Molecular Characterization of Podocytes in Podocyte-Specific Nup160 Knockout Mice

Background: Mutations in NUP160 cause steroid-resistant nephrotic syndrome (SRNS). Our previous study found that podocyte-specific Nup160 knockout mice developed nephrotic syndrome, but pathogenic mechanism remains unknown. Here, we characterized molecules of podocytes in podocyte-specific Nup160 knockout mice. Methods: Podocyte-specific Nup160 knockout mouse model was generated. This model incorporated double-fluorescent Cre reporter genes, allowing for the visualization of green fluorescence in podocytes and red fluorescence in non-podocytes. The mice were divided into three groups: Nup160podo-/-, Nup160podo+/- and Nup160podo+/+. Single-cell proteomics techniques were employed to detect the expression of podocyte molecules. The thresholds of differentially expressed genes were set at an adjusted P < 0.05 and |log2FC| ≥ 0.15. Subsequently, enrichment analysis of Gene Ontology and the Kyoto Encyclopedia of Genes and Genomes was conducted on these differentially expressed genes to characterize their molecular signature. Results: A total of 1414 proteins were identified in the podocytes of Nup160podo+/+ mice, and 1416 proteins in the podocytes of Nup160pod+/- mice, and 1413 proteins in the podocytes of Nup160podo-/- mice. Thirteen genes were found to be upregulated and 49 downregulated in the Nup160podo-/- mice compared with the Nup160podo+/+ mice. The downregulated differentially expressed genes were significantly associated with pathways that regulate synaptic structure modification, synapse organization, nucleocytoplasmic transport, actin binding, and SMAD binding. The differentially expressed genes were closely linked to pathways related to actin filaments, actin binding, organization, RNA splicing, and spliceosome function. Additionally, 13 genes were upregulated and 19 genes downregulated in the Nup160podo-/- mice compared with the Nup160podo+/- mice. The downregulated differentially expressed genes were closely associated with cell adhesion molecule binding, integrin binding, and GTPase activating protein binding. The upregulated differentially expressed genes were related to regulating Ras protein signal transduction, small GTPase-mediated signal transduction, and neurotransmitter transport pathways. Conclusion: Single-cell proteomics reveals that loss of Nup160 impairs actin cytoskeleton organization, cell adhesion, and nucleocytoplasmic transport of the podocytes. Funding: Government Support – Non-U.S.