Profilin1 is required for prevention of mitotic catastrophe in murine and human glomerular diseases
Xuefei Tian, … , Rongguo Fu, Shuta Ishibe
Xuefei Tian, … , Rongguo Fu, Shuta Ishibe
Published October 17, 2023
Citation Information: J Clin Invest. 2023;133(24):e171237. https://doi.org/10.1172/JCI171237.
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Research Article Cell biology Nephrology

Profilin1 is required for prevention of mitotic catastrophe in murine and human glomerular diseases

Abstract

The progression of proteinuric kidney diseases is associated with podocyte loss, but the mechanisms underlying this process remain unclear. Podocytes reenter the cell cycle to repair double-stranded DNA breaks. However, unsuccessful repair can result in podocytes crossing the G1/S checkpoint and undergoing abortive cytokinesis. In this study, we identified Pfn1 as indispensable in maintaining glomerular integrity — its tissue-specific loss in mouse podocytes resulted in severe proteinuria and kidney failure. Our results suggest that this phenotype is due to podocyte mitotic catastrophe (MC), characterized histologically and ultrastructurally by abundant multinucleated cells, irregular nuclei, and mitotic spindles. Podocyte cell cycle reentry was identified using FUCCI2aR mice, and we observed altered expression of cell-cycle associated proteins, such as p21, p53, cyclin B1, and cyclin D1. Podocyte-specific translating ribosome affinity purification and RNA-Seq revealed the downregulation of ribosomal RNA-processing 8 (Rrp8). Overexpression of Rrp8 in Pfn1-KO podocytes partially rescued the phenotype in vitro. Clinical and ultrastructural tomographic analysis of patients with diverse proteinuric kidney diseases further validated the presence of MC podocytes and reduction in podocyte PFN1 expression within kidney tissues. These results suggest that profilin1 is essential in regulating the podocyte cell cycle and its disruption leads to MC and subsequent podocyte loss.

Authors

Xuefei Tian, Christopher E. Pedigo, Ke Li, Xiaotao Ma, Patricia Bunda, John Pell, Angela Lek, Jianlei Gu, Yan Zhang, Paulina X. Medina Rangel, Wei Li, Eike Schwartze, Soichiro Nagata, Gabriel Lerner, Sudhir Perincheri, Anupama Priyadarshini, Hongyu Zhao, Monkol Lek, Madhav C. Menon, Rongguo Fu, Shuta Ishibe

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Figure 3

Loss of podocyte Pfn1 results in morphologic MC appearance, chromosomal instability, and dsDNA damage.

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Loss of podocyte Pfn1 results in morphologic MC appearance, chromosomal ...
(A) Representative images of transmission electron micrography demonstrate abnormal MC podocytes in Pfn1-KO mice (arrow) compared with control glomeruli at 4 weeks of age. Scale bar: 1 μm. (B) Immunofluorescence images of primary podocytes isolated from control and Pfn1-KO mice at P7 stained with WT1 (red, podocyte marker) and Hoechst (blue, DNA marker) showing abnormal MC podocytes in Pfn1-KO mice (arrow) compared with control podocytes. Scale bar: 20 μm. (C) Immunofluorescence images of primary culture podocytes isolated from control and Pfn1-KO mice stained with Hoechst and anti-tubulin antibody showing the chromosome bridge (arrow) in a MC Pfn1-KO podocyte. Scale bar: 20 μm. (D) Quantification of the percentage of MC podocytes per field of view in B. Total of 100 fields of view in 5 independent experiments. (E) Quantification of the percentage of chromosome bridge per field of view in C. n = 5 independent experiments. *P < 0.05 vs. control. (F) Immunofluorescence images of primary culture podocytes stained with γ-H2AX (green, double-strand breaks [DSBs] marker) and WT1 (red) showing abnormal MC podocytes in Pfn1-KO mice, as indicated by the arrows. Scale bar: 20 μm. (G) Quantification of γH2AX foci per podocyte nucleus (left) and per podocyte nuclear area (right) in F. Total of 400 cells in 5 independent experiments. *P < 0.05 vs. control. Statistics were analyzed via a 2-tailed t test.