Mitochondria-Endoplasmic Reticulum Interplay Regulates Exo-Cytosis in Human Neuroblastoma Cells.

Giacomo Dentoni,Luana Naia,Maria Ankarcrona

doi:10.3390/cells11030514

Abstract

Mitochondria–endoplasmic reticulum (ER) contact sites (MERCS) have been emerging as a multifaceted subcellular region of the cell which affects several physiological and pathological mechanisms. A thus far underexplored aspect of MERCS is their contribution to exocytosis. Here, we set out to understand the role of these contacts in exocytosis and find potential mechanisms linking these structures to vesicle release in human neuroblastoma SH-SY5Y cells. We show that increased mitochondria to ER juxtaposition through Mitofusin 2 (Mfn2) knock-down resulted in a substantial upregulation of the number of MERCS, confirming the role of Mfn2 as a negative regulator of these structures. Furthermore, we report that both vesicle numbers and vesicle protein levels were decreased, while a considerable upregulation in exocytotic events upon cellular depolarization was detected. Interestingly, in Mfn2 knock-down cells, the inhibition of the inositol 1,4,5-trisphosphate receptor (IP3R) and the mitochondrial calcium (Ca2+) uniporter (MCU) restored vesicle protein content and attenuated exocytosis. We thus suggest that MERCS could be targeted to prevent increased exocytosis in conditions in which ER to mitochondria proximity is upregulated.

Highlights

Mitochondria are highly dependent on other organelles and pivotal is their interaction with the endoplasmic reticulum (ER)
To assess whether mitochondria-ER contact sites (MERCS) could affect vesicle dynamics in SH-SH5Y cells, we decided to knock-down Mitofusin 2 (Mfn2) as a well-established method to modulate the juxtaposition between mitochondria and ER
We found that the levels of several vesicular proteins and neuropeptides namely synaptophysin, synapsin-1, neuropeptide Y were significantly lower in Mfn2 knock-down (Mfn2 KD) cells compared to Negative Control (NC) (Figure 2B,C; p < 0.05), while no significant differences were found in synaptotagmin-1 and syntaxin-1 levels (Figure 2B,C)

Summary

Introduction

Mitochondria are highly dependent on other organelles and pivotal is their interaction with the endoplasmic reticulum (ER). Five to 20% of the mitochondrial surface is believed to be in contact with the ER [1], and the lipid-raft domain on the ER juxtaposed to mitochondria is referred to as the mitochondria-associated ER membrane (MAM) [2]. Mfn knock-out MEF cells studies showed decreased interaction between mitochondria and ERmembranes, as well as decreased Ca2+-shuttling between ER and mitochondria [12]. These studies were corroborated by observations in cell lines [14,15], brain [16] and neurons [17]

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Conclusion