Loss of the collagen IV modifier prolyl 3-hydroxylase 2 causes thin basement membrane nephropathy
Hande Aypek, … , Tobias B. Huber, Florian Grahammer
Hande Aypek, … , Tobias B. Huber, Florian Grahammer
Published May 2, 2022
Citation Information: J Clin Invest. 2022;132(9):e147253. https://doi.org/10.1172/JCI147253.
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Research Article Nephrology

Loss of the collagen IV modifier prolyl 3-hydroxylase 2 causes thin basement membrane nephropathy

Abstract

The glomerular filtration barrier (GFB) produces primary urine and is composed of a fenestrated endothelium, a glomerular basement membrane (GBM), podocytes, and a slit diaphragm. Impairment of the GFB leads to albuminuria and microhematuria. The GBM is generated via secreted proteins from both endothelial cells and podocytes and is supposed to majorly contribute to filtration selectivity. While genetic mutations or variations of GBM components have been recently proposed to be a common cause of glomerular diseases, pathways modifying and stabilizing the GBM remain incompletely understood. Here, we identified prolyl 3-hydroxylase 2 (P3H2) as a regulator of the GBM in an a cohort of patients with albuminuria. P3H2 hydroxylates the 3′ of prolines in collagen IV subchains in the endoplasmic reticulum. Characterization of a P3h2ΔPod mouse line revealed that the absence of P3H2 protein in podocytes induced a thin basement membrane nephropathy (TBMN) phenotype with a thinner GBM than that in WT mice and the development of microhematuria and microalbuminuria over time. Mechanistically, differential quantitative proteomics of the GBM identified a significant decrease in the abundance of collagen IV subchains and their interaction partners in P3h2ΔPod mice. To our knowledge, P3H2 protein is the first identified GBM modifier, and loss or mutation of P3H2 causes TBMN and focal segmental glomerulosclerosis in mice and humans.

Authors

Hande Aypek, Christoph Krisp, Shun Lu, Shuya Liu, Dominik Kylies, Oliver Kretz, Guochao Wu, Manuela Moritz, Kerstin Amann, Kerstin Benz, Ping Tong, Zheng-mao Hu, Sulaiman M. Alsulaiman, Arif O. Khan, Maik Grohmann, Timo Wagner, Janina Müller-Deile, Hartmut Schlüter, Victor G. Puelles, Carsten Bergmann, Tobias B. Huber, Florian Grahammer

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Figure 4

High-resolution imaging to visualize collagen IV α3α4α5 network formation.

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High-resolution imaging to visualize collagen IV α3α4α5 network formatio...
(A) High-resolution imaging of glomeruli from P3h2ΔPod and P3h2fl/fl mice with expansion microscopy. In WT GBM, a linear collagen IV localization was observed for collagen IV and collagen IV α3, α4, and α5 proteins, indicating proper network formation. However, in KO GBM, collagen IV alignment was split, and irregular network formation (white arrows) was observed for collagen IV, collagen IV α3, α4, and α5 stainings, which showed disrupted network formation. Microscopy was performed with a LSM 800 with Airyscan using a ×63 objective, a digital zoom of ×8, and ×4 linear expansion of tissues. (B) Quantification of irregular network formation of collagen IV. Randomly chosen glomeruli from WT and KO mice were evaluated for linear and irregular collagen IV network formation. The graph shows that collagen IV network formation in KO GBM was significantly disrupted when compared with that in WT GBM, indicating that collagen IV α3, α4, and α5 proteins were unable to form a proper network in the absence of P3H2. n = 3. Graphs show the mean ± SD. ****P < 0.0001, by unpaired, 2-tailed Student’s t test.