Retrograde transport defects in Munc18-1 null neurons explain abnormal Golgi morphology

Abstract

Loss of the exocytic Sec1/MUNC18 protein MUNC18-1 or its t-SNARE partners SNAP25 and syntaxin-1 results in rapid, cell-autonomous and unexplained neurodegeneration, which is independent of their known role in synaptic vesicle exocytosis. cis-Golgi abnormalities are the earliest cellular phenotypes before degeneration occurs. Here, we investigated whether these Golgi abnormalities cause defects in the constitutive and regulated secretory pathway that may explain neurodegeneration. Electron microscopy confirmed that loss of MUNC18-1 expression results in a smaller cis-Golgi. In addition, we now show that medial-Golgi and the trans-Golgi Network are also affected. However, stacking and cisternae ultrastructure of the Golgi were normal. Overall ultrastructure of null mutant neurons was remarkably normal just hours before cell death occurred. Anterograde ER-to-Golgi and Golgi exit of endogenous and exogenous proteins were normal. In contrast, loss of MUNC18-1 caused reduced retrograde Cholera Toxin transport from the plasma membrane to the Golgi. In addition, MUNC18-1-deficiency resulted in abnormalities in retrograde TrkB trafficking. We conclude that MUNC18-1 deficient neurons have normal anterograde yet reduced retrograde transport to the Golgi. This imbalance in transport routes provides a plausible explanation for the observed Golgi abnormalities and cell death in MUNC18-1 deficient neurons. Significance statementLoss of MUNC18-1 or its t-SNAREs SNAP25 and syntaxin-1 leads to massive, yet unexplained, neurodegeneration. Previous research showed that Golgi abnormalities are the earliest, shared phenotype. Golgi abnormalities are also an early feature in neurodegenerative diseases, such as Alzheimers Disease or Amyotrophic Lateral Sclerosis. This study elucidates the mechanism underlying the Golgi phenotype upon loss of MUNC18-1. By systematically assessing transport routes to and from the Golgi, we show that retrograde endosome-to-Golgi, but not anterograde transport from the Golgi, is disturbed. This imbalance in transport routes provides a plausible explanation for the Golgi phenotype, and may explain the neurodegeneration. The findings in this study contributes to new insights in cellular mechanisms of neurodegeneration.