Mitofusin 2 (Mfn2) is a large GTPase involved in the fusion of mitochondrial outer membrane. After the initial report from Luca Scorrano’s group suggesting the potential involvement of Mfn2 in ER-mitochondrial tethering, Mfn2 is among the most widely studied ER-mitochondria tethering protein factors but its exact role in MERC regulation is still in hot debate. In 2008, Luca Scoranno and colleagues found that Mfn2 ablation caused dramatic defects in ER morphology in vitro in mouse embryonic fibroblasts(MEFs) and HeLa cells [
18]. It turned out that Mfn2 is enriched at MERC and is required for the juxtaposition of ER to mitochondria [
18]. ER-located Mfn2 interacts
in trans with mitochondrial mitofusins (i.e., Mfn1 or Mfn2) to form trans-organelle hetero- or homo- dimer tethers to bridge the mitochondria onto ER which allows efficient calcium transfer between ER and mitochondria [
18,
19]. The confocal microscopy analysis revealed that the distance between ER and mitochondria increases in cells lacking Mfn2 which impairs mitochondrial calcium uptake [
18,
19]. Indeed, the presence and enrichment of Mfn2 in the interface between ER and mitochondria is directly demonstrated by immunoelectron microscopy in Melanocytes [
20]. Moreover, Mfn2-mediated ER-mitochondria tethering is regulated by MitoL-dependent activation of mitochondrial Mfn2 through ubiquitination and MitoL ablation inhibited Mfn2 complex formation and caused Mfn2 mislocalization from MAM to non-MAM ER [
21]. Owing to its well-known role in the tethering of adjacent membranes, Mfn2 is widely accepted as a major regulator of MERC in different tissues [
18,
19,
22]. However, such a view was challenged by a quantitative electron microscopic study followed by a series morphological and functional studies from multiple groups: In 2012, Pierre C et al. reported that the percentage of mitochondria in close contact with ER tubules was increased in Mfn2 defective MEFs when compared to wildtype MEFs (4.91% vs 2.25%, the distance between ER and mitochondria was restricted to <20 nm in the study). In 2015, Riccardo Filadia et al. confirmed this EM findings that the ER-mitochondria crosslink was increased in Mfn2 defective MEFs and SH-SY5Y cells and provided functional evidence of an elevated calcium flow from ER to mitochondria in Mfn2 defective MEFs [
23]. While they also confirmed the confocal microscopy results from Scoranno’s group that Mfn2 deficiency caused a net decrease in the overlapping area between ER and mitochondria, they noted that the dramatic changes in mitochondrial morphology caused by Mfn2 ablation resulted in changes in mitochondrial area which could make classical colocalization analysis unsuitable. Excluding the confounding effects of area change, they found that mitochondrial perimeter colocalizing with the ER actually increased in Mfn2 deficient cells [
23]. Other groups also reported that Mfn2 knockdown increased ER-mitochondria tethering and calcium transfer from ER to mitochondria [
24]. Along this line, deficiency in Gp78 (a.k.a. autocrine motility factor receptor, AMFR), an endoplasmic-reticulum (ER)-associated protein degradation E3 ubiquitin ligase involved in the degradation of Mfn2, caused significantly decreased rough ER-mitochondria crosstalk as evaluated by EM which could be blocked by Mfn2 knockdown, suggesting that Gp78 might promote ER-mitochondria interaction through degradation of Mfn2 [
25]. Collectively, these studies strongly challenges the role of Mfn2 as an essential component of ER-mitochondrial tethering but argues for a role of Mfn2 as a negative regulator of organelle apposition. Most recently, Luca Scorrano’s group responded to these challenges by providing further ultramorphometric and confocal microscopic evidence based on unbiased fluorescent probes of ER-mitochondrial proximity to demonstrate that Mfn2 ablation increases ER-mitochondria distance which resulted in impaired mitochondrial calcium uptake in an mitochondrial calcium uniporter (MCU)-independent manner in MEF cells [
22], yet the number of ER-mitochondria contacts were not directly addressed in this study and it is not clear whether and how larger ER-mitochondrial distance may affect ER-mitochondrial tethering. Further research is needed to fully elucidate the role of Mfn2 at MERC.