Targeting of Nir2 to Lipid Droplets Is Regulated by a Specific Threonine Residue within Its PI-Transfer Domain

Litvak, Vladimir; Shaul, Yoav D.; Shulewitz, Mark J.; Amarilio, Roy; Carmon, Shari; Lev, Sima

doi:10.1016/s0960-9822(02)01107-7

Cited by 49 publications

(40 citation statements)

References 24 publications

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“…using indirect immunofluorescence and confocal microscopy analysis. OSBP ( Figure 3A), CERT ( Figure 3B), and Nir2 ( Figure 3C) were localized mainly to the Golgi complex of control cells, consistent with previous reports (Storey et al, 1998;Litvak et al, 2002), and treatment with 25OH markedly enhanced their Golgi association, as determined by colocalization with Golgi markers. This treatment also enhanced their interaction with VAPs as demonstrated by coimmunoprecipitation studies ( Figure 3D).…”

Section: Depletion Of Vaps Affects the Golgi Targeting Of Nir2 Osbpsupporting

confidence: 91%

Coordinated Lipid Transfer between the Endoplasmic Reticulum and the Golgi Complex Requires the VAP Proteins and Is Essential for Golgi-mediated Transport

Peretti

Dahan

Shimoni

et al. 2008

MBoC

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286

272

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Lipid transport between intracellular organelles is mediated by vesicular and nonvesicular transport mechanisms and is critical for maintaining the identities of different cellular membranes. Nonvesicular lipid transport between the endoplasmic reticulum (ER) and the Golgi complex has been proposed to affect the lipid composition of the Golgi membranes. Here, we show that the integral ER-membrane proteins VAP-A and VAP-B affect the structural and functional integrity of the Golgi complex. Depletion of VAPs by RNA interference reduces the levels of phosphatidylinositol-4-phosphate (PI4P), diacylglycerol, and sphingomyelin in the Golgi membranes, and it leads to substantial inhibition of Golgimediated transport events. These effects are coordinately mediated by the lipid-transfer/binding proteins Nir2, oxysterolbinding protein (OSBP), and ceramide-transfer protein (CERT), which interact with VAPs via their FFAT motif. The effect of VAPs on PI4P levels is mediated by the phosphatidylinositol/phosphatidylcholine transfer protein Nir2, which is required for Golgi targeting of OSBP and CERT and the subsequent production of diacylglycerol and sphingomyelin. We propose that Nir2, OSBP, and CERT function coordinately at the ER-Golgi membrane contact sites, thereby affecting the lipid composition of the Golgi membranes and consequently their structural and functional identities. INTRODUCTIONThe unique lipid compositions of the secretory and endocytic organelles are critical for maintaining their distinct structural and functional identities (van Meer, 2000). Their identities are maintained despite extensive interconnecting vesicular-trafficking pathways, and they require tight regulation of lipid sorting, metabolism, and transport (van Meer, 1993;Pomorski et al., 2001;van Meer and Sprong, 2004). Increasing lines of evidence suggest that lipid-transfer proteins (LTPs) play a major role in regulating the lipid composition of membranous organelles (Sprong et al., 2001;De Matteis et al., 2007). These proteins facilitate lipid transfer between a donor and acceptor membrane (Holthuis and Levine, 2005), and they usually contain dual-targeting determinants for different membrane compartments. LTPs have been proposed to efficiently transfer lipids at membrane contact sites (MCSs) (Holthuis and Levine, 2005;Levine and Loewen, 2006).MCSs or zones of close apposition between the ER membranes and other cellular membranes, including the plasma membrane (PM), the membranes of the vacuoles, mitochondria, peroxisomes, lipid droplets, late endosomes, lysosomes, and the Golgi apparatus, have been identified in all eukaryotes by morphological and biochemical studies (Shore and Tata, 1977;Levine, 2004;Levine and Loewen, 2006). More recently, electron tomography studies have identified close contacts (10 -20 nm) between the ER and the outer mitochondrial membrane (Perkins et al., 1997) and between a specialized trans-ER and the three trans-most cisternae of the Golgi complex (Ladinsky et al., 1999;Marsh et al., 2001Marsh et al., , 2004. This ...

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Section: Depletion Of Vaps Affects the Golgi Targeting Of Nir2 Osbpsupporting

confidence: 91%

Coordinated Lipid Transfer between the Endoplasmic Reticulum and the Golgi Complex Requires the VAP Proteins and Is Essential for Golgi-mediated Transport

Peretti

Dahan

Shimoni

et al. 2008

MBoC

Self Cite

286

272

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“…We have previously shown that Nir2 mainly localizes in the Golgi apparatus, but it is also found in the ER in interphase cells (36). To determine the localization of VAP-B in HeLa cells, we used the anti-VAP-B antibody in indirect immunofluorescence and confocal microscopy analyses.…”

Section: Nirs Interact With Vap-b Via Their Conserved Ffat Motif-mentioning

confidence: 99%

Differential Regulation of Endoplasmic Reticulum Structure through VAP-Nir Protein Interaction

Amarilio

Ramachandran

Sabanay

et al. 2005

Journal of Biological Chemistry

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178

161

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The endoplasmic reticulum (ER) exhibits a characteristic tubular structure that is dynamically rearranged in response to specific physiological demands. However, the mechanisms by which the ER maintains its characteristic structure are largely unknown. Here we show that the integral ER-membrane protein VAP-B causes a striking rearrangement of the ER through interaction with the Nir2 and Nir3 proteins. We provide evidence that Nir (Nir1, Nir2, and Nir3)-VAP-B interactions are mediated through the conserved FFAT (two phenylalanines (FF) in acidic tract) motif present in Nir proteins. However, each interaction affects the structural integrity of the ER differently. Whereas the Nir2-VAP-B interaction induces the formation of stacked ER membrane arrays, the Nir3-VAP-B interaction leads to a gross remodeling of the ER and the bundling of thick microtubules along the altered ER membranes. In contrast, the Nir1-VAP-B interaction has no apparent effect on ER structure. We also show that the Nir2-VAP-B interaction attenuates protein export from the ER. These results demonstrate new mechanisms for the regulation of ER structure, all of which are mediated through interaction with an identical integral ER-membrane protein. The endoplasmic reticulum (ER)1 is an extensive network of membranes comprised of an array of interconnecting tubules and cisternae that emerges from the nuclear envelope (NE) and extends peripherally throughout the cell cytoplasm (1). It contains several structurally distinct domains, including the NE, the rough and smooth ER (rER and sER), and the regions that contact other organelles, such as the Golgi apparatus, the late endosomes, the lysosomes, mitochondria, peroxisomes, and the plasma membrane (2). The ER functions in diverse metabolic processes including lipid synthesis, carbohydrate metabolism, and the detoxification of drugs. It is responsible for the synthesis, translocation, glycosylation, folding, assembly, and processing of secretory and membrane proteins, and it functions in intracellular calcium storage and sequestering (3, 4).While the function of the ER in membrane trafficking, lipid biosynthesis, and calcium signaling have been extensively studied, the mechanism by which the ER maintains its characteristic structure in vivo remains largely unknown. Studies from yeast and mammalian cells have shown that the size and/or structure of the ER is extremely sensitive to certain cellular stress conditions, such as the unfolded protein response (UPR) or to the overexpression of a subset of ERresident membrane proteins (5, 6). Overexpression of 3-hydroxymethyl-3-methylglutaryl coenzyme A (HMG-CoA) reductase, microsomal aldehyde dehydrogenase (msALDH), cytochrome P-450, and malfolded cytochrome P-450, causes the proliferation of ER membranes and stacking of the ER cisternae into organized structures known as crystalloid ER, sinusoidal ER, or karmellae (7-12). The formation of these structures is easily visible and can be used as a quantitative method for studying ER membrane biogenesis (5). Neverth...

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“…Nir2 is not an integral, membrane-spanning protein but is associated to Golgi/endoplasmatic reticulum membranes. A T59E mutation in Nir2 changed the cellular localization to spherical cytosolic lipid droplets [30]. It is suggested that phosphorylation of Thr59 and relocalization of Nir2 is stimulated by incubation with oleic acid.…”

Section: Phosphorylation Of Pi-tpa and Pi-tpb In Vitro And In Vivomentioning

confidence: 99%

Structure–function relationships of phosphatidylinositol transfer proteins: involvement of phosphorylation sites

2004

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The mammalian low molecular weight phosphatidylinositol transfer proteins: PI-TPa and PI-TPb are extremely well conserved and highly homologous. Surprisingly, the two proteins clearly show different cellular localizations and display contrary physiological functions. Phosphorylation of the proteins might be the regulatory factor to ensure the selective cellular functions. A major difference between PI-TPa and PI-TPb is the capacity of PI-TPb to bind and transfer sphingomyelin (SM) in vivo. This activity is correlated with phosphorylation of Ser262, which is only present in PI-TPb. Structural aspects of phosphorylation sites of PI-TPs are analyzed in order to find an explanation for the functional data. We propose that phosphorylation of one serine residue (Ser165/166) in both PI-TPa and PI-TPb is involved in the regulation of membrane binding of all PI-TPs and that phosphorylation of the unique Ser262 in PI-TPb-like proteins ensures the right cellular localization of PI-TPbs that is necessary for the specific activity at the Golgi membrane.

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Targeting of Nir2 to Lipid Droplets Is Regulated by a Specific Threonine Residue within Its PI-Transfer Domain

Cited by 49 publications

References 24 publications

Coordinated Lipid Transfer between the Endoplasmic Reticulum and the Golgi Complex Requires the VAP Proteins and Is Essential for Golgi-mediated Transport

Coordinated Lipid Transfer between the Endoplasmic Reticulum and the Golgi Complex Requires the VAP Proteins and Is Essential for Golgi-mediated Transport

Differential Regulation of Endoplasmic Reticulum Structure through VAP-Nir Protein Interaction

Structure–function relationships of phosphatidylinositol transfer proteins: involvement of phosphorylation sites

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