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Abstract: FR-PO932

Studying the Role of MiRNA in Podocyte Maintenance, Inspired by a Nephrotic Syndrome-Related Point Mutation in XPO5

Session Information

Category: Glomerular Diseases

  • 1204 Podocyte Biology

Authors

  • Navon, Royi, Hadassah - Hebrew University Medical Center, Jerusalem, Israel
  • Ben-Dov, Iddo Z., Hadassah - Hebrew University Medical Center, Jerusalem, Israel
  • Gur-Wahnon, Devorah, Hadassah - Hebrew University Medical Center, Jerusalem, Israel

Group or Team Name

  • Laboratory of Medical Transcriptomics
Background

MicroRNA (miRNA) are small noncoding RNA molecules that regulate gene expression, and are crucial for the development and homeostasis of podocytes. Biogenesis of miRNA requires a multistage process of which Exportin5 (XPO5) plays a rate limiting step by exporting miRNA precursors from the nucleus to the cytoplasm. Recently, steroid-resistant nephrotic syndrome (SRNS) in a child was attributed to a point mutation in XPO5 (Braun DA 2016). However, whether or not the mutation affects the miRNA related function of Exportin-5 is not known. We hypothesized that the V552I mutation impedes miRNA maturation and that the association with SRNS may shed light on the roles of specific miRNA in podocytes.

Methods

We have successfully generated the p.V552I Exportin-5 (XPO5V552I) homozygous mutation in HEK293 cells via CRISPR-Cas9. In addition we have generated human podocyte clones with a heterozygous mutation.

Results

Small RNA sequencing of our HEK293 cells shows a significant decrease in global miRNA content in XPO5V552I compared to parental cells. Principal component analysis (Figure) also revealed population separation between wildtype HEK293 and XPO5V552I cells, indicating miRNA expression segregated by genotype. Moreover, Small RNA sequencing of the podocyte clones revealed distinct changes in miRNA profiles.

Conclusion

These results strongly suggest the involvement of the XPO5V552I mutation in the dysregulation seen in the miRNA. Further investigation will lead us to the discovery of the specific miRNA involved in maturation and maintenance of podocytes.

Figure: Principal component analysis plot by microRNA profiles showing samples arranging by cell type and genotype. hek, HEK293 cells; pods, podocytes; wt, control cells; mut, cells with mutated XPO5.

Funding

  • Government Support - Non-U.S.