Lipid Trafficking in Cells

Synonyms

Intracellular lipid transport; Membrane traffic; Nonvesicular transport

Definition

The processes by which lipids move among organelles by vesicular trafficking processes or by diffusion through the cytoplasm in association with carrier proteins.

Introduction

Lipids play many important roles in cell physiology. Lipid bilayer membranes provide the main barrier separating the interior and exterior of a cell, restricting the transport of ions and polar molecules in and out of the cell. Lipid bilayers similarly provide a permeability barrier between the cytoplasm and the interior of organelles. The lipid composition of biological membranes must be maintained in a range that allows rapid diffusion of proteins within the bilayer while maintaining the barrier function. Several lipid species can be acted upon enzymatically to produce signaling second messengers that activate downstream signaling networks. The organization of lipids in a membrane bilayer can also affect the signaling properties of receptors and thereby modulating intracellular signal transduction. Organelles within a cell maintain lipid compositions that are individually distinct, and aspects of these lipid differences are an important component of the “identity” of the organelles that is used by proteins to interact selectively with certain organelles. The distinct lipid composition of these organelles is maintained even though there is a very robust transport of lipids between organelles. The processes regulating lipid traffic and the mechanisms for establishing and maintaining lipid composition of organelles are discussed herein. This review will focus on mammalian cells, but similar trafficking occurs in all eukaryotic cells.

Membrane Composition Varies Among Organelles

Each type of organelle maintains a distinct protein composition, which allows the organelle to carry out its specific functions. There has been considerable study of the mechanisms for trafficking membrane proteins to specific organelles, and this selective trafficking is often based on peptide sequences in the cytoplasmic domains of proteins that are recognized by other proteins that form part of the scaffolds for budding transport vesicles or tubules from a parent organelle (Palade 1975; Bonifacino and Traub 2003). Lipid compositions also vary significantly among organelles (van Meer et al. 2008), but as discussed in a later section, the mechanisms for sorting lipids are not as well understood as for proteins. Some of the key differences in lipid composition are illustrated in Fig. 1. The plasma membrane is relatively enriched in sphingomyelin (SM), which is on the extracellular leaflet of the bilayer, and cholesterol (~30% of the lipid molecules). Phosphatidylserine (PS) is found on many organelles; on the plasma membrane, it is nearly exclusively on the cytoplasmic leaflet. External PS is found on apoptotic cells, and it is recognized by receptors on macrophages that engulf the dying cells (Savill and Fadok 2000). Phosphatidylinositol (PI), the precursor of phosphoinositides, is synthesized primarily in the endoplasmic reticulum (ER) and typically represents less than 15% of the total phospholipids found in eukaryotic cells. Phosphatidylinositol-4-phosphate (PI4P) and phosphatidylinositol-(4,5)-bisphosphate [PI(4,5)P₂] make up the bulk of phosphoinositides in mammalian cells, with PI(4,5)P₂ constituting ~1% of the phospholipids in the inner leaflet of the plasma membrane (Di Paolo and De Camilli 2006).

Lipid Trafficking in Cells, Fig. 1

Lipid distribution and movement among intracellular organelles. Phosphoinositides and special lipids involved in signaling and organelle recognition pathways are shown in ovals. PC, PE, PS, PI, and PA are synthesized in the ER and leave mainly by vesicular transport. SM and glycosphingolipids are synthesized in the Golgi and also move by vesicular transport. Each organelle is color-coded to reflect the relative level of cholesterol in its membranes. (The cholesterol levels for the limiting membrane of the late endosomes and lysosomes are uncertain because of the cholesterol associated with lipoproteins and internal membranes that are enriched in bis-monoacylglycerophosphate (BMP).) Circulating low-density lipoprotein (LDL) particles carrying cholesterol and cholesteryl ester are internalized through the LDL receptors and transported to sorting endosomes. Free cholesterol, which is released upon cholesteryl ester hydrolysis in late endosomes and lysosomes, requires proteins NPC1 and NPC2 to leave these organelles. It then moves to other organelles by poorly characterized mechanisms. Excess free cholesterol is esterified, and fatty acid sterol esters are packed into lipid droplets (LD). Pathways of intracellular sterol transport, both vesicular and nonvesicular, are shown. Phosphoinositides serve as organelle identity molecules: PI3P, phosphatidylinositol-3-phosphate; PI4P, phosphatidylinositol-4-phosphate; PI(3,5)P₂, phosphatidylinositol-(3,5)-bisphosphate; PI(4,5)P₂, phosphatidylinositol-(4,5)-bisphosphate; and PI(3,4,5)P₃, phosphatidylinositol-(3,4,5)-trisphosphate. ERC, endocytic-recycling compartment

Many lipids are synthesized in the ER and then must be transported to other cellular organelles. The ER contains lipids with a much higher level of unsaturation than the plasma membrane, and it maintains a low level of cholesterol (~5% of lipid molecules). The ER is unusual in that it does not contain a significant transbilayer asymmetry for most of its lipids, indicating that there are mechanisms for flipping lipids across this membrane relatively rapidly (Sanyal and Menon 2009).

The lipid composition shifts along the secretory pathway, as sterols and sphingolipids increase in concentration from the ER through the Golgi to the plasma membrane. For example, trans-Golgi network (TGN)-derived secretory vesicles in yeast contain 2.3-fold more ergosterol and sphingolipids than the TGN (Klemm et al. 2009). The overall lipid composition of early endosomes is similar to that found in the plasma membrane, with some differences in the fatty acid structure. Cholesterol and SM, as well as PS, are highly enriched in the recycling endosome. Late endosomes contain large amounts of neutral lipids such as triglycerides and cholesteryl ester, which are in the core of lipoproteins that are being digested. The membrane of late endosomes has a complex structure, with inward budding invaginations and detached internal vesicles. Bis-monoacylglycerophosphate (BMP, 15% of total lipids) is selectively enriched in the internal membranes of late endosomes (Gruenberg 2001; Kobayashi et al. 2001). Glycerolipids and sphingolipids are digested on these internal membranes to avoid damage to the limiting membrane (Kolter and Sandhoff 2010).

The lipid composition in the mitochondria varies little among different cell types, with phosphatidylcholine (PC) and phosphatidylethanolamine (PE) being the most abundant phospholipids (40% and 30% of total mitochondrial phospholipids, respectively), followed by glycerophospholipid cardiolipin (which is only found in mitochondria) and PI (10–15% of phospholipids), and phosphatidic acid (PA) and PS (5% of total mitochondrial phospholipids). Sphingolipids and sterols are not present in large amounts in mitochondrial membranes, although cholesterol, the precursor for steroid hormones, is delivered to mitochondria in steroidogenic cells (Osman et al. 2011).

Basic Mechanisms of Lipid Transport

There are two major methods for transport of lipids between organelles; they can move as part of a vesicle or tubule, or they can be transported by nonvesicular processes through the cytoplasm. Since lipids are poorly water soluble, nonvesicular transport is mediated by carrier proteins that can carry lipids between membranes. A specialized type of nonvesicular transport involves exchange of lipids between organelles at sites of close apposition (Fig. 1). Each of these transport mechanisms will be described briefly.

Vesicular Sorting

There is good evidence for sorting of lipids as vesicles and tubules bud off from a parent organelle. For example, fluorescent lipid analogs follow different endocytic routes after internalization from the plasma membrane depending upon the degree of unsaturation and the length of their hydrocarbon tails (Mukherjee et al. 1999). Lipid-anchored proteins, which are associated with membranes via a glycosyl-phosphatidylinositol (GPI) anchor, follow different intracellular itineraries than most other lipid molecules (Mayor and Riezman 2004). Additionally, toxins (e.g., shiga or cholera toxins) bound to glycolipids follow distinctive pathways after internalization that allow the toxins to reach the ER, where they are able to use the cell’s protein translocation machinery to cross into the cytoplasm (Sandvig et al. 2004).

The molecular basis for the sorting of lipids during membrane-trafficking processes is not well understood. In several cases, e.g., sorting of fluorescent lipid analogs and GPI-anchored proteins, changing cholesterol levels in cells alters the trafficking, indicating that membrane biophysical properties that are affected by cholesterol content are part of the sorting mechanism. One suggestion is that coexisting membrane microdomains are present in the parent organelle and that one type of microdomain might be preferentially incorporated into the budding vesicles or tubules. For example, if more ordered microdomains (sometimes called rafts) are enriched in a subset of proteins that are recruited by cytoplasmic coats on the budding membranes, then order-inducing lipids such as cholesterol and lipids with mostly saturated acyl chains would be selectively enriched in the budding transport vesicles (Fig. 2a). This mechanism has been suggested for lipid sorting in the biosynthetic secretory pathway (Simons and Ikonen 1997).

Lipid Trafficking in Cells, Fig. 2

Lipid sorting by vesicular mechanisms. This figure illustrates two putative mechanisms for sorting lipids during the vesicle-budding process in membrane transport. (a) Proteins associated with a particular type of microdomain, in this illustration cholesterol-enriched ordered lipid domains (pink), may also associate with coat proteins that form a budding vesicle. This would enrich the new vesicle in the types of lipids in the microdomain. (b) The shape of lipids can affect their ability to be accommodated in regions of high curvature present at sites of vesicle budding. Cone-shaped lipids (red) might be accommodated in the cytoplasmic leaflet of the neck region of a vesicle bud, whereas inverted cone lipids (green) might be better accommodated in the lumenal leaflet of the neck region. Different curvature (and the associated lipid preferences) are found in the more distal parts of the budding vesicle or tubule

Another possibility is that curvature preferences play an important role in lipid sorting. Sites of vesicle and tubule budding often involve regions of high membrane curvature. Lipid molecules can be nearly cylindrical in their overall shape (i.e., the same radius at the headgroup and at the end of the acyl chain), which would lead to a preference for planar bilayers. Cone-shaped lipids (with small headgroup area) or inverted cone lipids (with large headgroups compared to the acyl chain area) will have preferences to be in curved membranes (Fig. 2b). While the preference of individual lipid molecules for curved regions does not impose a strong selection, curvature can induce phase separation in lipid mixtures in which the composition is close to a phase separation boundary (Maxfield and van Meer 2010). Thus, curvature preferences and formation of microdomains may work synergistically to promote lipid sorting during vesicle formation.

A third mechanism for lipid sorting would involve specific binding of lipid species to proteins that are incorporated into (or excluded from) forming vesicles. There is little evidence that such a mechanism plays a general role in lipid sorting. However, there is mounting evidence that lipid-modifying enzymes are incorporated into forming vesicles. In particular, phosphatidylinositol kinases and phosphatases alter the phosphoinositide properties of the newly formed vesicles so that they no longer match the phosphoinositide signature of the donor membrane and begin to take on the characteristics of the target membrane with which they will fuse (Di Paolo and De Camilli 2006).

Nonvesicular Transport

Lipid molecules are held in membranes by noncovalent interactions and by the free energy associated with sequestering the hydrocarbon tails away from water. However, lipids in biological membranes are very dynamic, and they can have motions that transiently remove them partially from the bilayer, but complete spontaneous desorption is energetically unfavorable. It requires ~50 kJ/mol to remove cholesterol from membranes (Zhang et al. 2008) and ~80 kJ/mol to remove phospholipids (Marrink et al. 2009). Because of their poor solubility in water, lipid molecules, including cholesterol, must be associated with carrier proteins for efficient nonvesicular transport between organelles. Several such transport proteins have been identified. Members of the steroidogenic acute regulatory (StAR) protein-related lipid transfer (START) domain family of proteins have been shown to bind and transport cholesterol and some other lipids. The founding member of this family, StARD1, transports cholesterol to the inner membrane of mitochondria, where it can be converted to steroid hormones in steroidogenic tissues (Strauss et al. 2003; Miller and Auchus 2011). Other members of the START family have been shown to bind cholesterol (Alpy and Tomasetto 2005; Lavigne et al. 2010) and may play an important role in transport. Another member of this family, ceramide transfer protein (CERT), binds and transports ceramide (Hanada et al. 2009).

The oxysterol-binding protein (OSBP) was identified as a protein that binds 25-hydroxycholesterol. A related family of proteins (Osh proteins) contains OSBP homology domains, and many of these can bind sterols and transport them between membranes in vitro. The significance of this transport in cells is not known (Fairn and McMaster 2008). A family of PI/PC-transfer proteins (PITPs) can transfer lipids between membranes, but their main function is to coordinate lipid traffic and lipid metabolism (Mousley et al. 2010).

With their lower free energy for desorption from the membrane, sterols are a good candidate for nonvesicular transport, and there is evidence for substantial nonvesicular sterol transport in yeast and in mammalian cells. For example, sterol in the endocytic recycling compartment, which contains about 35–40% of the cholesterol in a cultured fibroblast line, exchanges with other sterol pools in the cell with a t_1/2 of about 2–3 min (Hao et al. 2002). This suggests that nonvesicular transport is the largest component of intracellular sterol transport. Similar conclusions were reached by studying the transport of newly synthesized cholesterol from the ER to the plasma membrane (Maxfield and Menon 2006). In yeast and in mammalian cells, treatments that block vesicular transport did not greatly alter transport of sterol from the ER to the plasma membrane. Unfortunately, the identity of the major cholesterol carriers has not been elucidated.

This rapid equilibration among organelles raises a question about how large differences in the sterol content of different organelle membranes is maintained (Mesmin and Maxfield 2009). Other membrane components of the various organelles can stabilize sterols to varying degrees, and this allows different concentrations of sterol to coexist in different organelles. For example, lipids with saturated acyl chains stabilize cholesterol in the membrane, and this type of lipid is enriched in the plasma membrane (high cholesterol) while unsaturated lipids predominate in the ER (low cholesterol).

The ER is the most important organelle for sterol regulation (Brown and Goldstein 2009). Cholesterol is synthesized in the ER. Esterification of cholesterol for storage in lipid droplets is the cellular high capacity, rapid response system for dealing with excess cholesterol, and acyl CoA cholesteryl acyltransferase, the enzyme that esterifies cellular cholesterol, resides in the ER. The complexes containing Insig, sterol regulatory element-binding protein (SREBP), and SREBP cleavage-activating protein (SCAP) also reside in the ER, and these proteins sense cholesterol levels and regulate the transcription of proteins involved in the uptake and synthesis of sterols. In high-cholesterol conditions, cholesterol binds to SCAP and Insig, leading to retention of SREBP in the ER. When cholesterol levels fall, SCAP and SREBP are recruited to transport vesicles and delivered to the Golgi apparatus. Two sequential proteolytic cleavages of SREBP in the Golgi lead to release of a cytoplasmic fragment of SREBP, which contains a transcriptional regulator that increases transcription of genes involved in cholesterol synthesis and the gene for the LDL receptor. In order for this system to work effectively, the ER must be able to sense the cholesterol levels in other organelles such as the plasma membrane. Since there is little vesicular transport from the plasma membrane to the ER, it is likely that nonvesicular sterol transport is an important component of this regulation, but the identity of the sterol carriers remains uncertain.

Endocytic Uptake of Lipoproteins

Nucleated cells can synthesize cholesterol and take it up by endocytosis of lipoproteins. Outside the central nervous system, LDLs are the main cholesterol carriers; the core of LDL is packed with cholesteryl esters, and the outer surface contains the protein (mainly apoB) and a monolayer of lipids and cholesterol. LDL binds to the LDL receptor and enters the cell by receptor-mediated endocytosis (Brown and Goldstein 1986). The LDL dissociates from its receptor in an acidified endosome, and the LDL receptor is recycled to the plasma membrane while the LDL is delivered to late endosomes and lysosomes and is degraded. Cholesteryl esters are hydrolyzed by lysosomal acid lipase. The unesterified cholesterol, which is very poorly soluble in water, is transported out of the lysosomes by a process that requires the proteins NPC1 and NPC2. It appears that cholesterol is first bound by NPC2, a soluble protein in the lumen of late endosomes. The cholesterol is then transferred to an N-terminal cholesterol-binding site on NPC1, a polytopic membrane protein in the late endosomes. It is unclear how NPC1 then facilitates the transport of cholesterol out of the late endosomes and lysosomes, but defects in either NPC1 or NPC2 cause Niemann-Pick disease type C, a severe lysosomal storage disorder in which cholesterol and other lipids accumulate in lysosomal storage organelles (Mesmin and Maxfield 2009).

Transport of Newly Synthesized Lipids

Most lipids are synthesized in the ER in mammalian cells, although significant lipid synthesis and remodeling also occurs in the Golgi apparatus and in mitochondria. An excellent review of transport of newly synthesized lipids has been presented recently (Blom et al. 2011). The predominant transport pathway for newly made phospholipids is by vesicular transport. (This must operate in such a way that the lipids with more saturated acyl chains are continually returned to the ER to maintain that organelle’s membrane characteristics.)

An important exception is ceramide, which can be transported by CERT, which has a ceramide-binding START domain. CERT also has a pleckstrin homology domain for targeting to Golgi membranes and a FFAT motif that binds the ER protein, VAP. Thus, CERT is able to efficiently shuttle ceramide from the cytoplasmic side of the ER, where it is made to the cytoplasmic side of the Golgi, where it can be converted to glucosylceramide. It is not entirely clear how the glucosylceramide is flipped to the lumenal leaflet of the biosynthetic organelles, which is essential to conversion to more complex glycolipids. One proposal (Halter et al. 2007) is that glucosylceramide is returned to the ER by nonvesicular transport on FAPP2 and that it is then flipped in the ER membrane and transported by vesicular transport to the Golgi. This somewhat baroque pathway may be used to regulate SM and glycolipid synthesis since the activity of the nonvesicular transport processes can modulate proteins that are involved in other aspects of lipid metabolism (D’Angelo et al. 2008).

Summary

Lipids move rapidly among cellular organelles by a combination of vesicular and nonvesicular transport processes. Despite all of this trafficking, organelles maintain very different lipid compositions. The maintenance of the unique lipid compositions of organelles requires sorting of lipids at many steps of vesicle formation. The underlying mechanisms for this lipid sorting are only partially understood. Some lipids (especially cholesterol) are transported by soluble carrier proteins among organelles.

Cross-References

Chemical Diversity of Lipids

Functional Roles of Lipids in Membranes

Glycerolipids: Chemistry

Glycosylphosphatidylinositol

Lipid Bilayer Asymmetry

Lipid Domains

Lipid Flip-Flop

Lipid Lateral Diffusion

Lipid Signaling and Phosphatidylinositols

Lipidomics

Sphingolipids and Gangliosides