Biology subjects

Aoyama-Ishiwatari, S.

Publications and source records attributed to Aoyama-Ishiwatari, S..

3 recordsLinked to original sources

bioRxiv2025

Live-cell imaging reveals nutrient-dependent dynamics of ER-mitochondria contact formation via PDZD8

Cells adapt to changes in nutrient conditions by reorganizing organelle activities. Mitochondria-endoplasmic reticulum contact sites (MERCS) have been proposed to play an essential role in this adaptive response. However, the lack of suitable tools for detecting these contact sites in living cells has hindered the studies of the mechanisms regulating MERCS in response to nutritional changes. Here, we establish a novel cell line, MERCdRED, which expresses a ddFP-based fluorescent MERCS sensor. Using correlative light-electron microscopy, we show that the MERCdRED signal accurately labels the contact sites as defined by electron microscopy. Live imaging of MERCdRED cells revealed that large MERCS are more stable than smaller ones. Furthermore, in combination with knockout of the ER-mitochondria tethering protein PDZD8, we revealed that nutrient deprivation reduces MERCS area in a PDZD8-dependent manner. This new tool, together with our findings, will contribute to a better understanding of the molecular basis of cellular metabolism. Summary StatementWe established a novel cell line enabling live imaging of MERCS, which revealed their dynamic nature and demonstrated nutrient deprivation-induced MERCS reduction along with its underlying mechanisms.

cell biology↗

bioRxiv2023

Mitochondrial protein FKBP8 captures PDZD8 to form mitochondria-ER contacts

Mitochondria-ER membrane contact sites (MERCS) represent a fundamental ultrastructural feature underlying unique biochemistry and physiology in eukaryotic cells. The ER protein PDZD8 is required for the formation of MERCS in many cell types, however, its tethering partner on the outer mitochondrial membrane (OMM) is currently unknown. Here we identified the OMM protein FKBP8 as the tethering partner of PDZD8 using a combination of unbiased proximity proteomics, CRISPR-Cas9 endogenous protein tagging, Cryo-Electron Microscopy (Cryo-EM) tomography, and correlative light-EM (CLEM). Single molecule tracking revealed highly dynamic diffusion properties of PDZD8 along the ER membrane with significant pauses and capture at MERCS. Overexpression of FKBP8 was sufficient to narrow the ER-OMM distance, whereas independent versus combined deletions of these two proteins demonstrated their interdependence for MERCS formation. Furthermore, PDZD8 enhances mitochondrial complexity in a FKBP8-dependent manner. Our results identify a novel ER-mitochondria tethering complex that regulates mitochondrial morphology in mammalian cells.

cell biology↗

bioRxiv2020

NUDT21 links mitochondrial IPS-1 to RLR-containing stress granules and activates host antiviral defense

Viral RNA in the cytoplasm of mammalian host cells is recognized by retinoic acid- inducible protein-I (RIG-I)-like receptors (RLRs), which localize to cytoplasmic stress granules (SGs). Activated RLRs associate with the mitochondrial adaptor protein IPS-1, which activates antiviral host defense mechanisms including type I interferon (IFN) induction. It has remained unclear, however, how RLRs in SGs and IPS-1 in the mitochondrial outer membrane associate physically and engage in information transfer. Here we show that NUDT21, an RNA binding protein that regulates alternative transcript polyadenylation, physically associates with IPS-1 and mediates its localization to SGs in response to transfection with poly(I:C), a mimic of viral double-stranded RNA. We found that, despite its well-established function in the nucleus, a fraction of NUDT21 localizes to mitochondria in resting cells and becomes localized to SGs in response to poly(I:C) transfection. NUDT21 was also found to be required for efficient type I IFN induction in response to viral infection. Our results together indicate that NUDT21 links RLRs in SGs to mitochondrial IPS-1 and thereby activates host defense responses to viral infection.

immunology↗