Die Bedeutung der Intramembranproteasen SPPL2a/b für die Homöostase von C-terminal verankerten Membranproteinen
Biochemie
Zusammenfassung der Projektergebnisse
Tail-anchored (TA) proteins are a topologically defined class of membrane proteins that include the large and functionally critical group of SNARE proteins involved in vesicular traffic by facilitating membrane fusion. Mechanisms controlling the degradation of SNARE proteins are currently poorly understood. We have systematically analysed if two intramembrane proteases Signal Peptide Peptidase-like 2a and b (SPPL2a/b) contribute to the homeostasis of TA proteins in the late secretory and endosomal/lysosomal pathway. As their transmembrane segment displays type II topology and their C-terminus is short, TA proteins fulfill all basic requirements to be substrates of these proteases. SPPL2a and SPPL2b are aspartyl intramembrane proteases with homology to presenilins that primarily reside in lysosomes/late endosomes and at the plasma membrane, respectively. We have systematically screened a wide spectrum of SNARE proteins and other TA proteins in cellular substrate-protease co-expression assays and identified cleavage of the Vesicle associated membrane proteins VAMP1-4 by SPPL2a/b. Cleavage of these four SNARE proteins was scrutinised at the endogenous level in different cell lines as well as by analysing VAMP1-4 levels in tissues and primary cells of SPPL2a/b double-deficient (dKO) mice. Loss of SPPL2a/b activity resulted in an accumulation of VAMP1-4 in a cell type- and tissue-dependent manner clearly identifying these proteins as SPPL2a/b substrates validated in vivo highlighting turnover by these intramembrane proteases. We could demonstrate that a dysregulation of SPPL2a/b in certain pathophysiological contexts can directly impact on cellular levels of VAMP2. We took major efforts to analyse how modulation of SNAREs by SPPL2a/b may impact on exocytotic processes. As a model process, we analysed the plasma membrane translocation of the insulin-sensitive transporter Glut4, for which a major role of VAMP2 has been established in the literature. In parallel, we had observed that SPPL2a/bdeficient (dKO) mice exhibit a metabolic phenotype characterised by an enhanced glucose tolerance which would be consistent with an augmented and/or accelerated Glut4 translocation. Experiments in cellular model systems of Glut4 translocation did not readily confirm this hypothesis. However, these models exhibit several major limitations and were therefore also not capable of formally disproving the described concept, which will followedup using in vivo approaches. Altogether, we have systematically characterised a novel biochemical mechanism for the turnover of SNARE proteins. However, more work will be needed to define its regulatory impact on membrane trafficking processes in a cell typedependent manner and to underpin a potential connection between the metabolic phenotype of SPPL2a/b dKO mice and their altered SNARE homeostasis.
Projektbezogene Publikationen (Auswahl)
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The intramembrane proteases SPPL2a and SPPL2b regulate the homeostasis of selected SNARE proteins. The FEBS Journal, 290(9), 2320-2337.
Ballin, Moritz; Griep, Wolfram; Patel, Mehul; Karl, Martin; Mentrup, Torben; Rivera‐Monroy, Jhon; Foo, Brian; Schwappach, Blanche & Schröder, Bernd
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The role of SPP/SPPL intramembrane proteases in membrane protein homeostasis. The FEBS Journal, 291(1), 25-44.
Mentrup, Torben; Leinung, Nadja; Patel, Mehul; Fluhrer, Regina & Schröder, Bernd
