Abstract: FR-PO236

APOL1 Facilitates Transition of Parietal Epithelial Cells (PECs) via Down-Regulation of miR193a

Session Information

Category: Cell Biology

  • 202 Apoptosis, Proliferation, Autophagy, Cell Senescence, Cell Transformation

Authors

  • Kumar, Vinod, Fienstine Institute for Medical Research, New York, New York, United States
  • Lan, Xiqian, Feinstein Institute for Medical Research, Great Neck, New York, United States
  • Marashi Shoshtari, Seyedeh Shadafarin, The Feinstein Institute for Medical Research, Manhasset, New York, United States
  • Meyer-Schwesinger, Catherine, University of Hamburg, Hamburg, Germany
  • Malhotra, Ashwani, Feinstein Inst.Med research and NSLIJ, Manhasset, New York, United States
  • Skorecki, Karl, Rambam Health Care Campus, Haifa, Israel
  • Singhal, Pravin C., Fienstine Institute for Medical Research, New York, New York, United States
Background

Progenitor cells play an important role to maintain podocyte (PD) homeostasis during podocyte cytotoxic environment. A subset of pareital epithelial cells (PEC) has been reported to act as progenitor cells for the maintenance of podocyte homeostasis. Down- regulation of microRNA (miR) 193a has been demonstrated to facilitate transition (acquirement of podocyte differentiatin markers) of cultured PECs. APOL1 is expressed intracellularly in podocytes, however, PECs do not express APOL1. APOL1 risk alleles (G1 and G2) have been reported to PD cytotoxic, however, the role of PD expression of APOL1G0 is far from clear. We hypothesize that APOL1 expression emerges in PECs to facilitate PECs transition through down-regulation of miR193a.

Methods

Human immortalized cultured PECs proliferate at 33°C and differentiate at 37°C. PECs were differentiated for variable time periods (4, 8, 12 days; n=4). To examine a causal relationship, PECs were transfected with either control or APOL1 siRNA and followed by differentiation at 37C (n=4). To study the feedback relationship between APOL1 and miR193a, PECs and Hep G2 cells (+ve control) were treated with different concentrations of an inhibitor of miR193a (25, 50, and 100 nM) followed by differentiation at 37C. To confirm the relationship between APOL1 and miR193a, HEK (human embryonic kidney) cells /mouse (M) PDs (-ve control for APOL1) were transfected with either control or APOL1 plasmids. Protein blots were probed for APOL1, WT1, podocalyxin, podocin, and reprobed for actin. RNAs were assayed for miR193a levels. To confirm binding of miR193a to the APOL1 gene, RIP-ChIP assay was carried out.

Results

PECs started displaying APOL1 on day 4 during their transition at 37°C. APOL1 expression was associated with down- regulation of miR193a and enhanced expression of WT1, podocalyxin, and podocin. Knockdown of APOL1 up regulated (2 fold) miR193a levels in PECs and Hep G2, whereas, expression of APOL1 in HEKs and mouse podocytes down-regulated (2.5 fold) miR193a expression. Since miR193a also inversely up-regulated APOL1 expression, this suggests a feedback loop relationship in PECs. RIP-ChIP assay confirmed binding of miR193a to APOL1 gene promoter.

Conclusion

APOL1 facilitates PEC transition through down-regulation of miR193a.

Funding

  • NIDDK Support