A proline to serine substitution at position 56 in the gene encoding vesicle-associated membrane protein-associated protein B (VAPB) causes some dominantly inherited familial forms of motor neuron disease including amyotrophic lateral sclerosis (ALS) type-8. VAPB is an integral endoplasmic reticulum (ER) protein whose amino-terminus projects into the cytosol. Overexpression of ALS mutant VAPBP56S disrupts ER structure but the mechanisms by which it induces disease are not properly understood. Here we show that VAPB interacts with the outer mitochondrial membrane protein, protein tyrosine phosphatase-interacting protein 51 (PTPIP51). ER and mitochondria are both stores for intracellular calcium (Ca2+) and Ca2+ exchange between these organelles occurs at regions of ER that are closely apposed to mitochondria. These are termed mitochondria-associated membranes (MAM). We demonstrate that VAPB is a MAM protein and that loss of either VAPB or PTPIP51 perturbs uptake of Ca2+ by mitochondria following release from ER stores. Finally, we demonstrate that VAPBP56S has altered binding to PTPIP51 and increases Ca2+ uptake by mitochondria following release from ER stores. Damage to ER, mitochondria and Ca2+ homeostasis are all seen in ALS and we discuss the implications of our findings in this context.

Signaling between the endoplasmic reticulum (ER) and mitochondria regulates a number of key neuronal functions. This signaling involves close physical contacts between the two organelles that are mediated by “tethering proteins” that function to recruit regions of ER to the mitochondrial surface. The ER protein, vesicle-associated membrane protein-associated protein B (VAPB) and the mitochondrial membrane protein, protein tyrosine phosphatase interacting protein-51 (PTPIP51), interact to form one such tether. Recently, damage to ER-mitochondria signaling involving disruption of the VAPB-PTPIP51 tethers has been linked to the pathogenic process in Parkinson’s disease, fronto-temporal dementia (FTD) and related amyotrophic lateral sclerosis (ALS). Loss of neuronal synaptic function is a key feature of Parkinson’s disease and FTD/ALS but the roles that ER-mitochondria signaling and the VAPB-PTPIP51 tethers play in synaptic function are not known. Here, we demonstrate that the VAPB-PTPIP51 tethers regulate synaptic activity. VAPB and PTPIP51 localise and form contacts at synapses, and stimulating neuronal activity increases ER-mitochondria contacts and the VAPB-PTPIP51 interaction. Moreover, siRNA loss of VAPB or PTPIP51 perturbs synaptic function and dendritic spine morphology. Our results reveal a new role for the VAPB-PTPIP51 tethers in neurons and suggest that damage to ER-mitochondria signaling contributes to synaptic dysfunction in Parkinson’s disease and FTD/ALS. Electronic supplementary material The online version of this article (10.1186/s40478-019-0688-4) contains supplementary material, which is available to authorized users.

A proline-to-serine substitution at position 56 in the gene encoding vesicle-associated membrane protein-associated protein B (VAPB; VAPBP56S) causes some dominantly inherited familial forms of motor neuron disease, including amyotrophic lateral sclerosis (ALS) type-8. Here, we show that expression of ALS mutant VAPBP56S but not wild-type VAPB in neurons selectively disrupts anterograde axonal transport of mitochondria. VAPBP56S-induced disruption of mitochondrial transport involved reductions in the frequency, velocity and persistence of anterograde mitochondrial movement. Anterograde axonal transport of mitochondria is mediated by the microtubule-based molecular motor kinesin-1. Attachment of kinesin-1 to mitochondria involves the outer mitochondrial membrane protein mitochondrial Rho GTPase-1 (Miro1) which acts as a sensor for cytosolic calcium levels ([Ca2+]c); elevated [Ca2+]c disrupts mitochondrial transport via an effect on Miro1. To gain insight into the mechanisms underlying the VAPBP56S effect on mitochondrial transport, we monitored [Ca2+]c levels in VAPBP56S-expressing neurons. Expression of VAPBP56S but not VAPB increased resting [Ca2+]c and this was associated with a reduction in the amounts of tubulin but not kinesin-1 that were associated with Miro1. Moreover, expression of a Ca2+ insensitive mutant of Miro1 rescued defective mitochondrial axonal transport and restored the amounts of tubulin associated with the Miro1/kinesin-1 complex to normal in VAPBP56S-expressing cells. Our results suggest that ALS mutant VAPBP56S perturbs anterograde mitochondrial axonal transport by disrupting Ca2+ homeostasis and effecting the Miro1/kinesin-1 interaction with tubulin.

FE65 is an adaptor protein that binds to the amyloid precursor protein (APP). As such, FE65 has been implicated in the pathogenesis of Alzheimer's disease. In addition, evidence suggests that FE65 is involved in brain development. It is generally believed that FE65 participates in these processes by recruiting various interacting partners to form functional complexes. Here, we show that via its first phosphotyrosine binding (PTB) domain, FE65 binds to the small GTPase ADP-ribosylation factor 6 (ARF6). FE65 preferentially binds to ARF6-GDP, and they colocalize in neuronal growth cones. Interestingly, FE65 stimulates the activation of both ARF6 and its downstream GTPase Rac1, a regulator of actin dynamics, and functions in growth cones to stimulate neurite outgrowth. We show that transfection of FE65 and/or ARF6 promotes whereas small interfering RNA knockdown of FE65 or ARF6 inhibits neurite outgrowth in cultured neurons as compared to the mock-transfected control cells. Moreover, knockdown of ARF6 attenuates FE65 stimulation of neurite outgrowth and defective neurite outgrowth seen in FE65-deficient neurons is partially corrected by ARF6 overexpression. Notably, the stimulatory effect of FE65 and ARF6 on neurite outgrowth is abrogated either by dominant-negative Rac1 or knockdown of Rac1. Thus, we identify FE65 as a novel regulator of neurite outgrowth via controlling ARF6-Rac1 signaling.

Fronto-temporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are two related and incurable neurodegenerative diseases. Features of these diseases include pathological protein inclusions in affected neurons with TAR DNA-binding protein 43 (TDP-43), dipeptide repeat proteins derived from the C9ORF72 gene, and fused in sarcoma (FUS) representing major constituent proteins in these inclusions. Mutations in C9ORF72 and the genes encoding TDP-43 and FUS cause familial forms of FTD/ALS which provides evidence to link the pathology and genetics of these diseases. A large number of seemingly disparate physiological functions are damaged in FTD/ALS. However, many of these damaged functions are regulated by signalling between the endoplasmic reticulum and mitochondria, and this has stimulated investigations into the role of endoplasmic reticulum-mitochondria signalling in FTD/ALS disease processes. Here, we review progress on this topic.