Overexpression of Myo1e promotes albumin endocytosis by mouse glomerular podocytes mediated by Dynamin

Huijun Shen; Yu Bao; Chunyue Feng; Haidong Fu; Jianhua Mao

doi:10.7717/peerj.8599

Overexpression of Myo1e promotes albumin endocytosis by mouse glomerular podocytes mediated by Dynamin

PeerJ. 2020 Mar 17:8:e8599. doi: 10.7717/peerj.8599. eCollection 2020.

Authors

Huijun Shen¹, Yu Bao¹, Chunyue Feng¹, Haidong Fu¹, Jianhua Mao¹

Affiliation

¹ Department of Nephrology, The Children's Hospital of Zhejiang University School of Medicine, Hangzhou, Zhejiang, China.

Abstract

Background: As a fundamental process internalizing molecules from the plasma membrane, endocytosis plays a crucial role in podocyte biology. Our previous study has identified that overexpression of Myole may enhance podocyte endocytosis. However, its potential mechanism has been not well understand. Thus, we aimed to analyze whether albumin endocytosis by mouse glomerular podocytes is dependent on Myo1e expression. Also, we aimed to elucidate whether the underlying mechanism is mediated by Dynamin.

Methods: Firstly, mouse podocyte cells (MPC5) were treated with different concentrations of FITC-bovine serum albumin (BSA). The fluorescence intensity and cell viability were detected by flow cytometry and MTT assays, respectively. Afterwards, the optimal concentration of FITC-BSA was determined. Secondly, MPC5 cells were treated with Myole overexpression or knockdown. Cell morphology was observed under microscope. Immunofluorescence assay was used to determine the expression of F-actin. The protein expression of nephrin and podocin was detected by western blot. Flow cytometry was used to detect MPC5 cell apoptosis with annexin V. Finally, MPC5 cells were treated with Myole overexpression and/or Dynasore (a GTPase inhibitor of Dynamin). The fluorescence intensity was detected using flow cytometry assay.

Results: MPC5 endocytosis BSA was elevated with a concentration-dependent manner. MTT results showed that MPC5 cell viability was inhibited with a concentration-dependent manner. Myo1e overexpression promoted podocyte endocytic FITC-BSA, which was contrary to its knockdown. Under microscope, after inhibition of Myo1e, podocyte foot process fusion was observed. Myo1e overexpression promoted the expression of cytoskeleton F-actin and podocyte-specific molecules (nephrin and podocin) in podocyte endocytic FITC-BSA. Furthermore, we found that Myo1e promoted the apoptosis of podocytes. Dynasore attenuated the increase in endocytosis of FITC-BSA induced by Myo1e overexpression, suggesting that podocytes might mediate albumin endocytosis via Myo1e-Dynamin-Albumin.

Conclusion: Our findings revealed that overexpression of Myo1e promotes albumin endocytosis in mouse glomerular podocyte endocytic albumin mediated by Dynamin.

Keywords: Albumin; Endocytosis; Glomerular albuminuria; MPC5; Podocytes.

Grants and funding

This work was funded by the Zhejiang Provincial Natural Science Foundation of China (LH14H050002). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.