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ECM stiffness regulates calcium influx into mitochondria via tubulin and VDAC1 activity.

ABSTRACT: Calcium ions (Ca²⁺) play pivotal roles in regulating numerous cellular functions, including metabolism and growth, in normal and cancerous cells. Consequently, Ca²⁺ signaling is a vital determinant of cell fate and influences both cell survival and death. These intracellular signals are susceptible to modulation by various factors, including changes in the extracellular environment, which leads to mechanical alterations. However, the effect of extracellular matrix (ECM) stiffness variations on intracellular Ca²⁺ signaling remains underexplored. In this study, we aimed to elucidate the mechanisms of Ca²⁺ regulation through the mitochondria, which are crucial to Ca²⁺ homeostasis. We investigated how Ca²⁺ regulatory mechanisms adapt to different levels of ECM stiffness by simultaneously imaging the mitochondria and endoplasmic reticulum (ER) in live cells using genetically encoded biosensors. Our findings revealed that the uptake of mitochondrial Ca²⁺ through Voltage-Dependent Anion Channel 1 (VDAC1), facilitated by intracellular tubulin, is influenced by ECM stiffness. Unraveling these Ca²⁺ regulatory mechanisms under various conditions offers a novel perspective for advancing biomedical studies involving Ca²⁺ signaling.

SUBMITTER: Kim M

PROVIDER: S-EPMC11363740 | biostudies-literature | 2024

REPOSITORIES: biostudies-literature

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ECM stiffness regulates calcium influx into mitochondria via tubulin and VDAC1 activity.

Kim Minji M Han Kiseok K Choi Gyuho G Ahn Sanghyun S Suh Jung-Soo JS Kim Tae-Jin TJ

Animal cells and systems 20240829 1

Calcium ions (Ca<sup>2+</sup>) play pivotal roles in regulating numerous cellular functions, including metabolism and growth, in normal and cancerous cells. Consequently, Ca<sup>2+</sup> signaling is a vital determinant of cell fate and influences both cell survival and death. These intracellular signals are susceptible to modulation by various factors, including changes in the extracellular environment, which leads to mechanical alterations. However, the effect of extracellular matrix (ECM) sti ...[more]

PMID: 39220629

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Project description:The voltage-dependent anion channel 1 (VDAC1) was first described as a mitochondrial porin that mediates the flux of metabolites and ions, thereby integrating both cell survival and death signals. In the nervous system, the functional roles of VDAC1 remain poorly understood. Herein, the rat retina was employed to study VDAC1. First, it was observed that even subtle changes in VDAC1 levels affect neuronal survival, inducing severe alterations in the retinal morphology. We next examined the regulation of VDAC1 after traumatic retinal injury. After mechanical trauma, SOD1 translocates towards the nucleus, which is insufficient to contain the consequences of oxidative stress, as determined by the evaluation of protein carbonylation. Using in vitro models of oxidative stress and mechanical injury in primary retinal cell cultures, it was possible to determine that inhibition of VDAC1 oligomerization by 4'-diisothiocyano-2,2'-disulfonic acid stilbene (DIDS) rescues cell viability, impacting microglial cell activation. We next focused on the regulation of VDAC1 after retinal mechanical injury. VDAC1 was promptly upregulated 2 h after lesion in the plasma membrane and endoplasmic reticulum rather than in the mitochondria, and multimers of VDAC1 were assembled after lesion. DIDS intraocular application decreased apoptosis and prevented microglial polarization, which confirmed in vitro observations. Considering the role of microglia in neuroinflammation, multiplex evaluation of cytokines showed that DIDS application disorganized the inflammatory response 2 h after the lesion, matching the fast regulation of VDAC1. Taken together, data disclosed that fine regulation of VDAC1 influences neuronal survival, and pharmacological inhibition after trauma injury has neuroprotective effects. This protection may be attributed to the effects on VDAC1 abnormal accumulation in the plasma membrane, thereby controlling the activation of microglial cells. We concluded that VDAC1 is a putative therapeutic target in neuronal disorders since it integrates both death and survival cellular signaling.

Project description:BackgroundInflammation is currently recognized as one of the major causes of premature delivery. As a member of the interleukin-1β (IL-1β) family, interleukin-33 (IL-33) has been shown to be involved in normal pregnancy as well as a variety of pregnancy-related disorder. This study aims to investigate the potential function of IL-33 in uterine smooth muscle cells during labor.MethodsMyometrium samples from term pregnant (≥37 weeks gestation) women were either frozen or cells were isolated and cultured. Immunohistochemistry and western blotting were used to assess the distribution of IL-33. Cultured cells were incubated with lipopolysaccharide (LPS) to mimic inflammation as well as in the presence of 4μ8C (IRE1 inhibitor III) to block endoplasmic reticulum (ER) stress and BAPTA-AM, a calcium chelator.ResultsLPS reduced the expression of nuclear IL-33 in a time-limited manner and induced ER stress. However, knockdown of IL-33 increased LPS-induced calcium concentration, ER stress and phosphorylation of nuclear factor kappa-B (NF-κB), and P38 mitogen-activated protein kinase (P38 MAPK). In addition, siRNA IL-33 further stimulates LPS enhanced cyclooxygenase-2 (COX-2) expression via NF-κB and p38 pathways. IL-33 expression was decreased in the nucleus with the onset of labor. LPS-induced ER stress and increased expression of the labor-associated gene, COX-2, as well as IL-6 and IL-8 in cultured myometrial cells. IL-33 also increased COX-2 expression, but after it was knocked down, the stimulating effect of LPS on calcium was enhanced. 4μ8C also inhibited the expression of COX-2 markedly. The expression of calcium channels on the membrane and intracellular free calcium ion were both increased which was accompanied by phosphorylated NF-κB and p38.ConclusionsThese data suggest that IL-33 may be involved in the initiation of labor by leading to stress of the ER via an influx of calcium ions in human uterine smooth muscle cells.FundingThis study was supported by grants from the National Natural Science Foundation of China (No. 81300507).

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ECM stiffness regulates calcium influx into mitochondria via tubulin and VDAC1 activity.

Publications

ECM stiffness regulates calcium influx into mitochondria via tubulin and VDAC1 activity.

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