8/19/2023 0 Comments Ipicture regina vossSecretion is generally believed to be due to proteolytic cleavage within the so-called stem region that tethers the catalytic glycosyltransferase ectodomain to its membrane anchor (Paulson & Colley, 1989 Ohtsubo & Marth, 2006 Varki et al, 2009). Secretion of the catalytically active glycosyltransferase domain has been discussed to negatively regulate cellular protein glycosylation, since the crucially required nucleotide- or lipid-linked sugar donor substrates that exclusively occur intracellularly are not available for secreted glycosyltransferases (Paulson & Colley, 1989 Ohtsubo & Marth, 2006 Varki et al, 2009). Secreted glycosyltransferases were observed in conditioned media of cultured cells (Elhammer & Kornfeld, 1986 Saito et al, 2002 El-Battari et al, 2003), but also in bodily fluids (Kim et al, 1972a, b Elhammer & Kornfeld, 1986 Kitazume et al, 2009) and tissues (Weinstein et al, 1987). Their large C-terminal ectodomain that harbours the glycosyltransferase activity faces the lumen of the Golgi (Paulson & Colley, 1989 Sears & Wong, 1998 Varki et al, 2009). Most glycosyltransferases are single-span type II transmembrane proteins with a short N-terminal cytoplasmic tail. Within the medial- and trans-Golgi network, numerous glycosyltransferase and glycosidases compete for these high-mannose-type precursor glycans converting them into higher-order, complex N-glycans (Sears & Wong, 1998 Moremen et al, 2012). In eukaryotic organisms, N-glycan synthesis is initiated in the ER, resulting in high-mannose-type glycans attached to the luminal domain of secretory and membrane proteins (Kornfeld & Kornfeld, 1985). Recently, we described the first SPP元 substrate in a cell culture model system, confirming the assumption that SPP元 indeed is proteolytically active (Voss et al, 2012). SPP元 most likely adopts the nine TMD topology conserved among GxGD proteases (Friedmann et al, 2004), localises to the Golgi network (Friedmann et al, 2006) and is not glycosylated (Friedmann et al, 2004). Highly conserved orthologues of mammalian SPP元, however, are found in most multicellular eukaryotes pointing to a fundamental cellular function of SPP元 (Voss et al, 2013). While recent studies on SPP, SPPL2a and SPPL2b identified substrates of these proteases in vitro and in vivo and consequently gave a first impression of the physiological function of these family members (reviewed in Voss et al, 2013), the physiological function of SPP元 has remained completely enigmatic. In contrast to presenilins, which exclusively accept type I transmembrane substrates (Kopan & Ilagan, 2004), SPP/SPPLs seem to be selective towards transmembrane substrates in type II orientation (Weihofen et al, 2002 Friedmann et al, 2004 Nyborg et al, 2004). Mutagenesis of either aspartate residue inactivates the respective SPP/SPPL (Weihofen et al, 2002 Fluhrer et al, 2006 Friedmann et al, 2006 Kirkin et al, 2007 Voss et al, 2012). The two catalytic aspartate residues required for the proteolytic activity of SPP/SPPL proteases are embedded in conserved YD and GxGD amino acid motifs in transmembrane domain (TMD) 6 and TMD7, respectively (Voss et al, 2013). Notably, all members of the SPP/SPPL family share characteristic structural features and catalytic motifs with the presenilins, the catalytically active subunits of the γ-secretase complex (Voss et al, 2013). Together with its mammalian paralogues, signal peptide peptidase (SPP) and the signal peptide peptidase-like (SPPL) proteases SPPL2a, SPPL2b and SPPL2c, it was first identified by database queries in 2002 (Grigorenko et al, 2002 Ponting et al, 2002 Weihofen et al, 2002). Signal peptide peptidase-like 3 (SPP元) is a multi-pass transmembrane protein that is highly conserved among multicellular eukaryotes and belongs to the family of intramembrane-cleaving GxGD proteases (Voss et al, 2013). Thus, SPP元 plays a central role in an evolutionary highly conserved post-translational process in eukaryotes. In line with that, reduced expression of SPP元 results in a hyperglycosylation phenotype, whereas elevated SPP元 expression causes hypoglycosylation. Cleavage of these enzymes leads to a reduction in their cellular activity. We demonstrate that SPP元 alters the pattern of cellular N-glycosylation by triggering the proteolytic release of active site-containing ectodomains of glycosidases and glycosyltransferases such as N-acetylglucosaminyltransferase V, β-1,3 N-acetylglucosaminyltransferase 1 and β-1,4 galactosyltransferase 1. Its physiological function, however, has remained enigmatic, since presently no physiological substrates have been identified. Signal peptide peptidase-like 3 (SPP元) is an intramembrane-cleaving aspartyl protease of the GxGD type. Protein N-glycosylation is involved in a variety of physiological and pathophysiological processes such as autoimmunity, tumour progression and metastasis.
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