Vimentin and Desmin Intermediate Filaments Maintain Mitochondrial Membrane Potential

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Abstract

Intermediate filaments (IF) are one of the three main components of the cytoskeleton in animal cells, characterized by the diverse range of proteins that represent them in different cell types. They provide mechanical strength to cells and help position the nucleus and various organelles within the cell. Desmin is an IF protein characteristic of muscle cells, while vimentin, which has a similar structure, is found in many mesenchymal cells. During myogenesis and the regeneration of damaged muscle tissue, both of these proteins can be expressed, forming a mixed IF network. Each protein is known to regulate mitochondrial function in the cells where it is present, including mitochondrial localization and the maintenance of mitochondrial membrane potential. However, the regulation of mitochondrial function in cells with mixed IFs remains unclear. To investigate how the simultaneous presence of these proteins affects mitochondrial membrane potential, we utilized BHK21 cells as a model system, expressing both vimentin and desmin IFs. The expression of either protein individually or both proteins simultaneously was suppressed using gene knockout and/or RNA interference. It was found that removal of either protein did not affect the mitochondrial membrane potential, which remained unchanged compared to when both proteins were present. Simultaneous removal of both proteins resulted in a 20% reduction in mitochondrial membrane potential, indicating that vimentin and desmin both play a critical role in its maintenance.

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About the authors

A. A. Dayal

Institute of Protein Research of the Russian Academy of Sciences

Email: alexminin@gmail.com
Russian Federation, 142290, Pushchino, Moscow Region

O. I. Parfentyeva

Institute of Protein Research of the Russian Academy of Sciences

Email: alexminin@gmail.com
Russian Federation, 142290, Pushchino, Moscow Region

W. Huiying

Institute of Protein Research of the Russian Academy of Sciences

Email: alexminin@gmail.com
Russian Federation, 142290, Pushchino, Moscow Region

A. S. Shakhov

Institute of Protein Research of the Russian Academy of Sciences; Lomonosov Moscow State University

Email: alexminin@gmail.com
Russian Federation, 142290, Pushchino, Moscow Region; 119992, Moscow

I. B. Alieva

Institute of Protein Research of the Russian Academy of Sciences; Lomonosov Moscow State University

Email: alexminin@gmail.com
Russian Federation, 142290, Pushchino, Moscow Region; 119992, Moscow

A. A. Minin

Institute of Protein Research of the Russian Academy of Sciences

Author for correspondence.
Email: alexminin@gmail.com
Russian Federation, 142290, Pushchino, Moscow Region

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Supplementary files

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2. Fig. 1. Vimentin (green) and desmin (red) PFs in VNK21 cells detected with the respective antibodies. Images were obtained using high-resolution microscopy. Scale bar 10 μm

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3. Fig. 2. Destruction of desmin (a, b) and vimentin (c, d) PFs by RNA interference in VNK21 cells transfected with plasmids pG-SHIN2-des and pG-SHIN2-vim, respectively. Control cells (e, f) were transfected with plasmid pG-SHIN2-scr. PFs were stained using antibodies against desmin (Des) and vimentin (Vim). Transfected cells were identified by GFP expression. Scale bar 10 μm

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4. Fig. 3. No change in mitochondrial membrane potential in VNA21 cells after knockdown of desmin (shRNA-des) and vimentin (shRNA-vim) by RNA interference. Control cells were transfected with plasmid pG-SHIN2-scr. Data are presented as mean values of mitochondrial fluorescence in the indicated number of cells with standard error as a percentage of the mean value in untransfected cells (-)

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5. Fig. 4. Immunoblotting of cell homogenates of the original VNK21 (WT) cell line and the VNK21(Des-/-) and VNK21(Vim-/-) cell lines resulting from knockout of desmin and vimentin genes, respectively. Alpha-tubulin was used as a control

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6. Fig. 5. PFs in cells of VNC21(Des-/-) (a, b) and VNC21(Vim-/-) (c, d) cell lines derived from knockout of desmin and vimentin genes, respectively, detected by immunofluorescence with antibodies against vimentin (a, c) or desmin (b, d). Scale of 10 μm

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7. Fig. 6. Removal of vimentin (a, b) in VNC21(Des-/-) cells and desmin (c, d) in VNC21(Vim-/-) cells transfected with plasmids pG-SHIN2-vim and pG-SHIN2-des, respectively, by RNA interference. PFs were stained using antibodies against vimentin (b) and desmin (d). Transfected cells were identified by GFP expression (a, c, shown with arrows). Scale bar 10 μm

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8. Fig. 7. Decreased mitochondrial membrane potential in VNA21(Vim-/-) cells (a) and in VNA21(Des-/-) cells (b) as a result of RNA interference of desmin or vimentin, respectively. Cells transfected with plasmid pG-SHIN2-scr were used as controls. Data are presented as mean values of mitochondrial fluorescence in the indicated number of cells with standard error as a percentage of the mean value in untransfected cells

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