Feline Lindhout

110 4 ABSTRACT In neurons, the continuous and dynamic endoplasmic reticulum (ER) network extends throughout the axon, and its dysfunction causes various axonopathies. However, it remains largely unknown how ER integrity and remodeling modulate presynaptic function in mammalian neurons. Here, we demonstrated that ER membrane receptors VAPA and VAPB are involved in modulating the synaptic vesicle (SV) cycle. VAP interacts with secernin-1 (SCRN1) at the ER membrane via a single FFAT-like motif. Similar to VAP, loss of SCRN1 or SCRN1-AP interactions resulted in impaired SV cycling. Consistently, SCRN1 or VAP depletion was accompanied by decreased action potential-evoked Ca 2+ responses. Additionally, we found that VAP-SCRN1 interactions play an important role in maintaining ER continuity and dynamics, as well as presynaptic Ca 2+ homeostasis. Based on these findings, we propose a model where the ER-ocalized VAP-SCRN1 interactions provide a novel control mechanism to tune ER remodeling and thereby modulate Ca 2+ dynamics and SV cycling at presynaptic sites. These data provide new insights into the molecular mechanisms controlling ER structure and dynamics, and highlight the relevance of ER function for SV cycling. INTRODUCTION The continuous and dynamic ER network is one of the most abundant organelles in cells. In neurons, somatodendritic domains contain both rough and smooth ER, whereas axons exclusively exhibit smooth ER. The smooth ER lacks ribosomes and is not involved in translation; instead, it is important for Ca 2+ homeostasis, lipid synthesis and delivery, and signaling. The relevance of axonal ER in particular is highlighted by various human axonopathies caused by mutations in different generic ER proteins. More specifically, dysfunction of ER-shaping proteins such as atlastin-1, reticulon-2, receptor expression- enhancing protein 1 (REEP1), and receptor expression-enhancing protein 2 (REEP2) leads to hereditary spastic paraplegia (HSP), whereas mutations in ER receptor VAMP-associated protein B (VAPB) cause amyotrophic lateral sclerosis (ALS; Hazan et al, 1999; Zhao et al, 2001; Nishimura et al, 2004; Zuchner et al, 2006; Montenegro et al, 2012; Esteves et al, 2014; Yalcin et al, 2017). Together, these pathologies hint for an increased sensitivity for proper ER structure and function in axons. Recent ultrastructural three-dimensional analysis revealed that the ER in axons is comprised of a conserved and unique organization (Wu et al, 2017; Yalcin et al, 2017; Terasaki, 2018). The axonal ER structure consists of narrow ER tubules, which occasionally form cisternae at tubular branch points with comparably small lumen (Wu et al, 2017; Yalcin et al, 2017; Terasaki, 2018). This distinctive ER network extends throughout all axon branches with a relative constant density of only 1–2 narrow tubules per diameter, while remaining continuous with the rest of the ER network (Wu et al, 2017; Yalcin et al, 2017; Terasaki, 2018). At presynaptic terminals, the ER forms a local tubular network opposing the active zone. This presynaptic ER structure often wraps around mitochondria and is in close proximity to the