bims-mecosi 2023-10-01 papers

Issue of 2023–10–01
five papers selected by
Verena Kohler, University of Graz

Cells. 2023 09 20. pii: 2322. [Epub ahead of print]12(18):

TRPV1 Channels Are New Players in the Reticulum-Mitochondria Ca2+ Coupling in a Rat Cardiomyoblast Cell Line.

Nolwenn Tessier, Mallory Ducrozet, Maya Dia, Sally Badawi, Christophe Chouabe, Claire Crola Da Silva, Michel Ovize, Gabriel Bidaux, Fabien Van Coppenolle, Sylvie Ducreux.

The Ca2+ release in microdomains formed by intercompartmental contacts, such as mitochondria-associated endoplasmic reticulum membranes (MAMs), encodes a signal that contributes to Ca2+ homeostasis and cell fate control. However, the composition and function of MAMs remain to be fully defined. Here, we focused on the transient receptor potential vanilloid 1 (TRPV1), a Ca2+-permeable ion channel and a polymodal nociceptor. We found TRPV1 channels in the reticular membrane, including some at MAMs, in a rat cardiomyoblast cell line (SV40-transformed H9c2) by Western blotting, immunostaining, cell fractionation, and proximity ligation assay. We used chemical and genetic probes to perform Ca2+ imaging in four cellular compartments: the endoplasmic reticulum (ER), cytoplasm, mitochondrial matrix, and mitochondrial surface. Our results showed that the ER Ca2+ released through TRPV1 channels is detected at the mitochondrial outer membrane and transferred to the mitochondria. Finally, we observed that prolonged TRPV1 modulation for 30 min alters the intracellular Ca2+ equilibrium and influences the MAM structure or the hypoxia/reoxygenation-induced cell death. Thus, our study provides the first evidence that TRPV1 channels contribute to MAM Ca2+ exchanges.

Keywords: Ca2+ homeostasis; ER–mitochondria contact sites; H9c2; TRP channels; TRPV1; hypoxia–reoxygenation

DOI: https://doi.org/10.3390/cells12182322

Front Cell Dev Biol. 2023 ;11 1171440

Reduced ER-mitochondrial contact sites and mitochondrial Ca2+ flux in PRKN-mutant patient tyrosine hydroxylase reporter iPSC lines.

Mutsumi Yokota, Yutaro Yoshino, Mitsuko Hosoi, Ryota Hashimoto, Soichiro Kakuta, Takahiro Shiga, Kei-Ichi Ishikawa, Hideyuki Okano, Nobutaka Hattori, Wado Akamatsu, Masato Koike.

Endoplasmic reticulum-mitochondrial contact sites (ERMCS) play an important role in mitochondrial dynamics, calcium signaling, and autophagy. Disruption of the ERMCS has been linked to several neurodegenerative diseases, including Alzheimer's disease (AD), Parkinson's disease (PD), and amyotrophic lateral sclerosis (ALS). However, the etiological role of ERMCS in these diseases remains unclear. We previously established tyrosine hydroxylase reporter (TH-GFP) iPSC lines from a PD patient with a PRKN mutation to perform correlative light-electron microscopy (CLEM) analysis and live cell imaging in GFP-expressing dopaminergic neurons. Here, we analyzed ERMCS in GFP-expressing PRKN-mutant dopaminergic neurons from patients using CLEM and a proximity ligation assay (PLA). The PLA showed that the ERMCS were significantly reduced in PRKN-mutant patient dopaminergic neurons compared to the control under normal conditions. The reduction of the ERMCS in PRKN-mutant patient dopaminergic neurons was further enhanced by treatment with a mitochondrial uncoupler. In addition, mitochondrial calcium imaging showed that mitochondrial Ca2+ flux was significantly reduced in PRKN-mutant patient dopaminergic neurons compared to the control. These results suggest a defect in calcium flux from ER to mitochondria is due to the decreased ERMCS in PRKN-mutant patient dopaminergic neurons. Our study of ERMCS using TH-GFP iPSC lines would contribute to further understanding of the mechanisms of dopaminergic neuron degeneration in patients with PRKN mutations.

Keywords: ER-mitochondrial contact sites; PRKN; dopaminergic neurons; iPSC; tyrosine hydroxylase reporter

DOI: https://doi.org/10.3389/fcell.2023.1171440

Int Immunopharmacol. 2023 Sep 22. pii: S1567-5769(23)01286-9. [Epub ahead of print]124(Pt B): 110961

Aflatoxin B1 exposure triggers inflammation and premature skin aging via ERMCS/Ca2+/ROS signaling cascade.

Deyi Wu, Yi Wu, Meng Zhang, Hainan Lan.

Aflatoxin B1 (AFB1) is a recognized hazard environmental contaminant mainly found in cereal and fruits. The toxicity of AFB1 exposure to various organs has been revealed in some literature. In current study, we explored the effect of AFB1 exposure on premature aging/senescence of skin. In vivo, 8-week-old C57 mice were used as models to evaluate the effect of dietary AFB1 exposure on premature skin aging. The results showed that AFB1 exposure caused premature skin aging by testing aging markers. Additionally, AFB1 led to oxidative stress and inflammatory response. In vitro, AFB1 exposure triggered premature cellular senescence in mouse skin fibroblasts cells (L929 cells) by assessing a range of cellular senescence-related markers. Further, the potential molecular mechanism by which AFB1 induce the premature skin aging was studied. ROS and Ca2+ is proven to be the key molecules in AFB1-induced cellular senescence. Further, through eliminating Ca2+, AFB1-caused oxidative stress and cellular senescence were both attenuated, suggesting that Ca2+ overload in the mitochondria results in cellular senescence by increasing ROS production. Next, we analyzed the causes of Ca2+ overload, and results showed that AFB1 exposure induces Ca2+ overload through increasing the formation of mitoguardin (Miga) and vesicle-associated membrane protein (VAMP)-associated protein (Vap33)-mediated endoplasmic reticulum (ER)-mitochondria contact sites (ERMCS). AFB1 exposure also inhibited mitophagy, leading to accelerate L929 cell senescence. In short, combining in vivo and in vitro results, we demonstrate that exposure to AFB1 causes premature skin aging, which is dependent on ERMCS/Ca2+/ROS/ signaling axis. The current study suggests that prolonged exposure to AFB1 makes skin more vulnerable to damage.

Keywords: AFB1; Ca(2+)overload; Inflammation; Oxidative stress; Premature aging; Skin

DOI: https://doi.org/10.1016/j.intimp.2023.110961

World J Gastroenterol. 2023 Sep 14. 29(34): 5038-5053

Suberoylanilide hydroxamic acid upregulates reticulophagy receptor expression and promotes cell death in hepatocellular carcinoma cells.

Jia-Yao Li, Tian Tian, Bing Han, Ting Yang, Yi-Xin Guo, Jia-Yu Wu, Yu-Si Chen, Qin Yang, Ru-Jia Xie.

BACKGROUND: Hepatocellular carcinoma (HCC) is a common clinical condition with a poor prognosis and few effective treatment options. Potent anticancer agents for treating HCC must be identified. Epigenetics plays an essential role in HCC tumorigenesis. Suberoylanilide hydroxamic acid (SAHA), the most common histone deacetylase inhibitor agent, triggers many forms of cell death in HCC. However, the underlying mechanism of action remains unclear. Family with sequence similarity 134 member B (FAM134B)-induced reticulophagy, a selective autophagic pathway, participates in the decision of cell fate and exhibits anticancer activity. This study focused on the relationship between FAM134B-induced reticulophagy and SAHA-mediated cell death.
AIM: To elucidate potential roles and underlying molecular mechanisms of reticulophagy in SAHA-induced HCC cell death.
METHODS: The viability, apoptosis, cell cycle, migration, and invasion of SAHA-treated Huh7 and MHCC97L cells were measured. Proteins related to the reticulophagy pathway, mitochondria-endoplasmic reticulum (ER) contact sites, intrinsic mitochondrial apoptosis, and histone acetylation were quantified using western blotting. ER and lysosome colocalization, and mitochondrial Ca2+ levels were characterized via confocal microscopy. The level of cell death was evaluated through Hoechst 33342 staining and propidium iodide colocalization. Chromatin immunoprecipitation was used to verify histone H4 lysine-16 acetylation in the FAM134B promoter region.
RESULTS: After SAHA treatment, the proliferation of Huh7 and MHCC97L cells was significantly inhibited, and the migration and invasion abilities were greatly blocked in vitro. This promoted apoptosis and caused G1 phase cells to increase in a concentration-dependent manner. Following treatment with SAHA, ER-phagy was activated, thereby triggering autophagy-mediated cell death of HCC cells in vitro. Western blotting and chromatin immunoprecipitation assays confirmed that SAHA regulated FAM134B expression by enhancing the histone H4 lysine-16 acetylation in the FAM134B promoter region. Further, SAHA disturbed the Ca2+ homeostasis and upregulated the level of autocrine motility factor receptor and proteins related to mitochondria-endoplasmic reticulum contact sites in HCC cells. Additionally, SAHA decreased the mitochondrial membrane potential levels, thereby accelerating the activation of the reticulophagy-mediated mitochondrial apoptosis pathway and promoting HCC cell death in vitro.
CONCLUSION: SAHA stimulates FAM134B-mediated ER-phagy to synergistically enhance the mitochondrial apoptotic pathway, thereby enhancing HCC cell death.

Keywords: Apoptosis; Autophagic cell death; Hepatocellular carcinoma; Histone H4 lysine-16; Reticulophagy; Suberoylanilide hydroxamic acid

DOI: https://doi.org/10.3748/wjg.v29.i34.5038

J Biol Chem. 2023 Sep 27. pii: S0021-9258(23)02323-2. [Epub ahead of print] 105295

Comparative proximity biotinylation implicates the small GTPase RAB18 in sterol mobilization and biosynthesis.

Loss of functional RAB18 causes the autosomal recessive condition Warburg Micro syndrome. To better understand this disease, we used proximity biotinylation to generate an inventory of potential RAB18 effectors. A restricted set of 28 RAB18-interactions were dependent on the binary RAB3GAP1-RAB3GAP2 RAB18-guanine nucleotide exchange factor (GEF) complex. 12 of these 28 interactions are supported by prior reports and we have directly validated novel interactions with SEC22A, TMCO4 and INPP5B. Consistent with a role for RAB18 in regulating membrane contact sites (MCSs), interactors included groups of microtubule/membrane-remodelling proteins, membrane-tethering and docking proteins, and lipid-modifying/transporting proteins. Two of the putative interactors, EBP and OSBPL2/ORP2, have sterol substrates. EBP is a Δ8-Δ7 sterol isomerase and ORP2 is a lipid transport protein. This prompted us to investigate a role for RAB18 in cholesterol biosynthesis. We find that the cholesterol precursor and EBP-product lathosterol accumulates in both RAB18-null HeLa cells and RAB3GAP1-null fibroblasts derived from an affected individual. Further, de novo cholesterol biosynthesis is impaired in cells in which RAB18 is absent or dysregulated, or in which ORP2 expression is disrupted. Our data demonstrate that GEF-dependent Rab-interactions are highly amenable to interrogation by proximity biotinylation and may suggest that Micro syndrome is a cholesterol biosynthesis disorder.

Keywords: BioID; EBP; ORP2; RAB18; Rab; SNARE proteins; cholesterol metabolism; lathosterol; lipid transport; protein-protein interaction

DOI: https://doi.org/10.1016/j.jbc.2023.105295