bims-mecosi 2022-05-01 papers

Issue of 2022–05–01
three papers selected by
Verena Kohler, Stockholm University

Front Cell Dev Biol. 2022 ;10 829545

Correlative Organelle Microscopy: Fluorescence Guided Volume Electron Microscopy of Intracellular Processes.

Sergey V Loginov, Job Fermie, Jantina Fokkema, Alexandra V Agronskaia, Cilia De Heus, Gerhard A Blab, Judith Klumperman, Hans C Gerritsen, Nalan Liv.

Intracellular processes depend on a strict spatial and temporal organization of proteins and organelles. Therefore, directly linking molecular to nanoscale ultrastructural information is crucial in understanding cellular physiology. Volume or three-dimensional (3D) correlative light and electron microscopy (volume-CLEM) holds unique potential to explore cellular physiology at high-resolution ultrastructural detail across cell volumes. However, the application of volume-CLEM is hampered by limitations in throughput and 3D correlation efficiency. In order to address these limitations, we describe a novel pipeline for volume-CLEM that provides high-precision (<100 nm) registration between 3D fluorescence microscopy (FM) and 3D electron microscopy (EM) datasets with significantly increased throughput. Using multi-modal fiducial nanoparticles that remain fluorescent in epoxy resins and a 3D confocal fluorescence microscope integrated into a Focused Ion Beam Scanning Electron Microscope (FIB.SEM), our approach uses FM to target extremely small volumes of even single organelles for imaging in volume EM and obviates the need for post-correlation of big 3D datasets. We extend our targeted volume-CLEM approach to include live-cell imaging, adding information on the motility of intracellular membranes selected for volume-CLEM. We demonstrate the power of our approach by targeted imaging of rare and transient contact sites between the endoplasmic reticulum (ER) and lysosomes within hours rather than days. Our data suggest that extensive ER-lysosome and mitochondria-lysosome interactions restrict lysosome motility, highlighting the unique capabilities of our integrated CLEM pipeline for linking molecular dynamic data to high-resolution ultrastructural detail in 3D.

Keywords: correlative light and electron microscopy; live-cell imaging; lysosome; organelle contact site; volume electron microscopy

DOI: https://doi.org/10.3389/fcell.2022.829545

Kidney Int. 2022 Apr 22. pii: S0085-2538(22)00336-2. [Epub ahead of print]

Reticulon-1A mediates diabetic kidney disease progression through endoplasmic reticulum-mitochondrial contacts in tubular epithelial cells.

Yifan Xie, Jing E, Hong Cai, Fang Zhong, Wenzhen Xiao, Ronald E Gordon, Lois Wang, Ya-Li Zheng, Aihua Zhang, Kyung Lee, John Cijiang He.

Recent epidemiological studies suggest that some patients with diabetes progress to kidney failure without significant albuminuria and glomerular injury, suggesting a critical role of kidney tubular epithelial cell (TEC) injury in diabetic kidney disease (DKD) progression. However, the major risk factors contributing to TEC injury and progression in DKD remain unclear. We previously showed that expression of endoplasmic reticulum-resident protein Reticulon-1A (RTN1A) increased in human DKD, and the increased RTN1A expression promoted TEC injury through endoplasmic reticulum (ER) stress response. Here, we show that TEC-specific RTN1A overexpression worsened DKD in mice, evidenced by enhanced tubular injury, tubulointerstitial fibrosis, and kidney function decline. But RTN1A overexpression did not exacerbate diabetes-induced glomerular injury or albuminuria. Notably, RTN1A overexpression worsened both ER stress and mitochondrial dysfunction in TECs under diabetic conditions by regulation of ER-mitochondria contacts. Mechanistically, ER-bound RTN1A interacted with mitochondrial hexokinase-1 and the voltage-dependent anion channel-1 (VDAC1), interfering with their association. This disengagement of VDAC1 from hexokinase-1 resulted in activation of apoptotic and inflammasome pathways, leading to TEC injury and loss. Thus, our observations highlight the importance of ER-mitochondrial crosstalk in TEC injury and the salient role of RTN1A-mediated ER-mitochondrial contact regulation in DKD progression.

Keywords: Diabetic kidney disease; and endoplasmic reticulum-mitochondrial contacts; endoplasmic reticulum stress; kidney tubular epithelial cells

DOI: https://doi.org/10.1016/j.kint.2022.02.038

Andrology. 2022 Apr 24.

Low androgen status inhibits erectile function by up-regulating the expression of proteins of mitochondria-associated membranes in rat corpus cavernosum.

Hao-Zhou Yang, Wen-Ju Xiong, Xiong Li, Jun Jiang, Rui Jiang.

OBJECTIVE: To investigate the effect of low androgen status on mitochondria-associated membranes (MAMs) and its relationship with erectile function.
METHODS: A total of 36 eight-week-old male Sprague-Dawley rats were randomly divided into 6 groups: the control (sham-operated) group, the castration group, the castration + testosterone (cast + T) group, the control + siRNA group, the cast + siRNA group, and the cast + empty vector group. Testosterone propionate (3 mg/kg) was subcutaneously injected into the rats in the cast + T group every other day starting from the second day after the surgery. Four weeks later, lentiviral vectors carrying phosphofurin acidic cluster sorting protein 2 (PACS-2) gene-specific siRNA (1 × 108 TU/ mL, 10 μL) were injected into the rats in the siRNA groups. At the 6th week of castration, the ratio of the maximum intracavernous pressure/the mean arterial pressure (ICPmax/MAP), the levels of nitric oxide (NO), endothelial nitric oxide synthase (eNOS), phospho-eNOS (p-eNOS), fatty acid-CoA ligase 4 (FACL-4), PACS-2, and inositol 1,4,5-trisphosphate receptor type 1 (IP3R1) in the penile corpus cavernosum were determined.
RESULTS: The FACL-4, PACS-2 and IP3R1 were primarily localized in the cytoplasm of endothelial cells and part of smooth muscle cells in the corpus cavernosum. The level of NO, the ratio of ICPmax/MAP, and p-eNOS/eNOS were decreased significantly in the castration group compared with the control group (P<0.01). The expressions of FACL-4, PACS-2, and IP3R1 were increased significantly in the castration group compared with the control group (P<0.01). The level of NO, the ratios of ICPmax/MAP, and the ratio of p-eNOS/eNOS were increased significantly in the cast + siRNA group compared with the castration group (P<0.01). The expressions of FACL-4 and PACS-2 were decreased significantly in the cast + siRNA group compared with the castration group (P<0.01).
CONCLUSION: Low androgen status up-regulated the expressions of patients in MAMs (FACL-4, PACS-2 and IP3R1) in the corpus cavernosum and inhibited the eNOS/NO/cGMP signaling pathway, resulting in impaired erectile function in rats. Erectile function may be improved by inhibiting the high expression of PACS-2 in the corpus cavernosum under low androgen state. This article is protected by copyright. All rights reserved.

Keywords: Androgen; RNA interference; corpus cavernosum; mitochondria-associated membranes

DOI: https://doi.org/10.1111/andr.13188