Transport of the Major Myelin Proteolipid Protein Is Directed by VAMP3 and VAMP7

Abstract

CNS myelination by oligodendrocytes requires directed transport of myelin membrane components and a timely and spatially controlled membrane expansion. In this study, we show the functional involvement of the R-soluble N-ethylmaleimide-sensitive factor attachment protein receptor (R-SNARE) proteins VAMP3/cellubrevin and VAMP7/TI-VAMP in myelin membrane trafficking. VAMP3 and VAMP7 colocalize with the major myelin proteolipid protein (PLP) in recycling endosomes and late endosomes/lysosomes, respectively. Interference with VAMP3 or VAMP7 function using small interfering RNA-mediated silencing and exogenous expression of dominantnegative proteins diminished transport of PLP to the oligodendroglial cell surface. In addition, the association of PLP with myelin-like membranes produced by oligodendrocytes cocultured with cortical neurons was reduced. We furthermore identified Syntaxin-4 and Syntaxin-3 as prime acceptor Q-SNAREs of VAMP3 and VAMP7, respectively. Analysis of VAMP3-deficient mice revealed no myelination defects. Interestingly, AP-3␦-deficient mocha mice, which suffer from impaired secretion of lysosome-related organelles and missorting of VAMP7, exhibit a mild dysmyelination characterized by reduced levels of select myelin proteins, including PLP. We conclude that PLP reaches the cell surface via at least two trafficking pathways with distinct regulations: (1) VAMP3 mediates fusion of recycling endosomederived vesicles with the oligodendroglial plasma membrane in the course of the secretory pathway; (2) VAMP7 controls exocytosis of PLP from late endosomal/lysosomal organelles as part of a transcytosis pathway. Our in vivo data suggest that exocytosis of lysosomerelated organelles controlled by VAMP7 contributes to myelin biogenesis by delivering cargo to the myelin membrane.

Transcriptional Expression of Myelin Basic Protein in Oligodendrocytes Depends on Functional Syntaxin 4: a Potential Correlation with Autocrine Signaling

“…However, quite unexpectedly, effective downregulation of syntaxin 4 did not result in significant downregulation of vesicular transport of the myelin-specific protein PLP, the viral model protein VSV G, or integrin ␣6, known to regulate MBP mRNA expression (54). Yet, we cannot exclude that in this particular case, sheet-directed transport, mediated by the SNARE machinery, might have exploited alternatives for syntaxin 4, e.g., syntaxin 2, which is present in mature OLGs (18,19), in a nonpolarized manner (our unpublished observations) or for the VAMP3 v-SNARE, e.g., VAMP2 (18,19) or VAMP7, which has been implicated in PLP trafficking (32). If so, it should also be emphasized that "t-SNARE substitution" does not apply to the apical machinery, driven by syntaxin 3, as in this case its downregulation did effectively preclude PLP trafficking to the plasma membrane of the cell body, which precedes subsequent transport to the myelin sheet (1,34,36,(63)(64)(65).…”

“…Interacting binding partners of syntaxin 4 in vesicular transport are v-SNAREs, known as VAMPs. In OLGs, the t-SNARE syntaxin 4 is recognized by VAMP3 (18,32). To assess, therefore, whether VAMP3 might be involved in facilitating the expression of MBP, we next examined the effect of VAMP3 downregulation on MBP expression.…”

“…Although MBP mRNA is assembled in nonmembranous granules (26)(27)(28), a causal relationship with the vesicular transport ma-chinery is likely, given that membrane trafficking appears to be involved in RNA transport and/or anchoring (29)(30)(31). In contrast, PLP is an integral membrane protein that is synthesized at the endoplasmic reticulum and subsequently processed by vesicular transport, reaching the myelin membrane via a transcytotic transport mechanism (1,(32)(33)(34)(35)(36). Furthermore, a distinct role for the v-SNAREs vesicle-associated membrane protein 7 (VAMP7) and VAMP3, cognate binding partners of syntaxins 3 and 4, respectively, in PLP trafficking has been recently demonstrated (32).…”

“…In contrast, PLP is an integral membrane protein that is synthesized at the endoplasmic reticulum and subsequently processed by vesicular transport, reaching the myelin membrane via a transcytotic transport mechanism (1,(32)(33)(34)(35)(36). Furthermore, a distinct role for the v-SNAREs vesicle-associated membrane protein 7 (VAMP7) and VAMP3, cognate binding partners of syntaxins 3 and 4, respectively, in PLP trafficking has been recently demonstrated (32). PLP plays a major role in assembly and stabilization of the myelin sheath in that the protein brings about the correct apposition of the extracellular leaflets of the membrane (37,38).…”

Regulation of Oligodendrocyte Differentiation: Relevance for Remyelination

“…For PLP there is good evidence that the newly synthesized protein is first transported to the plasma membrane of the cell soma. From there it is internalized and stored in late endosomes before it is finally transported by a transcytotic pathway to the myelin sheath (Trajkovic et al, 2006;Feldmann et al, 2011). Surprisingly, although PLP deletion results in neuronal degeneration, it has only minor effects on myelin formation (Garbern et al 2002).…”