providing the first example of a cholesterol-degron collaboration.
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Colesterol , Retículo Endoplásmico , Humanos , Complejo de la Endopetidasa Proteasomal , Escualeno-MonooxigenasaRESUMEN
Mitochondria constantly divide and fuse. Homotypic fusion of the outer mitochondrial membranes requires the mitofusin (MFN) proteins, a family of dynamin-like GTPases. MFNs are anchored in the membrane by transmembrane (TM) segments, exposing both the N-terminal GTPase domain and the C-terminal tail (CT) to the cytosol. This arrangement is very similar to that of the atlastin (ATL) GTPases, which mediate fusion of endoplasmic reticulum (ER) membranes. We engineered various MFN-ATL chimeras to gain mechanistic insight into MFN-mediated fusion. When MFN1 is localized to the ER by TM swapping with ATL1, it functions in the maintenance of ER morphology and fusion. In addition, an amphipathic helix in the CT of MFN1 is exchangeable with that of ATL1 and critical for mitochondrial localization of MFN1. Furthermore, hydrophobic residues N-terminal to the TM segments of MFN1 play a role in membrane targeting but not fusion. Our findings provide important insight into MFN-mediated membrane fusion.