Diversity in unconventional protein secretion

Abstract

Eukaryotic cells use the endoplasmic reticulum (ER)-to-Golgi membrane pathway for the secretion of the vast majority of secretory proteins. This process is initiated by signal-peptide-dependent protein translocation into the lumen of the ER followed by vesicular transport of secretory cargo to the Golgi membranes and thence to the cell surface. However, a substantial number of proteins that lack a signal peptide are secreted without entering the conventional ER-to-Golgi pathway of protein secretion. These proteins are generally involved in cell survival, immune surveillance and tissue organization and, therefore, are of fundamental importance. Recent studies have uncovered the existence of four principal types of unconventional protein secretion that can be distinguished into non-vesicular and vesicular pathways (see Poster). The non-vesicular pathways encompass self-sustained protein translocation across plasma membranes (type I) and ABC-transporter-based secretion (type II). Vesicular pathways are characterized by autophagy-based secretion (type III) and proteins that bypass the Golgi complex (hereafter referred to as the Golgi bypass) for trafficking to the plasma membrane (type IV). Importantly, types I, II and III are pathways that mediate unconventional secretion of cytoplasmic proteins – that, in case of type II, are acylated. By contrast, all known examples of the type IV pathway are integral membrane proteins.The type I of unconventional secretion refers to direct translocation of cytoplasmic proteins across the plasma membrane without the involvement of vesicular intermediates (see Poster). The best studied example for this type of secretion is the release of fibroblast growth factor 2 (FGF2) (Nickel, 2011), which involves interaction of FGF2 with the phosphoinositide phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P2) at the inner leaflet and with heparan sulfates at the outer leaflet of the plasma membrane (Schäfer et al., 2004; Zehe et al., 2006; Temmerman et al., 2008). The mechanism by which FGF2 traverses the plasma membrane depends on a fully folded conformation of FGF2 (Backhaus et al., 2004; Torrado et al., 2009), a key finding that argues against the participation of a classic protein-conducting channel at the level of the plasma membrane. Recently, membrane translocation of FGF2 was revealed to involve PtdIns(4,5)P2-induced oligomerization at the inner membrane leaflet as well as insertion of FGF2 into the membrane (Steringer et al., 2012). This process is strongly stimulated by phosphorylation of FGF at tyrosine residue 82 through Tec kinase (Ebert et al., 2010). Reconstitution experiments – that were carried out to analyze transbilayer diffusion of membrane lipids by using model membranes and purified proteins – have established that PtdIns(4,5)P2-dependent oligomerization and membrane insertion of FGF2 is triggered by the formation of a lipidic membrane pore with a putative toroidal architecture (Steringer et al., 2012). We speculate this structure is a ring-like FGF2 hexamer that is located in the center of the pore (see Poster), and believe that it represents a transient translocation intermediate in the process of FGF2 secretion that is rapidly altered when FGF2 is extracellularly trapped by binding to heparan sulfate proteoglycans (see Poster).In conclusion, membrane translocation of FGF2 is mediated by a self-sustained mechanism that is driven by PtdIns(4,5)P2-dependent oligomerization and tyrosine phosphorylation of FGF2. This process generates an intermediate structure of FGF2 oligomers that open and stabilize a lipidic membrane pore without contact of the FGF2 oligomer to the inner hydrophobic core of the plasma membrane (see Poster) (Steringer et al., 2012). In this context, one may speculate that – thus far unidentified – membrane proteins exist that prevent a sustained breakdown of the membrane potential or a substantial loss of lipid asymmetry during membrane translocation of FGF2. Such a mechanism could explain how cells can tolerate a mechanism of translocation through the plasma membrane that involves transient disturbance of the regular lipid bilayer structure.These observations indicate a mechanism of unconventional secretion of FGF2 that is based on (i) recruitment by acidic membrane lipids at the inner leaflet, (ii) oligomerization-induced membrane insertion and (iii) tyrosine phosphorylation. This mechanism might be relevant for other unconventionally secreted proteins, such as HIV-1 Tat (Rayne et al., 2010a; Rayne et al., 2010b), annexin A2 (Anxa2) (Deora et al., 2004), and FGF1 (Prudovsky et al., 2008). Specifically, these proteins are known to bind to acidic membrane lipids, including phosphoinositides (Tarantini et al., 1995; Rescher et al., 2004; Rayne et al., 2010b). Further, they appear to be able to traverse the plasma membrane (Prudovsky et al., 2002; Deora et al., 2004; Rayne et al., 2010a; Rayne et al., 2010b; Kirov et al., 2012) and, in case of HIV