Proteomic Analysis of Retinal Mitochondria-Associated ER Membranes Identified Novel Proteins of Retinal Degeneration in Long-Term Diabetes

Wang, Joshua J.; Park, Karen Sophia; Dhimal, Narayan; Shen, Shichen; Tang, Xixiang; Qu, Jun; Zhang, Sarah X.

doi:10.3390/cells11182819

Cited by 9 publications

(4 citation statements)

References 66 publications

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“…The mitochondria's strategic localization close to ER/SR calcium channels makes it possible to decode calcium signals. Recent reports in cardiomyocytes indicated increased MAMs in db/db mice and a type 1 diabetic animal model [71]. Both models with muscular systolic dysfunction show increased mitochondrial calcium uptake.…”

Section: Discussionmentioning

confidence: 98%

The SIRT3-ATAD3A axis regulates MAM dynamics and mitochondrial calcium homeostasis in cardiac hypertrophy

Li,

Hu,

et al. 2024

Int. J. Biol. Sci.

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Mitochondria are energy-producing organelles that are mobile and harbor dynamic network structures. Although mitochondria and endoplasmic reticulum (ER) play distinct cellular roles, they are physically connected to maintain functional homeostasis. Abnormal changes in this interaction have been linked to pathological states, including cardiac hypertrophy. However, the exact regulatory molecules and mechanisms are yet to be elucidated. Here, we report that ATPase family AAA-domain containing protein 3A (ATAD3A) is an essential regulator of ER-mitochondria interplay within the mitochondria-associated membrane (MAM). ATAD3A prevents isoproterenol (ISO)-induced mitochondrial calcium accumulation, improving mitochondrial dysfunction and ER stress, which preserves cardiac function and attenuates cardiac hypertrophy. We also find that ATAD3A is a new substrate of NAD + -dependent deacetylase Sirtuin 3 (SIRT3). Notably, the heart mitochondria of SIRT3 knockout mice exhibited excessive formation of MAMs. Mechanistically, ATAD3A specifically undergoes acetylation, which reduces self-oligomerization and promotes cardiac hypertrophy. ATAD3A oligomerization is disrupted by acetylation at K134 site, and ATAD3A monomer closely interacts with the IP3R1-GRP75-VDAC1 complex, which leads to mitochondrial calcium overload and dysfunction. In summary, ATAD3A localizes to the MAMs, where it protects the homeostasis of ER-mitochondria contacts, quenching mitochondrial calcium overload and keeping mitochondrial bioenergetics unresponsive to ER stress. The SIRT3-ATAD3A axis represents a potential therapeutic target for cardiac hypertrophy.

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Section: Discussionmentioning

confidence: 98%

The SIRT3-ATAD3A axis regulates MAM dynamics and mitochondrial calcium homeostasis in cardiac hypertrophy

Li,

Hu,

et al. 2024

Int. J. Biol. Sci.

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“…An initial proteomic study of MAM was published in 2011, identifying 991 unique proteins by using SILAC quantitation [ 39 ]. After that, a few of studies have been performed on MAM proteomics, such as the cerebral cortex of mouse in Alzheimer’s Disease model [ 40 ], retina and brain MAMs isolated from diabetes mellitus mice [ 41 , 42 ],and so on. Currently, research on MAM proteomics has primarily concentrated on neurological disorders, with relatively limited exploration in other fields, particularly in the context of organ ischemic reperfusion injury.…”

Section: Discussionmentioning

confidence: 99%

Proteomic analysis of mitochondria associated membranes in renal ischemic reperfusion injury

Li,

Wang,

Cao

et al. 2024

J Transl Med

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Background The mitochondria and endoplasmic reticulum (ER) communicate via contact sites known as mitochondria associated membranes (MAMs). Many important cellular functions such as bioenergetics, mitophagy, apoptosis, and calcium signaling are regulated by MAMs, which are thought to be closely related to ischemic reperfusion injury (IRI). However, there exists a gap in systematic proteomic research addressing the relationship between these cellular processes. Methods A 4D label free mass spectrometry-based proteomic analysis of mitochondria associated membranes (MAMs) from the human renal proximal tubular epithelial cell line (HK-2 cells) was conducted under both normal (N) and hypoxia/reperfusion (HR) conditions. Subsequent differential proteins analysis aimed to characterize disease-relevant signaling molecules. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis was applied to total proteins and differentially expressed proteins, encompassing Biological Process (BP), Cell Component (CC), Molecular Function (MF), and KEGG pathways. Further, Protein–Protein Interaction Network (PPI) exploration was carried out, leading to the identification of hub genes from differentially expressed proteins. Notably, Mitofusion 2 (MFN2) and BCL2/Adenovirus E1B 19-kDa interacting protein 3(BNIP3) were identified and subsequently validated both in vitro and in vivo. Finally, the impact of MFN2 on MAMs during hypoxia/reoxygenation was explored through regulation of gene expression. Subsequently, a comparative proteomics analysis was conducted between OE-MFN2 and normal HK-2 cells, providing further insights into the underlying mechanisms. Results A total of 4489 proteins were identified, with 3531 successfully quantified. GO/KEGG analysis revealed that MAM proteins were primarily associated with mitochondrial function and energy metabolism. Differential analysis between the two groups showed that 688 proteins in HR HK-2 cells exhibited significant changes in expression level with P-value < 0.05 and HR/N > 1.5 or HR/N < 0.66 set as the threshold criteria. Enrichment analysis of differentially expressed proteins unveiled biological processes such as mRNA splicing, apoptosis regulation, and cell division, while molecular functions were predominantly associated with energy metabolic activity. These proteins play key roles in the cellular responses during HR, offering insights into the IRI mechanisms and potential therapeutic targets. The validation of hub genes MFN2 and BNIP3 both in vitro and vivo was consistent with the proteomic findings. MFN2 demonstrated a protective role in maintaining the integrity of mitochondria associated membranes (MAMs) and mitigating mitochondrial damage following hypoxia/reoxygenation injury, this protective effect may be associated with the activation of the PI3K/AKT pathway. Conclusions The proteins located in mitochondria associated membranes (MAMs) are implicated in crucial roles during renal ischemic reperfusion injury (IRI), with MFN2 playing a pivotal regulatory role in this context.

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“…[ 36 ] MAM dysfunction has been proven associated with neurodegeneration and retinal degeneration. [ 44,45 ] Given the fact that MAM dysregulation serves as the key point of cell damage upon lethal stimulation, [ 21,46,47 ] we wondered if there is an interrelationship between PARP‐1 and MAMs under oxidative stress. Here, the application of olaparib significantly alleviated H 2 O 2 ‐induced retinal cell viability loss.…”

Section: Discussionmentioning

confidence: 99%

Mitochondria‐associated endoplasmic reticulum membranes dysfunction contributes to PARP‐1‐dependent cell death under oxidative stress in retinal precursor cells

Yang

Lü

et al. 2023

J Biochem & Molecular Tox

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Persistent poly (ADP-ribose) polymerase 1 (PARP-1) activation has proven detrimental and can lead to PARP-1-dependent cell death. Mitochondria-associated endoplasmic reticulum (ER) membranes (MAMs) serve as essential hubs for many biological pathways, such as autophagy and mitochondria fission and fusion. This study aimed to alleviate the effects of hydrogen peroxide (H 2 O 2 )-induced persistent PARP-1 activation and MAM dysregulation by the usage of a PARP-1 inhibitor. Results showed that receptorinteracting protein kinase (RIPK) 1 inhibitor (necrostatin-1) and PARP-1 inhibitor (olaparib) protected retinal precursor cells from H 2 O 2 -induced death, while a pan-caspase inhibitor (Z-VAD-FMK) failed to protect R28 cells. Olaparib also alleviated H 2 O 2 -induced MAM dysregulation, as evidenced by decreased VDAC1/ITPR3 interactions and reduced mitochondrial membrane potential collapse. Additionally, olaparib also inhibited H 2 O 2induced autophagy. Inhibiting autophagic flux increased MAM signaling under both normal and oxidative conditions. Furthermore, H 2 O 2 treatment caused a reduction in the protein level of mitofusin-2 (MFN2) in a dose-and time-dependent manner. Mfn2 knockdown was found to further magnify MAM dysregulation and mitochondrial dysfunction under normal and oxidative conditions. Mfn2 overexpression surprisingly enhanced H 2 O 2 -induced MAM signaling and failed to rescue H 2 O 2 -induced mitochondrial dysfunction. These results indicate that MAMs probably serve as a membrane source for oxidative stress-associated autophagy. MAM dysregulation also contributed to H 2 O 2induced PARP-1-dependent cell death. However, more studies are required to decipher the link between the modulation of Mfn2 expression, changes in MAM integrity, and alterations in mitochondrial performances.

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Proteomic Analysis of Retinal Mitochondria-Associated ER Membranes Identified Novel Proteins of Retinal Degeneration in Long-Term Diabetes

Cited by 9 publications

References 66 publications

The SIRT3-ATAD3A axis regulates MAM dynamics and mitochondrial calcium homeostasis in cardiac hypertrophy

The SIRT3-ATAD3A axis regulates MAM dynamics and mitochondrial calcium homeostasis in cardiac hypertrophy

Proteomic analysis of mitochondria associated membranes in renal ischemic reperfusion injury

Mitochondria‐associated endoplasmic reticulum membranes dysfunction contributes to PARP‐1‐dependent cell death under oxidative stress in retinal precursor cells

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