Loss of nephronophthisis-associated nephrocystin-1 impairs DNA damage repair in kidney organoids

  1. E. Sendino Garví
  2. S. Biermans
  3. N.V.A.M. Knoers
  4. A.M. van Eerde
  5. R. Masereeuw
  6. G.G. Slaats
  7. A.M. van Genderen
  8. M.J. Janssen

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Abstract

Nephronophthisis (NPH) is a heterogeneous, autosomal recessive ciliopathy and an important cause of end-stage renal disease (ESRD) in children and young adults. Since its classification as ciliopathy in 2003, NPH disease causal attribution had been focused primarily on ciliary dysfunction. The finding that ciliopathy players are involved in the DNA damage response (DDR) signaling resulted in a paradigm shift in thinking on NPH disease aetiology. Mutations in NPHP1 are the leading cause of NPH, but the underlying mechanisms that lead to the disease phenotype remain poorly understood. Here, nephrocystin-1 depleted kidney organoids were generated and characterized to address this knowledge gap. We used CRISPR/Cas9 to generate NPHP1 control ( NPHP1 WT ) and two mutant ( NPHP1 ko1 and NPHP1 ko2. ) cell lines from healthy human induced pluripotent stem cells (iPSC), differentiated into kidney organoids in an air-liquid interface following an optimized protocol. Upon loss of nephrocystin-1, kidney organoids showed impaired nephron structures and loss of glomerular mesangial and distal tubular cells. Furthermore, NPHP1 depleted organoids exhibited a persistent inability to repair DNA lesions and showed increased senescence and fibrosis characteristics. Dynamic subcellular localization of nephrocystin-1 in NPHP1 WT , particularly its translocation to nuclei 15 min post-UVC light exposure, suggested its direct involvement in the DDR. In conclusion, a novel NPHP1 -depleted kidney organoid model was established, providing a platform to comprehensively study DNA damage, senescence and fibrosis simultaneously upon nephrocystin-1 loss. This advanced model aids in the understanding of the pathophysiology of NPH and paves the way towards identifying novel druggable targets.

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  1. PREreview

    This Zenodo record is a permanently preserved version of a Structured PREreview. You can view the complete PREreview at https://prereview.org/reviews/17304528.

    Does the introduction explain the objective of the research presented in the preprint? Yes
    Are the methods well-suited for this research? Somewhat appropriate
    Are the conclusions supported by the data? Highly supported
    Are the data presentations, including visualizations, well-suited to represent the data? Highly appropriate and clear
    How clearly do the authors discuss, explain, and interpret their findings and potential next steps for the research? Very clearly
    Is the preprint likely to advance academic knowledge? Highly likely
    Would it benefit from language editing? No
    Would you recommend this preprint to others? Yes, it's of high quality
    Is it ready for attention from an editor, publisher or broader audience? Yes, as it is

    Competing interests

    The author declares that they have no competing interests.

    Use of Artificial Intelligence (AI)

    The author declares that they did not use generative AI to come up with new ideas for their review.

  2. Version published to 10.1101/2025.06.09.658557 on bioRxiv