ER-PM connections: sites of information transfer and inter-organelle communication

Abstract

Eukaryotic cells are divided into distinct membrane-bound organelles with unique identities and specialized metabolic functions. Communication between organelles must take place to regulate the size, shape, and composition of individual organelles, as well as to coordinate transport between organelles. The endoplasmic reticulum (ER) forms an expansive membrane network that contacts and participates in crosstalk with several other organelles in the cell, most notably the plasma membrane (PM). ER-PM junctions have well-established functions in the movement of small molecules, such as lipids and ions, between the ER and PM. Recent discoveries have revealed additional exciting roles for ER-PM junctions in the regulation of cell signaling, ER shape and architecture, and PM domain organization.

Figure 1

Membrane contact sites between the endoplasmic reticulum (ER) and plasma membrane (PM) are conserved cellular structures. (a) The ER consists of a continuous membrane meshwork throughout the cell. Peripheral ER membranes form close associations with the PM without undergoing membrane fusion. These membrane contact sites allow for direct ER–PM crosstalk independently of membrane trafficking through the secretory and endocytic pathways. (b) In yeast cells, the PM is extensively associated with cortical ER (cER) membranes, as revealed by electron microscopy [12^•], © West et al., 2011. Originally published in J Cell Biol, 193:333-346. The cortical ER (cER) membrane (black arrow) and PM (white arrow) are in close apposition, approximately 30 nm apart. The lumen of the cER compartment is shaded in blue. Notably, electron-dense ribosomes in the cytoplasm are excluded from the ER–PM contact zone. The cell wall (CW) and scale bar (100 nm) are indicated. ER–PM contact sites share conserved features and also display unique properties in different cell types. (c) Electron microscopy of a mammalian Hela cell expressing the STIM proteins [14], © Orci et al. and the National Academy of Sciences, 2009. Originally published in Proc Natl Acad Sci USA 2009, 106:19358-19362. Similar to yeast cells, the cortical ER (cER, asterisk) is depleted of ribosomes as compared to internal ER membranes. Notably, the PM-associated cER and internal ER membranes display additional distinctions. The cER structures associated with the PM appear more flattened as compared to internal ER. Remarkably, the distance between the opposing membranes at ER–PM contacts can narrow to within 10 nm. An example of a tight membrane contact site (MCS) enriched in electron-dense material is shown (arrow). The scale bar indicates 200 nm.

Figure 2

ER–PM membrane contact sites regulate calcium (Ca²⁺) dynamics. Associations between the ER and other organelles, such as the PM and mitochondria (m), were first observed by Keith Porter and George Palade in electron microscopy experiments on muscle cells [5]. (a) An illustration of cardiac muscle from this study is shown, © Rockefeller University Press, 1957. Originally published in J Biophys Biochem Cytol, 3:269-300. Large invaginations of the sarcolemma (the PM), termed transverse tubules (T-tubules), were found to be closely associated with membranes of the sarcoplasmic reticulum (SR, the ER in muscle cells). These structures were termed triads (a T-tubule closely apposed by two terminal SR compartments) and were later shown to be involved in Ca²⁺-mediated and cytoskeletal-mediated muscle contraction, known as excitation–coupled contraction. (b) Molecular components responsible for excitation-coupled contraction have been identified. At ER–PM contacts, voltage-gated Ca²⁺ channels (VGCC) localized in T-tubules interact with and activate ryanodine receptors (RyR) in the ER [6]. Upon activation, the RyR releases Ca²⁺ stores from the ER lumen to generate Ca²⁺ signals in the cytoplasm necessary for cytoskeletal-mediated muscle cell contraction. The protein junctophilin (JP) is involved in ER–PM contact formation in muscle cells [15]. Junctophilin is an integral ER membrane protein with several cytoplasmic MORN repeats proposed to bind PIP lipids in the PM. (c) Following release, luminal ER Ca²⁺ stores must be replenished for normal ER function (e.g. lipid synthesis, protein folding and secretion). In many cell types, the STIM proteins function in the store-operated Ca²⁺ entry (SOCE) pathway to maintain Ca²⁺ homeostasis in the ER lumen [4]. The ER-localized STIM protein senses Ca²⁺ levels in the ER lumen via a Ca²⁺-binding EF hand domain. Upon depletion of Ca²⁺ in the ER lumen, STIM proteins oligomerize via cytoplasmic coiled-coil domains (CC) and translocate to ER–PM contact sites. A polybasic region rich in lysine residues (K) within the cytoplasmic tail of STIM binds to PIP lipids in the PM. The STIM proteins subsequently interact with and recruit PM-localized Orai1 Ca²⁺ channels (also known as Ca²⁺ release-activated channels, CRAC) to ER–PM junctions. The CRAC-activation domain (CAD) within the STIM proteins directly binds and activates Orai1 channels, resulting in Ca²⁺ influx necessary for cytoplasmic Ca²⁺ signals and for reloading Ca²⁺ stores into the ER lumen through SERCA (smooth ER Ca²⁺ ATPase) transporters.

Figure 3

Essential cellular processes occur at ER–PM contact sites. (a) Non-vesicular sterol lipid transfer is facilitated at ER–PM junctions. Oxysterol-binding protein related proteins (ORPs) are recruited to ER–PM junctions by protein–protein and protein–lipid interactions [28]. ORP family members contain a FFAT motif that binds integral ER membrane proteins termed VAPs and a PH domain that binds the PIP isoform PI4P in the PM [29,30]. The conserved ORP sterol-binding domain extracts newly synthesized sterol lipids (magenta) in the ER. Upon subsequent delivery of sterol to the PM, interactions between the mouth of the ORP sterol-binding domain and PI4P may inhibit extraction of sterol from the PM [32^•]. (b) ER–PM contacts are also sites for regulation of PI4P levels in the PM [39^•,50^••]. The ER-localized PIP phosphatase Sac1 is activated by ORPs family members at ER–PM contacts, resulting in PI4P turnover at the PM. This process is facilitated by VAPs in the ER. (c) In addition to lipid transfer and metabolism, growth factor receptor signaling is regulated at ER–PM junctions. The ER-anchored protein tyrosine phosphatase isoform PTP1B interacts with and dephosphorylates ligand-bound receptor tyrosine kinases (RTKs) at ER–PM contacts. Dephosphorylation of active RTKs terminates signaling events at the PM and promotes RTK endocytosis and down-regulation [42,43,44^•,58].

Figure 4

Three conserved protein families are involved in ER–PM contact site formation and function. The tricalbin proteins (Tcb, orthologs of the extended synaptotagmin-like proteins), VAP family members, and Ist2 (related to the TMEM16 channel family) tether the ER to the PM in yeast cells [49^•,50^••,51^••,52^•]. (a) Electron microscopy of wild type and Δtether yeast cells [50^••]. Originally published in Dev Cell, 2012, 23: 1129–1140. In wild type cells, large regions of the PM are associated with cortical ER (purple). In cells lacking the ER–PM tether proteins (Δtether), cortical ER structures are depleted, and a meshwork of cytoplasmic ER accumulates (green). The nuclear ER (blue), mitochondria (M), nucleus (N), cell wall (CW) and vacuole (V) are also labeled. Scale bars, 500 nm. (b) All of the ER–PM tether proteins are integral ER membrane proteins that contain conserved cytoplasmic lipid-binding and protein-binding domains. The tricalbin proteins possess C2 domains that display Ca²⁺-stimulated lipid binding activity and a conserved SMP domain that is sufficient for lipid binding and ER targeting. The Ist2 protein contains a carboxyl terminal polybasic domain (PB) proposed to bind PIP lipids at the PM. The lipid binding activities in the tether proteins may facilitate the transmission of signals between the PM and ER. The VAP proteins recruit lipid-binding ORP family members to ER–PM contact sites; additional interactions with PM proteins and lipids may be involved in VAP tethering function. (c) ER–PM contact sites serve key roles in Ca²⁺ dynamics, sterol and phosphoinositide lipid signaling, growth factor receptor signaling, and membrane trafficking pathways. In addition to specific signaling roles, the ER–PM tether proteins may have general housekeeping functions in several of these essential cellular processes.